Microbial

J Biol Chem, 2001 Jun 15, 276(24), 21311 - 6 Epub 2001 Apr 03.
Evidence for direct interaction between enzyme I(Ntr) and aspartokinase to regulate bacterial oligopeptide transport; King ND et al.; Bradyrhizobium japonicum transports oligopeptides and the heme precursor delta-aminolevulinic acid (ALA) by a common mechanism . Two Tn5-induced mutants disrupted in the lysC and ptsP genes were identified based on the inability to use prolyl-glycyl-glycine as a proline source and were defective in {(14)C}ALA uptake activity . lysC and ptsP were shown to be proximal genes in the B . japonicum genome . However, RNase protection and in trans complementation analysis showed that lysC and ptsP are transcribed separately, and that both genes are involved in oligopeptide transport . Aspartokinase, encoded by lysC, catalyzes the phosphorylation of aspartate for synthesis of three amino acids, but the lysC strain is not an amino acid auxotroph . The ptsP gene encodes Enzyme I(Ntr) (EI(Ntr)), a paralogue of Enzyme I of the phosphoenolpyruvate:sugar phosphotransferase (PTS) system . In vitro pull-down experiments indicated that purified recombinant aspartokinase and EI(Ntr) interact directly with each other . Expression of ptsP in trans from a multicopy plasmid complemented the lysC mutant, suggesting that aspartokinase normally affects Enzyme I(Ntr) in a manner that can be compensated for by increasing the copy number of the ptsP gene . ATP was not a phosphoryl donor to purified EI(Ntr), but it was phosphorylated by ATP in the presence of cell extracts . This phosphorylation was inhibited in the presence of aspartokinase . The findings demonstrate a role for a PTS protein in the transport of a non-sugar solute and suggest an unusual regulatory function for aspartokinase in regulating the phosphorylation state of EI(Ntr).

Placenta, 2001 Apr, 22(4), 360 - 71
Study on risk factors for transplacental viral infections; effect of bacterial factors and double viral infections on virus replication in placenta and amniotic membranes; Jatczak B et al.; Among risk factors for vertical transmission of HIV there are listed concomitant viral and bacterial infections . Therefore the influence on the viruses replication in human placenta and amniotic membrane cultures of double viral infection with two unrelated viruses - encephalomyocarditis (EMCV) and vesicular stomatitis virus (VSV) - was studied and compared with the replication of the viruses in single virus infection (EMCV or VSV) in the same organ cultures . Additionally effect of bacterial factors - lipopolysaccharide (LPS) Escherichia coli and sonicated Treponema pallidum antigens (Tpa) - on VSV replication in the same culture system was studied and compared with VSV replication in untreated explants . Two effects were observed in double-virus infected cultures and also in bacterial factors treated cultures: inhibition and stimulation of virus replication . The kind of effect in the both cases was dependent on the presence or absence of innate antiviral immunity . In virus-sensitive organs double infected or treated with LPS or Tpa, inhibition of virus titer (2-5 log TCID(50)/ml) was observed . In the organs expressing the innate immunity, stimulation (1-4 log TCID(50)/ml) of virus replication was noticed . Contribution of endogenous TNFalpha in both reactions (stimulation and inhibition) was confirmed using antibodies against the TNF .

Int J Infect Dis, 2001, 5(1), 9 - 18
Effect of surfactant and specific antibody on bacterial proliferation and lung function in experimental pneumococcal pneumonia; Gan X et al.; OBJECTIVE: To investigate the effect of surfactant and specific antibody on bacterial proliferation in experimental pneumococcal pneumonia . METHODS: Near-term newborn rabbits received a standard dose (10(7)) of type 3 pneumococci via the airways . Control animals were sacrificed 1 minute later . Other animals were ventilated for 5 hours and treated via the tracheal cannula with surfactant (Curosurf 200 mg/kg), a mixture of surfactant and a polyclonal antipneumococcal antibody, the antibody without surfactant, or saline . RESULTS: There was a significant bacterial proliferation in lung tissue in all animals ventilated for 5 hours . Bacterial growth, expressed as log10 colony forming units (CFU) per gram of lung tissue was less prominent in animals treated with a mixture of surfactant and specific antibody than in animals treated with antibody alone (median, 7.51, range, 6.80--7.70 vs . median, 7.92, range, 7.07--8.50; P < 0.05) . Dynamic lung-thorax compliance was improved with surfactant or surfactant plus antibody in comparison with saline or antibody alone . CONCLUSIONS: The data suggest that the suppressive effect of the antibody on bacterial proliferation becomes evident only when surfactant is administered together with the antibody.

Eur J Neurosci, 2001 Mar, 13(6), 1155 - 65
Central monoamine and plasma corticosterone changes induced by a bacterial endotoxin: sensitization and cross-sensitization effects; Hayley S et al.; Low doses of lipopolysaccharide, tumour necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), or exposure to a stressor (restraint) increased plasma corticosterone levels . In animals pretreated with lipopolysaccharide, a marked sensitization of the corticosterone response was evident upon subsequent exposure to lipopolysaccharide, TNF-alpha, or restraint, 1 day later . As well, the sickness-inducing effects of lipopolysaccharide, TNF-alpha and IL-1 beta were markedly increased in mice pretreated with lipopolysaccharide . The sensitization effects were marked when the second treatment was administered 1 day after lipopolysaccharide administration, but not when a 28-day interval elapsed . In a second experiment, TNF-alpha influenced monoamine functioning in the paraventricular nucleus of the hypothalamus and within extrahypothalamic regions, including the central amygdala, locus coeruleus, prefrontal cortex . Moreover, serotonin activity within the central amygdala, as well as dopamine activity within the prefrontal cortex, were subject to a sensitization effect in animals pretreated with lipopolysaccharide 1 day earlier . Macrophage depletion by a suspension of clodronate liposomes attenuated the plasma corticosterone changes induced by TNF-alpha, but did not affect the sensitization . In contrast, the acute effects of TNF-alpha on central neurotransmitters were unaffected by the liposome suspension, but this treatment prevented the sensitization . These data may be relevant to clinical situations in which individuals exposed to bacterial infections may be rendered more susceptible to the behavioural and neurochemical effects of subsequently encountered stressors and immunological challenges.

Eur J Neurosci, 2001 Mar, 13(6), 1101 - 6
Bacterial endotoxin sensitizes the immature brain to hypoxic--ischaemic injury; Eklind S et al.; Epidemiological studies show a markedly increased risk of cerebral palsy following the combined exposure of infection and birth asphyxia . However, the underlying mechanisms of this increased vulnerability remain unclear . We have examined the effects of a low dose of bacterial endotoxin on hypoxic--ischaemic injury in the immature brain of rats . Bacterial endotoxin (lipopolysaccharide 0.3 mg/kg) was administered to 7-day-old rats 4 h prior to unilateral hypoxia--ischaemia and the neurological outcome was determined 3 days later . Rectal temperature and cerebral blood flow was measured during the study and the expression of CD14 and toll-like receptor-4 mRNA in the brain was examined . We found that a low dose of endotoxin dramatically sensitizes the immature brain to injury and induces cerebral infarction in response to short periods of hypoxia--ischaemia that by themselves caused no or little injury . This effect could not be explained by a reduction in cerebral blood flow or hyperthermia . In association with the sensitization of injury we found an altered expression of CD14 mRNA and toll-like receptor-4 mRNA in the brain . These results suggest that the innate immune system may be involved in the vulnerability of the immature brain following the combination of infection and hypoxia--ischaemia.

JPEN J Parenter Enteral Nutr, 2001 Mar-Apr, 25(2), 60 - 4
Liquid enteral diets induce bacterial translocation by increasing cecal flora without changing intestinal motility; Haskel Y et al.; The aim of this study was to determine the contribution of intestinal motility and cecal bacterial overgrowth to liquid diet-induced bacterial translocation (BT) . Three different commercially available liquid diets were offered to mice for 1 week . BT to the mesenteric lymph nodes (MLN), spleen, and liver were examined as well as cecal bacterial counts and populations, small bowel length and weight, and histopathologic changes in the ileal and jejunal mucosa . In addition, the effect of the various diets on intestinal motility was measured by the transit index of a charcoal mixture introduced into the stomach . The incidence of BT to the mesenteric lymph nodes was significantly and similarly increased (p < .05) in mice fed Vivonex (30%), Ensure (30%), and Osmolite (33%) compared with chow-fed controls (0%) . Compared with chow-fed controls, all three liquid diets were associated with the development of cecal bacterial overgrowth (p < .01) . There were no significant changes in the transit index for the three liquid diet groups compared with the chow-fed controls . BT to the MLN was induced by all three liquid diets tested, casting some doubts as to their role in preventing BT in clinical use . BT was associated with a statistically significant increase in cecal bacterial count but was not associated with gut motility changes in this model . In fact, no significant changes in intestinal motility were noted in all groups tested.

Appl Environ Microbiol, 2001 Apr, 67(4), 1830 - 8
Rapid method of determining factors limiting bacterial growth in soil; Alden L et al.; A technique to determine which nutrients limit bacterial growth in soil was developed . The method was based on measuring the thymidine incorporation rate of bacteria after the addition of C, N, and P in different combinations to soil samples . First, the thymidine incorporation method was tested in two different soils: an agricultural soil and a forest humus soil . Carbon (as glucose) was found to be the limiting substance for bacterial growth in both of these soils . The effect of adding different amounts of nutrients was studied, and tests were performed to determine whether the additions affected the soil pH and subsequent bacterial activity . The incubation time required to detect bacterial growth after adding substrate to the soil was also evaluated . Second, the method was used in experiments in which three different size fractions of straw (1 to 2, 0.25 to 1, and <0.25 mm) were mixed into the agricultural soil in order to induce N limitation for bacterial growth . When the straw fraction was small enough (<0.25 mm), N became the limiting nutrient for bacterial growth after about 3 weeks . After the addition of the larger straw fractions (1 to 2 and 0.25 to 1 mm), the soil bacteria were C limited throughout the incubation period (10 weeks), although an increase in the thymidine incorporation rate after the addition of C and N together compared with adding them separately was seen in the sample containing the size fraction from 0.25 to 1 mm . Third, soils from high-pH, limestone-rich areas were examined . P limitation was observed in one of these soils, while tendencies toward P limitation were seen in some of the other soils.

Appl Environ Microbiol, 2001 Apr, 67(4), 1775 - 82
Does the high nucleic acid content of individual bacterial cells allow us to discriminate between active cells and inactive cells in aquatic systems?
Lebaron P, Servais P, Agogue H, Courties C, Joux F.
The nucleic acid contents of individual bacterial cells as determined with three different nucleic acid-specific fluorescent dyes (SYBR I, SYBR II, and SYTO 13) and flow cytometry were compared for different seawater samples . Similar fluorescence patterns were observed, and bacteria with high apparent nucleic acid contents (HNA) could be discriminated from bacteria with low nucleic acid contents (LNA) . The best discrimination between HNA and LNA cells was found when cells were stained with SYBR II . Bacteria in different water samples collected from seven freshwater, brackish water, and seawater ecosystems were prelabeled with tritiated leucine and then stained with SYBR II . After labeling and staining, HNA, LNA, and total cells were sorted by flow cytometry, and the specific activity of each cellular category was determined from leucine incorporation rates . The HNA cells were responsible for most of the total bacterial production, and the specific activities of cells in the HNA population varied between samples by a factor of seven . We suggest that nucleic acid content alone can be a better indicator of the fraction of growing cells than total counts and that this approach should be combined with other fluorescent physiological probes to improve detection of the most active cells in aquatic systems.

Appl Environ Microbiol, 2001 Apr, 67(4), 1565 - 74
Bacterial diversity and community structure in an aerated lagoon revealed by ribosomal intergenic spacer analyses and 16S ribosomal DNA sequencing; Yu Z et al.; We investigated the bacterial community structure in an aerated plug-flow lagoon treating pulp and paper mill effluent . For this investigation, we developed a composite method based on analyses of PCR amplicons containing the ribosomal intergenic spacer (RIS) and its flanking partial 16S rRNA gene . Community percent similarity was determined on the basis of RIS length polymorphism . A community succession was evident in the lagoon, indicated by a progressive community transition through seven sample locations . The most abrupt changes in community structure were associated with a temperature change from 39 to 35 degrees C and with increases in dissolved oxygen . The temporal differences in community structure, based on summer and winter samplings, were greater than the spatial differences during either season . Clone libraries of rDNA-RIS amplicons were constructed from each of three summer samples . Among 90 clones analyzed (30 clones from each sample), 56 phylotypes were distinguished by restriction fragment length polymorphism . Indices of phylotype richness, evenness, and diversity all increased in clone libraries from the beginning to the end of the lagoon . A representative clone of each phylotype was phylogenetically analyzed on the basis of its partial 16S rRNA gene sequence (ca . 450 bp) . Phylogenetic analysis confirmed the increase in diversity and further indicated increasing richness of bacterial divisions . Pioneers in the community spatial succession appeared to include thermotolerant, microaerophilic methanol-oxidizing bacteria related to the genus Methylobacillus, as well as thermotolerant, microaerophilic nitrogen-fixing bacteria related to the genus Azospirillum.

Eur Cytokine Netw, 2001 Mar, 12(1), 187 - 93
Rat interleukin-10: production and characterisation of biologically active protein in a recombinant bacterial expression system; Ball C et al.; Interleukin-10 is an anti-inflammatory Th1 immunosuppressive cytokine, the active form of which is a non-covalent homodimer, and which exhibits species-specificity both with respect to structure and biological activity . The rat homologue of IL-10 shares 73% identity with human IL-10 at the amino-acid sequence level, and has, in addition to the two disulphide bonds present in human IL-10, a fifth, unpaired cysteine (cys-149) . Preparation of rat IL-10 by bacterial expression followed by solubilisation and refolding in a glutathione redox system, results in a molecule in which cys-149 is almost entirely oxidised, existing either as disulphide dimer or as a mixed disulphide with glutathione, and which has less than 1% of the activity of the native (cys-149-SH) form of the molecule . Site directed mutagenesis of rat IL-10 to replace cys-149 with tyrosine produces a molecule which readily adopts the active conformation upon solubilisation and refolding, and which is recoverable in good yield from bacterial expression systems . Comparison of the biological activities of rat IL-10tyr149 and commercial rat IL-10 preparations confirms that the activity of native-sequence rat IL-10 is either reduced or absent . It is proposed therefore that the biosynthetic analogue rat IL-10tyr149 is a more useful molecule to investigate the biological actions of IL-10 in the rat.

Ann Biol Clin (Paris), 2001 Mar-Apr, 59(2), 177 - 82
{Molecular tools in the epidemiology of tick-borne bacterial diseases}; Parola P et al.; Molecular tools have been used to detect rickettsiae in ticks . In Ixodes ricinus ticks collected in France, we detected for the first time there an emerging pathogen, Rickettsia helvetica, and an Ehrlichia sp, closely related to the agent of human granulocytic ehrlichiosis . In Guadeloupe (French West Indies), we described the occurrence of African tick-bite fever due to Rickettsia africae, which had been previously reported in sub-Saharan Africa only . In Africa, we completed our knowledge about the distribution of R . africae (Mali, Niger, Sudan, Burundi), and detected for the first time Rickettsia mongolotimonae, an emerging pathogen . Anaplasma marginale the agent of bovine anaplasmosis was detected in Mali . Rickettsiae of unknown pathogenicity were detected in Mali and Niger.

J Hepatol, 2001 Feb, 34(2), 215 - 21
Ascitic fluid carcinoembryonic antigen and alkaline phosphatase levels for the differentiation of primary from secondary bacterial peritonitis with intestinal perforation; Wu SS et al.; BACKGROUND/AIMS: In cirrhotic patients, spontaneous bacterial peritonitis (SBP) may be difficult to distinguish from secondary peritonitis with occult intestinal perforation; Runyon's criteria (based on ascitic fluid glucose, protein and lactate dehydrogenase levels) are sensitive but not specific . Ascitic fluid carcinoembryonic antigen (CEA) and alkaline phosphatase (AP) are potential markers for secondary peritonitis . METHODS: Ascitic fluid CEA and AP levels were prospectively compared among three subject groups--cirrhotic patients with sterile ascites, cirrhotic patients with SBP, and patients (cirrhotic and non-cirrhotic) with perforation-related secondary peritonitis . RESULTS: The secondary peritonitis group (n = 38 including 11 cirrhotic patients) had significantly higher mean CEA and AP levels than the SBP (n = 34) and sterile ascites patients (n = 63) . Of secondary peritonitis patients, 92% fulfilled predetermined criteria (either CEA >5 ng/ml or AP >240 units/l) versus only 12% of SBP patients; sensitivity was 92% and specificity 88% for differentiating secondary peritonitis from SBP . Runyon's criteria had a sensitivity of 97% and specificity of 56% . Stratification of secondary peritonitis patients by the presence or absence of cirrhosis did not alter our results . CONCLUSIONS: Ascitic fluid CEA or AP elevations appear to be sensitive and specific markers for perforation-related secondary peritonitis in cirrhotic as well as non-cirrhotic patients.

Res Microbiol, 2001 Jan-Feb, 152(1), 11 - 6
Second-order selection in bacterial evolution: selection acting on mutation and recombination rates in the course of adaptation; Tenaillon O et al.; The increase in genetic variability of a population can be selected during adaptation, as demonstrated by the selection of mutator alleles . The dynamics of this phenomenon, named second-order selection, can result in an improved adaptability of bacteria through regulation of all facets of mutation and recombination processes.

Yi Chuan Xue Bao, 2001, 28(3), 236 - 43
{Construction of a bacterial artificial chromosome (BAC) contig encompassing the bacterial blight resistance gene Xa4 locus in rice}; Jiang GH et al.; The gene Xa4 confers dominantly resistance to rice bacterial blight, which has been finely mapped between RFLP markers G181 and L1044, and co-segregated with the resistance gene homologues sequence marker RS13 . The three markers were used to screen a rice Bacterial Artificial Chromosome (BAC) library constructed from IRBB56, a Xa4-harborring indica variety, resulting in the detection of totally 128 positive clones . Of the 18 positive clones picked out by RS13, 4 and 6 clones were simultaneously detected by G181 and L1044, respectively . Based on their HindIII restriction patterns, 12 clones were selected out to construct a contig that spanned about 420 kb covering the Xa4 locus, which is a solid base for the isolation of Xa4 gene.

Compr Ther, 2001 Spring, 27(1), 72 - 7
Utility of prognostic stratification in adults with community-acquired bacterial meningitis; Aronin SI et al.; Prognostic stratification uses baseline clinical features to subdivide patients into subgroups with different risks for a particular outcome . We review the importance of prognostic stratification in internal medicine, in infectious diseases, and in adults with community-acquired bacterial meningitis.

J Biol Chem, 2001 Apr 27, 276(17), 14393 - 9 Epub 2001 Jan 22.
Overproduction of bacterial protein disulfide isomerase (DsbC) and its modulator (DsbD) markedly enhances periplasmic production of human nerve growth factor in Escherichia coli; Kurokawa Y et al.; Production of eukaryotic proteins with multiple disulfide bonds in the Escherichia coli periplasm often encounters difficulty in obtaining soluble products with native structure . Human nerve growth factor beta (NGF) contains three disulfide bonds between nonconsecutive cysteine residues and forms insoluble aggregates when expressed in E . coli . We now report that overexpression of Dsb proteins known to catalyze formation and isomerization of disulfide bonds can substantially enhance periplasmic production of NGF . A set of pACYC184-based plasmids that permit dsb expression under the araB promoter were introduced into cells carrying a compatible plasmid that expresses NGF . The efficiency of periplasmic production of NGF fused to the OmpT signal peptide was strikingly improved by coexpression of DsbCD or DsbABCD proteins (up to 80% of total NGF produced) . Coexpression of DsbAB was hardly effective, whereas that of DsbAC increased the total yield but not the periplasmic expression . These results suggest synergistic roles of DsbC and DsbD in disulfide isomerization that appear to become limiting upon NGF production . Furthermore, recombinant NGF produced with excess DsbCD (or DsbABCD) was biologically active judged by the neurite outgrowth assay using rat PC12 cells.

Virology, 2001 Mar 15, 281(2), 281 - 93
Chemical modification patterns of active and inactive as well as procapsid-bound and unbound DNA-packaging RNAof bacterial virus Phi29; Zhang C et al.; During replication, the lengthy genome of dsDNA viruses is translocated with remarkable velocity into the limited space within the preformed procapsid . We previously found that a viral-encoded RNA (pRNA) played a key role in bacterial virus phi29 DNA translocation . Design of mutant pRNA sets containing two and three inactive mutant pRNAs, respectively, led to the conclusion that the stoichiometry of pRNA in DNA packaging is the common multiple of 2 and 3 . Together with studies using binomial distribution of mutant and wild-type pRNA, it has been confirmed that six pRNAs of phi29 form a hexagonal complex to drive the DNA translocating machine . These findings have brought about commonality between viral DNA packaging and other universal DNA/RNA-riding processes including DNA replication and RNA transcription . Chemical modification was used to compare the structures of active and inactive as well as free and procapsid-bound pRNA . Our results explain why certain pRNA mutants are inactive in DNA packaging while remaining competent in procapsid binding, since the mutations were located in a domain involved in DNA translocation that is dispensable for procapsid binding . A mutant pRNA that had reduced procapsid binding was revealed to have a structural alteration within the procapsid-binding region that may account for the binding deficiency . Chemical probing of procapsid-bound pRNA revealed a large area of protection, while a 3-base bulge, C(18)C(19)A(20), was accessible to chemicals . A pRNA with a deletion of this 3-base bulge was fully competent to form dimers, bind procapsids, and inhibit phi29 virion assembly in vitro; however, its activity in DNA packaging and virion assembly was completely lost . The results suggest that this bulge is not involved in procapsid binding but may interact with other DNA-packaging components . A computer model showing the location of the CCA bulge was presented .

J Ethnopharmacol, 2001 Mar 3, 74(3), 231 - 7
Cyclooxygenase inhibiting and anti-bacterial activities of South African Erythrina species; Pillay CC et al.; Aqueous, ethanolic and ethyl acetate extracts of the bark and leaves of five South African Erythrina species Erythrina caffra, Erythrina humeana, Erythrina latissima, Erythrina lysistemon and Erythrina zeyheri were screened for prostaglandin synthesis-inhibitory and anti-bacterial activity . The bark generally displayed higher activity than the leaves in both bioassays . The highest cyclooxygenase inhibiting activity and anti-bacterial activity was recorded for the ethanol and ethyl acetate bark extracts of E . caffra, E . latissima and E . lysistemon . An anti-bacterial compound, 4',5,7-trihydroxy-6-prenylisoflavone, was isolated by bioassay-guided fractionation from bark of E . lysistemon.

Proc Natl Acad Sci U S A, 2001 Mar 27, 98(7), 4243 - 8
Vipp1 deletion mutant of Synechocystis: a connection between bacterial phage shock and thylakoid biogenesis?
Westphal S, Heins L, Soll J, Vothknecht UC.
Plant chloroplasts originated from an endosymbiotic event by which an ancestor of contemporary cyanobacteria was engulfed by an early eukaryotic cell and then transformed into an organelle . Oxygenic photosynthesis is the specific feature of cyanobacteria and chloroplasts, and the photosynthetic machinery resides in an internal membrane system, the thylakoids . The origin and genesis of thylakoid membranes, which are essential for oxygenic photosynthesis, are still an enigma . Vipp1 (vesicle-inducing protein in plastids 1) is a protein located in both the inner envelope and the thylakoids of Pisum sativum and Arabidopsis thaliana . In Arabidopsis disruption of the VIPP1 gene severely affects the plant's ability to form properly structured thylakoids and as a consequence to carry out photosynthesis . In contrast, Vipp1 in Synechocystis appears to be located exclusively in the plasma membrane . Yet, as in higher plants, disruption of the VIPP1 gene locus leads to the complete loss of thylakoid formation . So far VIPP1 genes are found only in organisms carrying out oxygenic photosynthesis . They share sequence homology with a subunit encoded by the bacterial phage shock operon (PspA) but differ from PspA by a C-terminal extension of about 30 amino acids . In two cyanobacteria, Synechocystis and Anabaena, both a VIPP1 and a pspA gene are present, and phylogenetic analysis indicates that VIPP1 originated from a gene duplication of the latter and thereafter acquired its new function . It also appears that the C-terminal extension that discriminates VIPP1 proteins from PspA is important for its function in thylakoid formation.

Folia Microbiol (Praha), 2000, 45(3), 197 - 203
From no-confidence to nitric oxide acknowledgement: a story of bacterial nitric-oxide reductase; Koutny M; The review briefly summarizes current knowledge of the bacterial nitric-oxide reductase (NOR) . This membrane enzyme consists of two subunits, the smaller one contains haem C and the larger one two haems B and nonhaem iron . The protein sequence and structure of metal centres demonstrate the relationship of NOR to the family of terminal oxidases . The binuclear Fe-Fe reaction centre, consisting of antiferromagnetically coupled haem B and nonhaem iron, is analogous to Fe-Cu centre of terminal oxidases . The data on the structure and function of NOR and terminal oxidases suggest that all these enzymes are closely evolutionally related . The catalytic properties are determined most of all by the relatively high toxicity of nitric oxide as a substrate and the resulting strong need to maintain its concentration at nanomolar levels . A kinetic model of the action of the enzyme comprises substrate inhibition . NOR does not conserve the free energy of nitric oxide reduction because it does not work as a proton pump and, moreover, the protons coming into the reaction are taken from periplasm, i.e . they do not cross the membrane.

Fish Shellfish Immunol, 2001 Jan, 11(1), 23 - 37
Enhanced lysozyme production in Atlantic salmon (Salmo salar L.) macrophages treated with yeast beta-glucan and bacterial lipopolysaccharide; Paulsen SM et al.; Atlantic salmon head kidney macrophages grown in the presence of particulate yeast beta-glucan and bacterial lipopolysaccharide (LPS) showed increased production of lysozyme in the culture supernatants compared to non-treated controls . The increased lysozyme production started at day 3 and was five- to six-fold higher compared to controls at day 6 in culture . Beta-glucan showed an approximate linear dose-response curve between 1 and 250 microg x ml(-1) whereas LPS showed a dose-response curve with a well-defined optimum concentration (10 microg x ml(-1)) . The increase in lysozyme activity was accompanied by an accumulation of lysozyme gene transcript in the stimulated cells . Recombinant human tumor necrosis factor alpha, known for its ability to stimulate lysozyme in human macrophages and to elevate respiratory burst activity of rainbow trout macrophages, failed to stimulate lysozyme production of Atlantic salmon macrophages . Macrophages isolated from fish suffering from a non-lethal Ichthyobodo necator infection displayed a highly increased ability to produce lysozyme in response to both beta-glucan and LPS . As in higher vertebrates, lysozyme production may reflect the differentiation stage of the Atlantic salmon macrophages as well as a direct activation of lysozyme gene transcription by biological response modifiers . The rather late increase in lysozyme production induced by beta-glucan and LPS may thus be explained by stimulation of differentiation of the macrophages in culture eventually combined with direct activation of transcription of the lysozyme gene.

Proc R Soc Lond B Biol Sci, 2001 Feb 22, 268(1465), 393 - 8
Coevolution between a cockroach and its bacterial endosymbiont: a biogeographical perspective; Clark JW et al.; Cryptocercus are subsocial, xylophagous cockroaches that live in temperate forests . Like other cockroaches, Cryptocercus harbour endosymbiotic bacteria in their fat bodies . Two species of Cryptocercus occur in the palaearctic, one each in eastern Russia and south-central China . In the USA, there are five species: one in the north-west and four in the south-east . Little is known about the relationship between the Eurasian and North American Cryptocercus or the causes of the disjunct distribution . Here, a molecular phylogeny for six out of the seven Cryptocercus species and their endosymbionts is inferred in an attempt to understand the evolution and biogeography of the genus . Our analysis showed that the North American Cryptocercus are monophyletic, suggesting that a single colonization event was followed by vicariance . There was complete concordance between the host and endosymbiont phylogenetic trees . Divergence estimates based on endosymbiont DNA sequences suggested that the palaearctic and nearctic Cryptocercus diverged 70-115 million years (Myr) ago and the eastern- and western-USA species diverged 53-88 Myr ago . These divergence estimates were correlated with biogeographical events, and a hypothesis is presented to explain the current distribution of Cryptocercus . Our findings suggest that Cryptocercus has had a long evolutionary history, dating back to the Jurassic.

Ann N Y Acad Sci, 2000, 917, 165 - 8
Increased sensitivity of the baroreceptor reflex after bacterial endotoxin; Rogausch H et al.; Lipopolysaccharide (LPS), an endotoxin that elicits the production of several cytokines, induces cardiovascular changes characterized by increased perfusion of immune organs and compensatory sympathetic vasoconstriction in other tissues . We therefore hypothesized that to adapt to altered blood flow distribution following LPS administration, changes in the sensitivity of reflexes that control blood pressure would occur . Our data show that the sensitivity of the baroreceptor reflex increases significantly two and three hours after the intravenous administration of a subpyrogenic dose of the endotoxin . This change in sensitivity that could occur at peripheral or central levels may underlie necessary adjustments of cardiovascular mechanisms during the course of certain immune responses.

FEMS Microbiol Lett, 2001 Mar 15, 196(2), 135 - 9
Inhibition of bacterial RNA polymerase by the cyanobacterial metabolites 12-epi-hapalindole E isonitrile and calothrixin A; Doan NT et al.; The alkaloid 12-epi-hapalindole E isonitrile, from a cyanobacterial Fischerella species, and the indolophenanthridine calothrixin A, from Calothrix, inhibited Escherichia coli RNA polymerase competitively with respect to ATP, and non-competitively with respect to UTP . The inhibition was dependent on the order of addition of the inhibitors . The K(I) values, with ATP as the variable substrate, were 1.3+/-0.2 mM and 0.23+/-0.11 mM, respectively . Based on comparisons with the sensitivity of whole cells to these inhibitors, it is concluded that other targets in addition to RNA polymerase may also be implicated in their action.

Nucleic Acids Res, 2001 Apr 1, 29(7), 1602 - 7
Phylogenetic analysis of tmRNA genes within a bacterial subgroup reveals a specific structural signature; Felden B et al.; Bacterial tmRNA mediates a trans-translation reaction, which permits the recycling of stalled ribosomes and probably also contributes to the regulated expression of a subset of genes . Its action results in the addition of a small number of C-terminal amino acids to protein whose synthesis had stalled and these constitute a proteolytic recognition tag for the degradation of these incompletely synthesized proteins . Previous work has identified pseudoknots and stem-loops that are widely conserved in divergent bacteria . In the present work an alignment of tmRNA gene sequences within 13 beta-proteobacteria reveals an additional sub-structure specific for this bacterial group . This sub-structure is in pseudoknot Pk2, and consists of one to two additional stem-loop(s) capped by stable GNRA tetraloop(s) . Three-dimensional models of tmRNA pseudoknot 2 (Pk2) containing various topological versions of the additional sub-structure suggest that the sub-structures likely point away from the core of the RNA, containing both the tRNA and the mRNA domains . A putative tertiary interaction has also been identified.

EMBO Rep, 2001 Mar, 2(3), 229 - 33
Bacterial Na(+)-ATP synthase has an undecameric rotor; Stahlberg H et al.; Synthesis of adenosine triphosphate (ATP) by the F(1)F(0) ATP synthase involves a membrane-embedded rotary engine, the F(0) domain, which drives the extra-membranous catalytic F(1) domain . The F(0) domain consists of subunits a(1)b(2) and a cylindrical rotor assembled from 9-14 alpha-helical hairpin-shaped c-subunits . According to structural analyses, rotors contain 10 c-subunits in yeast and 14 in chloroplast ATP synthases . We determined the rotor stoichiometry of Ilyobacter tartaricus ATP synthase by atomic force microscopy and cryo-electron microscopy, and show the cylindrical sodium-driven rotor to comprise 11 c-subunits.

Lett Appl Microbiol, 2001 Mar, 32(3), 211 - 4
Sensitive plate assay for screening and detection of bacterial polyurethanase activity; Howard GT et al.; AIMS: A plate assay to screen and detect bacterial polyurethanase in agar medium containing a colloidal polyester-polyurethane and rhodamine B is presented . METHODS AND RESULTS: Substrate hydrolysis causes the formation of orange fluorescent halos visible upon u.v . irradiation . The logarithm of polyurethanase activity from a purified polyurethanase protein is linearly correlated with the diameter of halos, thereby allowing quantification of polyurethanase activities ranging from 0.81 to 7.29 Units . CONCLUSIONS: The potential advantages of this system are in identification and recovery of viable polyurethanolytic bacteria and quantification of polyurethanase activity . SIGNIFICANCE AND IMPACT OF THE STUDY: These advantages are derived largely from the intense fluorescence observed due to the hydrolysis of substrate reacting with rhodamine B allowing for the use of low substrate concentrations and corresponding decrease in time required detecting low levels of enzyme activity.

J Biomater Sci Polym Ed, 2000, 11(11), 1227 - 37
The influence of thrombus components in mediating bacterial adhesion to biomaterials; Lamba NM et al.; Thrombosis and infection represent the two largest limiting factors determining the long term success of implanted biomaterials . Infections associated with biomaterials are difficult to treat, and appear to evade the host defense systems . Mechanisms relating infection to thrombosis are described . Investigations into the role of receptors in mediating adhesion to thrombi are also discussed, in addition to strategies to reduce bacterial adhesion to biomaterial surfaces.

MMWR Morb Mortal Wkly Rep, 1999 Sep 10, 48(35), 773 - 7
Resurgent bacterial sexually transmitted disease among men who have sex with men--King County, Washington, 1997-1999; Development of two bacterial artificial chromosome shuttle vectors for a recombination-based cloning and regulated expression of large genes in mammalian cells; Harvard Institute of Human Genetics, Harvard Medical School, Beth Israel Deaconess Medical Center, 4 Blackfan Circle, Boston, MA 02115, USAMost conditional expression vectors designed for mammalian cells have been valuable systems for studying genes of interest by regulating their expressions . The available vectors, however, are reliable for the short-length cDNA clones and not optimal for relatively long fragments of genomic DNA or long cDNAs . Here, we report the construction of two bacterial artificial chromosome (BAC) vectors, capable of harboring large inserts and shuttling among Escherichia coli, yeast, and mammalian cells . These two vectors, pEYMT and pEYMI, contain conditional expression systems which are designed to be regulated by tetracycline and mouse interferons, respectively . To test the properties of the vectors, we cloned in both vectors the green fluorescence protein (GFP) through an in vitro ligation reaction and the 17.8-kb-long X-inactive-specific transcript (Xist) cDNA through homologous recombination in yeast . Subsequently, we characterized their regulated expression properties using real-time quantitative RT-PCR (TaqMan) and RNA-fluorescent in situ hybridization (FISH) . We demonstrate that these two BAC vectors are good systems for recombination-based cloning and regulated expression of large genes in mammalian cells .

Biosens Bioelectron, 2001 Jan, 16(1-2), 109 - 13
A fiber-optic lactate sensor based on bacterial cytoplasmic membranes; Ignatov SG et al.; A new type of fiber-optic biosensor based on bacterial cytoplasmic membranes (CPM) as the biological recognition element and an oxygen sensitive dye layer as the transducer is described for the detection of lactate . CPMs from bacteria with an induced lactate oxidase system are adsorbed onto a cellulose disk . The disk is fixed mechanically over an oxygen sensitive siloxane layer on the distal end of an optical fiber . This system detects lactate with no interference from glucose, fructose or glutamic acid.

Am Surg, 2000 Oct, 66(10), 947 - 51
Isoproterenol inhibits bacterial lipopolysaccharide-stimulated release of tumor necrosis factor-alpha from human heart tissue; Smart KR Jr et al.; Recent evidence suggests that inflammatory cytokines, particularly tumor necrosis factor alpha (TNF-alpha), may play a role in heart disease . Elevated plasma levels of the cytokine have been reported in congestive heart failure and severe angina and after myocardial infarction . The exact role of TNF-alpha in heart disease and how production is stimulated and regulated in the heart are current areas of investigation . Regarding regulation of production, isoproterenol elevates cyclic AMP and inhibits TNF-alpha release in macrophages . Therefore we hypothesized that stimulation of beta-adrenergic receptors of the sympathetic nervous system would inhibit release of the cytokine from heart tissue . With Institutional Review Board approval and patient consent atrial tissue was obtained during preparation for cardiac bypass . The tissue was divided into segments, placed in culture medium, and incubated for various times in the presence or absence of lipopolysaccharide (LPS) (20 microg/mL) and/or isoproterenol (1 microM) . The medium was removed and analyzed for biologically active TNF-alpha by the L929 cell cytotoxicity assay . Tissue samples were weighed and TNF-alpha release was expressed as pg TNF-alpha/mg tissue . Initially, to determine the time course of release, measurements were made at 2, 5, 10, 15, 30, 60, 120, 180, and 360 minutes after the addition of LPS . Elevated TNF-alpha levels in the culture medium were reliably detected at 360 minutes after exposure to LPS . In atrial tissue obtained from seven patients TNF-alpha released into the culture medium at 360 minutes was 6 +/- 3 pg/mg tissue . In the presence of LPS, levels of the cytokine in the culture medium increased to 604 +/- 233 pg/mg tissue (P < 0.05 vs LPS alone) . When isoproterenol and LPS were simultaneously added to the culture medium release of TNF-alpha was reduced by 87 per cent to 82 +/- 40 pg/mg tissue (P < 0.05 vs LPS alone) . Our results show that activation of the beta-adrenergic receptor inhibits myocardial production of TNF-alpha . This finding suggests that the sympathetic nervous system inhibits production of the cytokine and that impaired sympathetic function in heart failure may play a role in the elevated levels of TNF-alpha.

J Math Biol, 2001 Feb, 42(2), 120 - 44
Mathematical models and simulations of bacterial growth and chemotaxis in a diffusion gradient chamber; Chiu C et al.; The diffusion gradient chamber (DGC) is a novel device developed to study the response of chemotactic bacteria to combinations of nutrients and attractants {7} . Its purpose is to characterize genetic variants that occur in many biological experiments . In this paper, a mathematical model which describes the spatial distribution of a bacterial population within the DGC is developed . Mathematical analysis of the model concerning positivity and boundedness of the solutions are given . An ADI (Alternating Direction Implicit) method is constructed for finding numerical solutions of the model and carrying out computer simulations . The numerical results of the model successfully reproduced the patterns that were observed in the experiments using the DGC.

Vet Rec, 2001 Feb 10, 148(6), 172 - 5
Acute phase protein responses to uterine bacterial contamination in cattle after calving; Sheldon IM et al.; Repeated ultrasonographic examinations and collections of blood samples and uterine lumenal swabs between seven and 28 days after calving were used to examine the relative effects of bacterial contamination and involution of the uterus on the concentrations of acute phase proteins in the blood of 26 dairy cows . The severity of bacterial contamination, as determined by the total bacterial growth score, was a significant variable for the concentrations of the acute phase proteins alpha1-acid glycoprotein (P < 0.0001), haptoglobin (P < 0.05) and ceruloplasmin (P < 0.0001) . In addition, the concentrations of alpha1-acid glycoprotein and ceruloplasmin were increased in the cows from which Escherichia coli (P < 0.0001) and Arcanobacterium pyogenes (P < 0.05), respectively, were isolated from the uterine lumen . Uterine involution, as determined by the decreasing diameter of the previously gravid uterine horn, was associated with a decrease in the concentrations of alpha1-acid glycoprotein (P < 0.005), haptoglobin (P < 0.05) and ceruloplasmin (P < 0.01) . However, the response of the acute phase proteins to bacterial contamination was independent of the day on which the samples were collected, indicating that their concentrations were increased by bacterial contamination in addition to the changes associated with uterine involution.

Dig Dis Sci, 2000 Dec, 45(12), 2313 - 9
Administration of bacterial lipopolysaccharide to rats induces heme oxygenase-1 and formation of antioxidant bilirubin in the intestinal mucosa; Otani K et al.; Heme oxygenase (HO)-1, the rate-limiting enzyme in heme degradation, is induced by oxidative stress and its major end product, bilirubin, is a potent physiological antioxidant . We studied the induction of HO-1 and bilirubin production in intestinal mucosa using a rat model of sepsis . E . coli lipopolysaccharide was administered intraperitonealy to male Wistar rats and intestinal mucosa was harvested . Intestinal lipid peroxides increased significantly at 1 hr and peaked at 170% of the control value at 5 hr . GSH significantly decreased at 3 hr, reaching the nadir of 50% of the control value at 5 hr . HO-1 mRNA was maximally induced fivefold at 3 hr and HO-1 protein maximally increased to 10 times the control value at 7.5 hr . Both bilirubin and bilirubin oxidative metabolites were maximally increased at 10 hr, to 4.3 and 3.7 times the control value, respectively . These data suggest that oxidative stress in sepsis quickly induces HO-1 in intestinal mucosa and that subsequent production of bilirubin works as an antioxidant . The small intestinal mucosa is an active participant in the general response to sepsis.

Vaccine, 2001 Mar 21, 19(17-19), 2323 - 8
Peptide and recombinant antigens for protection against bacterial middle ear infection; Bakaletz LO; Passive immunization of chinchillas with serum specific for either LB1 or for LPD-LB1 (f)(2,1,3) prior to challenge with heterologous NTHI isolates (relative to diversity in region three of P5-fimbrin), significantly inhibited the signs and incidence of otitis media (P < or = 0.01) induced by any of the challenge isolates . The ability of these antisera to induce total eradication of NTHI from the nasopharynx was not however equivalent among challenged cohorts . The data thus suggested that while early, complete eradication of NTHI from the nasopharynx was highly protective, reduction of the bacterial load to below a critical threshold level appeared to be similarly effective . Both immunogens thus remain strong vaccine candidates.

EMBO Rep, 2000 Sep, 1(3), 239 - 43
Point mutation of bacterial artificial chromosomes by ET recombination; Muyrers JP et al.; Bacterial artificial chromosomes (BACs) offer many advantages for functional studies of large eukaryotic genes . To utilize the potential applications of BACs optimally, new approaches that allow rapid and precise engineering of these large molecules are required . Here, we describe a simple and flexible two-step approach based on ET recombination, which permits point mutations to be introduced into BACs without leaving any other residual change in the recombinant product . Introduction of other modifications, such as small insertions or deletions, is equally feasible . The use of ET recombination to achieve site-directed mutagenesis opens access to a powerful use of BACs and is extensible to DNA molecules of any size in Escherichia coli, including the E . coli chromosome.

Biochimie, 2001 Jan, 83(1), 117 - 20
Conserved sequence motif at the C-terminus of the bacterial cell-division protein FtsA; Lowe J et al.; FtsA is an essential part of the septal ring structure in bacterial cell division . Two peptide-protein interactions are known in this process: FtsA and ZipA bind the C-terminus of FtsZ, the bacterial tubulin homologue, which is the first septal component to appear at the septum . Our recent crystal structure of FtsA revealed a possible peptide binding site on FtsA and a long disordered C-terminal region . Here we show that all FtsA proteins contain a conserved 10-13 residue motif at the C-terminal end that may facilitate targeting of downstream septal components.

Biochimie, 2001 Jan, 83(1), 91 - 7
Hypothesis: membrane domains and hyperstructures control bacterial division; Norris V et al.; The mechanism responsible for creating the division site in the right place at the right time in bacteria is unknown . It has been attributed to the formation of proteolipid domains in the cytoplasmic membrane surrounding the nucleoids . We interpret the growing evidence for this hypothesis by invoking hyperstructures, which exist at a level of organization intermediate between macromolecules and genes . Non-equilibrium hyperstructures comprise the genes, mRNA proteins and lipids required for a particular function such as cell division, and assemble and disassemble according to the needs of the cell.

Dis Aquat Organ, 2001 Jan 26, 44(1), 7 - 16
Pathology associated with an aquareovirus in captive juvenile Atlantic halibut Hippoglossus hippoglossus and an experimental treatment strategy for a concurrent bacterial infection; Cusack RR et al.; A large-scale mortality of larval and juvenile halibut Hippoglossus hippoglossus occurred at a semi-commercial halibut farm in Atlantic Canada . Investigation of the cause revealed aquareovirus particles in necrotic liver tissue of affected fish . Cytopathic effect on CHSE-214 cell lines occurred from all fish cultured for viruses, and the viral morphology of the particles in culture was consistent with that observed in necrotic host tissue . The virus was placed in the family of Reoviridae, genus Aquareovirus based on morphology and RT-PCR results . Multifocal hepatocellular necrosis was a consistent finding in all fish as well as acute necrosis of proximal renal tubules . Concurrent bacterial infections were present in some specimens . Fish experimentally treated with oxytetracycline or a combination of oxytetracycline and chloramine-T had a significantly lower mortality rate than untreated fish . Fish treated with chloramine-T alone had a significantly elevated mortality rate compared to controls . Despite supportive medical therapy, mortality levels in treated and untreated groups remained elevated, supporting the hypothesis that the primary pathogen was of viral origin . This is the first report of elevated mortalities in Atlantic halibut associated with an aquareovirus.

Genetika, 2001 Feb, 37(2), 183 - 9
{Effect of integrating the pJFF350 vector into the 85-MDa plasmid of Azospirillum brasilense Sp245 on bacterial flagellation and mobility}; Katsy EI et al.; Results of genetic analysis of three derivatives of Azospirillum brasilense Sp245 (strains BK570, SK051, and SK248) carrying cointegrates of plasmids 85-MDa and pJFF350 (the vector for omegon mutagenesis), which manifest abnormalities in flagellation and motility, are presented . It was shown for the first time that the integration of the suicide vector into one of Azospirillum resident plasmids is accompanied by the formation of various fusion products and changes in flagellation and motility of these bacteria, such as the loss of the polar (Fla) and lateral (Laf) flagella in SK051; inactivation of Fla and Laf in SK248; and Fla-dependent acceleration of expansion in semiliquid media in BK570.

Nat Rev Mol Cell Biol, 2000 Nov, 1(2), 110 - 9
Secrets of actin-based motility revealed by a bacterial pathogen; Cameron LA et al.; Actin-based cell motility is a complex process involving a dynamic, self-organizing cellular system . Experimental problems initially limited our understanding of this type of motility, but the use of a model system derived from a bacterial pathogen has led to a breakthrough . Now, all the molecular components necessary for dynamic actin self-organization and motility have been identified, setting the stage for future mechanistic studies.

Biologist (London), 2001 Feb, 48(1), 27 - 9
What in earth? Analysing soil bacterial communities; Hodder K; Walk into any field and you will at once see dozens of plant and animal species . However, below your feet and largely unnoticed, lies a population of unseen millions . Can the introduction of powerful molecular biological techniques increase our understanding of how these hidden bacterial communities function in their natural environment?

Mol Biotechnol, 2000 Nov, 16(3), 261 - 9
Terminal restriction fragment length polymorphism monitoring of genes amplified directly from bacterial communities in soils and sediments; Bruce KD et al.; Terminal Restriction Fragment Length Polymorphism (T-RFLP) or Fluorescent Polymerase Chain Reaction/Restriction Fragment Length Polymorphism (FluRFLP) have made a significant impact on the way in which PCR products amplified from mixed community DNA extracts have been assessed . Technically, these approaches are essentially the same . PCR products are generated that contain at one 5' end label, typically a fluorescent moiety, that will be detected by a DNA sequencing machine . Upon digestion using a specific restriction endonuclease, labeled and unlabeled fragments are generated . This restriction endonuclease is chosen such that following this digestion, each labeled fragment corresponds to a different sequence variant . During electrophoretic separation, the DNA sequencing machine detects only these labeled fragments and therefore detects only the sequence variants . The aim of this article is to describe the protocols and demonstrate that this profiling can be performed using different DNA sequencing machines . The analysis and applications of this approach are also discussed.

Physiol Res, 2000, 49(6), 703 - 10
Recovery of peripheral blood cells in irradiated mice pretreated with bacterial extract IRS-19; Mackova NO et al.; The effect of antigenic bacterial lysate IRS-19 on the recovery of blood cells was studied in mice injured by a single dose of 7 Gy irradiation . The preirradiation administration of IRS-19 accelerated the recovery of leukocytes, reticulocytes and platelets in peripheral blood . The recovery of leukocytes 9-14 days after irradiation in protected animals was accompanied by a higher level of band forms of granulocytes as well as activated lymphoid and monocytoid cells.

Transgenic Res, 2001, 10(1), 13 - 9
Accumulation of barley stripe mosaic virus is significantly reduced in transgenic wheat plants expressing a bacterial ribonuclease; Zhang L et al.; An rnc70 gene encoding a mutant bacterial ribonuclease III (RNase III) was introduced into wheat (Triticum aestivum cv . Bobwhite) by microprojectile bombardment . T1, T2, and T3 plants regenerated from three transgenic callus lines were challenged with barley stripe mosaic virus . Plants expressing RNase III exhibited a high level of resistance to the virus infection . This resistance was evidenced by the absence of virus symptoms and reduced accumulation of virions in these plants . The result demonstrates that this pathogen-targeted resistance strategy can be effectively employed in conferring resistance to viral diseases of cereal crops.

Scand J Immunol, 2001 Mar, 53(3), 240 - 4
T-cell enforced invariance of the antibody repertoire in the immune response against a bacterial carbohydrate antigen; Rademaekers A et al.; The humoral response against the bacterial polysaccharide antigen alpha(1-->3) dextran (Dex) is controlled by J558 idiotype-(Id) specific T cells . These T cells of which the cell clone 178-4 Ts is a representative by all relevant criteria, recognize J558 Id-bearing B cells in an I-Ed-restricted manner . Costimulation via CD28/B7-1 but not via CD40/CD40L leads to T-cell activation . These T cells do not only suppress B cells producing the immunoglobulin (Ig)G3 isotype but also support the survival and clonal expansion of J558 Id positive B cells both in vivo and in vitro . This T-cell mediated dominance of the J558 idiotype limits the appearance of antibodies carrying other more diverse idiotypes which appear in immunized BALB/c nu/nu mice where no regulatory T cells occur . This T-cell mediated antibody invariance could be a strategy of the immune system responding to highly conserved antigens like polysaccharides, different from those against protein antigens, where diversity is assumed to be the basis for a successful response.

Scand J Immunol, 2001 Mar, 53(3), 218 - 26
Immunomodulation using bacterial enterotoxins; Simmons CP et al.; Immunologic unresponsiveness (tolerance) is a key feature of the mucosal immune system, and deliberate vaccination by a mucosal route can effectively induce immune suppression . However, some bacterial-derived proteins, e.g . cholera toxin and the heat labile toxin of Escherichia coli, are immunogenic and immunomodulatory at mucosal surfaces and can effectively adjuvant immune responses to codelivered bystander antigens . This review summarizes some of the structural and biological characteristics of these toxins and provides examples of how these properties have been exploited for tolerance induction and mucosal vaccine development.

Scand J Immunol, 2001 Mar, 53(3), 211 - 7
Genetically manipulated bacterial toxin as a new generation mucosal adjuvant; Yamamoto M et al.; Cholera toxin (CT) and heat-labile toxin (LT) of Escherichia coli act as adjuvants for the enhancement of mucosal and serum antibody (Ab) responses to mucosally co-administered protein antigen (Ag) . Both LT and CT induce B7-2 expression on antigen-presenting cells (APCs) for subsequent co-stimulatory signalling to CD4+ T cells . CT directly affects CD4+ T cells activated via the TCR-CD3 complex with selective inhibition of Th1 responses whereas LT maintains Th1 cytokine responses with inhibition of interleukin (IL)-4 production . Interestingly, while CT failed to induce mucosal adjuvant activity in the absence of IL-4, LT did so . Nontoxic mutant (m)CTs (S61F and E112K) retain adjuvant properties by inducing CD4+ Th2 cells, which provided effective help for the Ag-specific mucosal immunoglobulin (Ig)A, as well as serum IgG1, IgE and IgA Ab responses . The mCT E112K has been shown to exhibit two distinct mechanisms for its adjuvanticity . Firstly, mCT enhanced the B7-2 expression of APCs . Secondly, this nontoxic CT derivative directly affected CD4+ T cells and selectively inhibited Th1 cytokine responses . Thus, several lines of evidence indicate that enzyme activity can be separated from adjuvant properties of CT and this offers promise for the development of safe delivery of vaccines for mucosal IgA responses.

Mol Microbiol, 2001 Mar, 39(5), 1272 - 84
Type 1 pili-mediated adherence of Escherichia coli strain LF82 isolated from Crohn's disease is involved in bacterial invasion of intestinal epithelial cells; Boudeau J et al.; We previously characterized the invasive ability of Escherichia coli strain LF82, isolated from an ileal biopsy of a patient with Crohn's disease . In the present study, we performed TnphoA insertion mutagenesis to identify genes involved in LF82 invasion of intestinal epithelial cells . Most of the non-invasive mutants had an insertion mutation within the type 1 pili-encoding operon . Two non-invasive fim mutants, which harboured an insertion within the fimI and fimF genes, still adhered but had lost the ability to induce host cell membrane elongations at the sites of contact with the epithelial cells . Transcomplementation experiments with a fim operon cloned from E . coli K-12 restored both invasive ability and the ability to induce host cell membrane elongations . Expression of the cloned LF82 or K-12 fim operon into the non-invasive laboratory strain JM109 did not confer invasive properties . Thus, these findings showed that: (i) type 1 pili-mediated adherence is involved in LF82-induced perturbation of host cell signalling responsible for membrane elongations; (ii) native shafts are required for type 1 pilus-mediated induction of membrane elongations; (iii) this active phenomenon is a key step in the establishment of the invasive process; and (iv) type 1 pili alone are not sufficient to trigger bacterial internalization.

Anaesthesia, 2001 Mar, 56(3), 231 - 4
Retention of airborne latex particles by a bacterial and viral filter used in anaesthesia apparatus; Barbara J et al.; We have developed a series of laboratory tests to evaluate the efficiency of a heat and moisture exchanger filter (Pall BB25) in retaining latex particles in order to protect allergic patients during anaesthesia . Latex particles were nebulised with cornstarch as a support and collected for assay in a flask, with or without the filter integrated into the experimental circuit . With the Pall BB25 filter in the circuit, no natural latex proteins were detected by measurement of either total protein or antigenic latex proteins . The Pall BB25 filter may represent a useful means of preventing inhalation of latex particles during anaesthesia in susceptible patients.

Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 2001 Mar, 91(3), 353 - 8
Bacterial penetration of restored cavities; Zivkovic S et al.; The aim of this study was to assess the quality of the marginal seals of 7 restoratives by means of a bacterial penetration test in vitro . Sixty intact premolars and third molars that were scheduled for extraction were used in the test . There were 2 experimental groups of teeth, as follows: (1) A class V conventional cavity and a wedge erosion cavity were prepared on the buccal surface and the lingual surface, respectively, of each tooth . (2) A class V conventional cavity and a wedge erosion cavity were prepared on the buccal surface and the lingual surface, respectively, of each tooth with a completely removed enamel layer . The cavities were then reconstructed with different restorative materials . The quality of the marginal seals was evaluated by submerging the teeth in a bacterial suspension and incubating them in an anaerobic milieu at 37 degrees C for 20 hours . The teeth were subsequently processed for histologic data and bacterial staining . The best marginal sealing in both the wedge erosion and the class V cavities was provided by the Herculite/Optibond system and the Valux Plus/Scotchbond Multipurpose system . Bacterial penetration was slightly greater with the Luxat compomer and the Dyrect compomer, as well as with Vitremer glass ionomer cement and Fuji LC glass ionomer cement . The bacterial penetration test showed that the use of restorative material does not entirely eliminate microleakage.

Comp Biochem Physiol B Biochem Mol Biol, 2001 Mar, 128(3), 565 - 73
Bacterial expression and characterization of starfish phospholipase A(2); Kishimura H et al.; Phospholipase A(2) (PLA(2)) from the pyloric ceca of the starfish Asterina pectinifera showed high specific activity and characteristic substrate specificity, compared with commercially available PLA(2) from porcine pancreas . To investigate enzymatic properties of the starfish PLA(2) in further detail, we constructed a bacterial expression system for the enzyme . The starfish PLA(2) cDNA isolated previously (Kishimura et al., 2000b . cDNA cloning and sequencing of phospholipase A(2) from the pyloric ceca of the starfish Asterina pectinifera . Comp . Biochem . Physiol . 126B, 579-586) was inserted into the expression plasmid pET-16b and the PLA(2) protein was expressed in Escherichia coli BL21 (DE3) by induction with isopropyl-beta-D(-)-thiogalactopyranoside . The recombinant PLA(2) produced as inclusion bodies was dissociated with 8 M urea and 10 mM 2-mercaptoethanol and renatured by dialyzing against 10 mM Tris--HCl buffer (pH 8.0) . Renatured PLA(2) was purified by subsequent column chromatographies on DEAE--cellulose (DE-52) and Sephadex G-50 . Although an N-terminal Ser in the native starfish PLA(2) was replaced by an Ala in the recombinant PLA(2), the recombinant enzyme showed essentially the same properties as did the native PLA(2) with respect to specific activity, substrate specificity, optimum pH and temperature, and Ca(2+) requirement.

Curr Opin Mol Ther, 2000 Feb, 2(1), 94 - 9
Live bacterial delivery systems for development of mucosal vaccines; Thole JE et al.; By expression of foreign antigens in attenuated strains derived from bacterial pathogens and in non-pathogenic commensal bacteria, recombinant vaccines are being developed that aim to stimulate mucosal immunity . Recent advances in the pathogenesis and molecular biology of these bacteria have allowed rational development of new and improved bacterial carriers and more effective gene expression systems . These advances have improved the performance and versatility of these delivery systems to induce mucosal immunity to recombinant antigens in animal models . Application of these (improved) technologies for development of human vaccines is still limited and awaits further exploration.

Anal Chem, 2001 Feb 15, 73(4), 746 - 50
Characterization of the protein subset desorbed by MALDI from whole bacterial cells; Ryzhov V et al.; This study characterizes various features of the proteins that are detected in MALDI mass spectra when whole bacteria cells are analyzed, in an effort to understand why some proteins are successfully detected and many others are not . Forty peaks observed in the mass range 4,000-20,000 Da in the spectra of Escherichia coli K-12 and 11775 are tentatively assigned to proteins in a protein database, and these proteins are characterized by cell location, copy number, pI, and hydropathicity . Those detected originate in the cytosol and generally share the traits of high abundance within the cell, strong bacisity, and medium hydrophilicity.

FEMS Microbiol Ecol, 2001 Mar, 35(1), 97 - 103
Bacterial populations and processes involved in acetate and propionate consumption in anoxic brackish sediment; Boschker HT et al.; Bacterial populations and pathways involved in acetate and propionate consumption were studied in anoxic brackish sediment from the Grosser Jasmunder Bodden, German Baltic Sea . Uptake of acetate and propionate from the porewater was studied using stable carbon isotope-labeled compounds . Labeled acetate was not produced as an intermediate during propionate uptake experiments, and propionate consumption was not affected by the addition of acetate . In parallel, incorporation of labeled acetate and propionate into phospholipid-derived fatty acids (PLFA) was studied to indicate bacterial populations involved in the consumption of these substrates . The (13)C-acetate label was mainly recovered in even-numbered PLFA (16:1omega7c, 16:0 and 18:1omega7c) . In contrast, primarily odd-numbered PLFA (a15:0, 15:0, 17:1omega6 and 17:0) and the even-numbered i16:0 were labeled after incubation with (13)C-propionate . Although single PLFA labeled with propionate are commonly found in sulfate reducers, the complete PLFA-labeling pattern does not resemble any of the know strains . However, the acetate-labeling pattern is similar to Desulfotomaculum acetoxidans and Desulfofrigus spp., two acetate-consuming, sulfate reducers . In conclusion, our data suggest that acetate and propionate were predominantly consumed by different, specialized groups of sulfate-reducing bacteria.

FEBS Lett, 2001 Mar 9, 492(1-2), 160 - 5
Crystal structure of the bacterial cell division regulator MinD; Cordell SC et al.; In bacterial cell division MinD plays a pivotal role, selecting the mid-cell over other sites . With MinC, MinD forms a non-specific inhibitor of division, that interacts with FtsZ . Specificity is provided by MinD's interaction with MinE at the mid-cell . We have solved the crystal structure of MinD-1 from Archaeoglobus fulgidus to 2.6 A by multiple anomalous dispersion . MinD is a classic nucleotide binding protein, related to nitrogenase iron proteins, which have a fold of a seven-stranded parallel beta-sheet, surrounded by alpha-helices . Although MinD, unlike the proteins it interacts with and those it is structurally related to, is a monomer, not a dimer.

Proc Natl Acad Sci U S A, 2001 Mar 13, 98(6), 3150 - 5
Direct observation of the enhancement of noncooperative protein self-assembly by macromolecular crowding: indefinite linear self-association of bacterial cell division protein FtsZ; Rivas G et al.; Recent measurements of sedimentation equilibrium and sedimentation velocity have shown that the bacterial cell division protein FtsZ self-associates to form indefinitely long rod-like linear aggregates in the presence of GDP and Mg(2+) . In the present study, the newly developed technique of non-ideal tracer sedimentation equilibrium was used to measure the effect of high concentrations-up to 150 g/liter-of each of two inert "crowder" proteins, cyanmethemoglobin or BSA, on the thermodynamic activity and state of association of dilute FtsZ under conditions inhibiting (-Mg(2+)) and promoting (+Mg(2+)) FtsZ self-association . Analysis of equilibrium gradients of both FtsZ and crowder proteins indicates that, under the conditions of the present experiment, FtsZ interacts with each of the two crowder proteins essentially entirely via steric repulsion, which may be accounted for quantitatively by a simple model in which hemoglobin, albumin, and monomeric FtsZ are modeled as effective spherical hard particles, and each oligomeric species of FtsZ is modeled as an effective hard spherocylinder . The functional dependence of the sedimentation of FtsZ on the concentrations of FtsZ and either crowder indicates that, in the presence of high concentrations of crowder, both the weight-average degree of FtsZ self-association and the range of FtsZ oligomer sizes present in significant abundance are increased substantially.

Clin Infect Dis, 2001 Mar 15, 32(6), 897 - 928 Epub 2001 Mar 14.
Ticks and tickborne bacterial diseases in humans: an emerging infectious threat; Parola P et al.; Ticks are currently considered to be second only to mosquitoes as vectors of human infectious diseases in the world . Each tick species has preferred environmental conditions and biotopes that determine the geographic distribution of the ticks and, consequently, the risk areas for tickborne diseases . This is particularly the case when ticks are vectors and reservoirs of the pathogens . Since the identification of Borrelia burgdorferi as the agent of Lyme disease in 1982, 15 ixodid-borne bacterial pathogens have been described throughout the world, including 8 rickettsiae, 3 ehrlichiae, and 4 species of the Borrelia burgdorferi complex . This article reviews and illustrate various aspects of the biology of ticks and the tickborne bacterial diseases (rickettsioses, ehrlichioses, Lyme disease, relapsing fever borrelioses, tularemia, Q fever), particularly those regarded as emerging diseases . Methods are described for the detection and isolation of bacteria from ticks and advice is given on how tick bites may be prevented and how clinicians should deal with patients who have been bitten by ticks.

Dev Comp Immunol, 2001 May, 25(4), 269 - 77
Bacterial formyl peptide mediated chemotaxis and extracellular acidification in shrimp haemocytes; Yip EC et al.; The bacterial formyl peptide N-formylmethionine-leucine-phenylalanine (fMLP) is a potent chemoattractant for mammalian neutrophils . In this study, we demonstrated the binding of fluorescent dye-conjugated-fMLP to haemocytes of the penaeid shrimp Penaeus penicillatus (Alcock), through the use of flow cytometry . Fluorescence microscopy with rhodamine-fMLP suggested that fMLP receptors are present only in sub-populations of the haemocytes: granulocytes and the semi-granular cells . In addition, fMLP dose-dependently mediated chemotaxis in sub-populations of haemocytes . Microphysiometry experiments demonstrated rapid extracellular acidification upon addition of fMLP, which is in agreement with the observation in neutrophils . t-BOC, the specific fMLP receptor antagonist, was able to block the binding, chemotaxis and extracellular acidification induced by the peptide . The ability of shrimp haemocytes to migrate toward fMLP in vitro suggests that this mechanism may be important for the accumulation of these cells in infected tissues of the shrimps.

Gene, 2001 Feb 7, 264(1), 11 - 8
Construction of a PAC vector system for the propagation of genomic DNA in bacterial and mammalian cells and subsequent generation of nested deletions in individual library members; Coren JS et al.; The BAC and PAC cloning systems allow investigators to propagate large genomic DNA fragments up to 300 kb in size in E . colicells.We describe a new PAC shuttle vector that can be propagated in both bacterial and human cells . Specifically, the P1 cloning vector pAd10sacBII was modified by the insertion of a puromycin-resistance gene (pac), the Epstein-Barr Virus (EBV) latent replication origin oriP,and the EBV EBNA1 gene . Transfection studies in HEK 293 cells demonstrated that the modified vector was stably maintained as an episome for at least 30 generations . And since pJCPAC-Mam1 contains a loxP site, genomic DNA cloned into this vector can be subjected to loxP-Cre -mediated deletion events . The transposon vector pTnPGKpuro/loxP was modified to make this system amenable to propagation in human cells by inserting pac, oriP, and EBNA1 elements into the vector (Chatterjee, P.K., Coren, J.C., 1997 . Isolating large nested deletions in PACs and BACs by in vivo selection of P1 headful-packaged products of Cre-catalyzed recombination between the loxP site in PAC and BAC and one introduced in transposition . NAR 25, 2205-2212.) . pTnPGKpuro/loxP-EBV was then used to generate deletions in an individual library member to demonstrate that all of the deletions still contain the required eukaryotic elements and that they were nested . All library members constructed in pJCPAC-Mam1 can be directly transformed into human cells to assess function . And the deletion technology can be used to aid in delineating the boundaries of genes and other cis-acting elements.

Biochim Biophys Acta, 2001 Apr 2, 1504(2-3), 311 - 8
Long-lived charge-separated states in bacterial reaction centers isolated from Rhodobacter sphaeroides; van Mourik F et al.; We studied the accumulation of long-lived charge-separated states in reaction centers isolated from Rhodobacter sphaeroides, using continuous illumination, or trains of single-turnover flashes . We found that under both conditions a long-lived state was produced with a quantum yield of about 1% . This long-lived species resembles the normal P(+)Q(-) state in all respects, but has a lifetime of several minutes . Under continuous illumination the long-lived state can be accumulated, leading to close to full conversion of the reaction centers into this state . The lifetime of this accumulated state varies from a few minutes up to more than 20 min, and depends on the illumination history . Surprisingly, the lifetime and quantum yield do not depend on the presence of the secondary quinone, Q(B) . Under oxygen-free conditions the accumulation was reversible, no changes in the normal recombination times were observed due to the intense illumination . The long-lived state is responsible for most of the dark adaptation and hysteresis effects observed in room temperature experiments . A simple method for quinone extraction and reconstitution was developed.

Eur J Clin Microbiol Infect Dis, 2001 Jan, 20(1), 65 - 7
Relationship of Ureaplasma urealyticum biovars to the presence or absence of bacterial vaginosis in pregnant women and to the time of delivery; Povlsen K et al.; In a nested case-control study, the occurrence of Ureaplasma urealyticum in cervical specimens from 84 women with idiopathic preterm delivery and from 400 women delivering at term was investigated . The two potential risk factors for preterm delivery, colonization with Ureaplasma urealyticum and bacterial vaginosis, were found to be interdependent variables . The association between these factors and preterm delivery was assessed by regression analysis . Neither colonization with Ureaplasma urealyticum (odds ratio {OR} 0.7, 95% confidence interval {CI} 0.4-1.2) nor bacterial vaginosis (OR 0.8, 95% CI 0.3-1.8) was associated with preterm delivery . In women who delivered preterm, biovar 2 was found significantly more often in those with the clinical diagnosis of bacterial vaginosis (43%) than in those without (5%) (OR 15, 95% CI 1.2-209).

Plant Physiol, 2001 Mar, 125(3), 1342 - 53
Comparative sequence analysis of colinear barley and rice bacterial artificial chromosomes; Dubcovsky J et al.; Colinearity of a large region from barley (Hordeum vulgare) chromosome 5H and rice (Oryza sativa) chromosome 3 has been demonstrated by mapping of several common restriction fragment-length polymorphism clones on both regions . One of these clones, WG644, was hybridized to rice and barley bacterial artificial chromosome (BAC) libraries to select homologous clones . One BAC from each species with the largest overlapping segment was selected by fingerprinting and blot hybridization with three additional restriction fragment-length polymorphism clones . The complete barley BAC 635P2 and a 50-kb segment of the rice BAC 36I5 were completely sequenced . A comparison of the rice and barley DNA sequences revealed the presence of four conserved regions, containing four predicted genes . The four genes are in the same orientation in rice, but the second gene is in inverted orientation in barley . The fourth gene is duplicated in tandem in barley but not in rice . Comparison of the homeologous barley and rice sequences assisted the gene identification process and helped determine individual gene structures . General gene structure (exon number, size, and location) was largely conserved between rice and barley and to a lesser extent with homologous genes in Arabidopsis . Colinearity of these four genes is not conserved in Arabidopsis compared with the two grass species . Extensive similarity was not found between the rice and barley sequences other than within the exons of the structural genes, and short stretches of homology in the promoters and 3' untranslated regions . The larger distances between the first three genes in barley compared with rice are explained by the insertion of different transposable retroelements.

J Bacteriol, 2001 Apr, 183(7), 2343 - 7
Heteromeric interactions among nucleoid-associated bacterial proteins: localization of StpA-stabilizing regions in H-NS of Escherichia coli; Johansson J et al.; The nucleoid-associated proteins H-NS and StpA in Escherichia coli bind DNA as oligomers and are implicated in gene regulatory systems . There is evidence for both homomeric and heteromeric H-NS-StpA complexes . The two proteins show differential turnover, and StpA was previously found to be subject to protease-mediated degradation by the Lon protease . We investigated which regions of the H-NS protein are able to prevent degradation of StpA . A set of truncated H-NS derivatives was tested for their ability to mediate StpA stability and to form heteromers in vitro . The data indicate that H-NS interacts with StpA at two regions and that the presence of at least one of the H-NS regions is necessary for StpA stability . Our results also suggest that a proteolytically stable form of StpA, StpA(F21C), forms dimers, whereas wild-type StpA in the absence of H-NS predominantly forms tetramers or oligomers, which are more susceptible to proteolysis.

J Microbiol Methods, 2001 Apr, 44(3), 253 - 62
Design and evaluation of PCR primers to amplify bacterial 16S ribosomal DNA fragments used for community fingerprinting; Watanabe K et al.; Denaturing gradient gel electrophoresis of PCR-amplified 16S ribosomal DNA (rDNA) fragments has frequently been applied to the fingerprinting of natural bacterial populations (PCR/DGGE) . In this study, sequences of bacterial universal primers frequently used in PCR/DGGE were compared with 16S rDNA sequences that represent recently proposed divisions in the domain Bacteria . We found mismatches in 16S rDNA sequences from some groups of bacteria . Inosine residues were then introduced into the bacterial universal primers to reduce amplification biases caused by these mismatches . Using the improved primers, phylotypes affiliated with Verrucomicrobia and candidate division OP11, were detected in DGGE fingerprints of groundwater populations, which have not been detected by PCR/DGGE with conventional universal primers.

Cell Biol Int, 2001, 25(1), 71 - 81
Mycobacterial growth in human macrophages: variation according to donor, inoculum and bacterial strain; Hoal-van Helden EG et al.; The microbicidal capacity of the macrophage is frequently evaded by mycobacteria, leading to tuberculosis (TB) . We investigated a number of parameters affecting the rate of growth of mycobacteria in human monocyte-derived macrophages (MDM) . The results show a great deal of variation in the growth of both Mycobacterium bovis BCG and M . tuberculosis H37Rv, using a large number of human macrophage donors, (132 and 40, respectively), but no correlation was seen with the TB status of the MDM donor . Clumping of the mycobacteria resulted in more vigorous growth in MDM, suggesting that inoculum size could affect disease progression . The growth rates of 17 clinical isolates of M . tuberculosis were measured in macrophages derived from three donors and no consistent or marked differences between isolates were observed over the 5-day period of growth measurement . However, all 17 clinical strains grew consistently faster than H37Rv in the same experiments .

Zh Mikrobiol Epidemiol Immunobiol, 2001 Jan-Feb, (1), 82 - 9
{Role of bacterial toxins in pathogenesis of hemolytic-uremic syndrome, caused by enterohemorrhagic Escherichia}; Egorova TN et al.; The pathogenesis of the hemolytico-uremic syndrome (HUS) caused by enterohemorrhagic E . coli (EHEC) has been studied previously rather completely . HUS is characterized by the signs of microangiopathic hemolytic anemia, thrombocytopenia with renal lesions and manifestations of transient disturbances in the functions of the central nervous system . The adherence of EHEC to enterocytes was found to occur in the terminal section of the ileum and the large intestine . This process is realized with involvement intimin, EHEC outer membrane protein . Shiga-like toxins (SLT) produced by EHEC are the leading factor of their pathogenicity . The mechanism of the toxin translocation through enterocytes is not yet clear, still there is no doubt that SLT penetrates into the systemic blood stream . This is indicated by the results of histopathological studies it possible to find the toxin traces on the membranes of endothelial cells of blood vessels . The study reveals that the cells of the vascular epithelium are highly sensitive to SLT . These cells carry receptors Gb3, also known as CD77, on their membranes . Enterohemolysin, serine protease, causing disturbances in the barrier function of the intestine, can be regarded in the pathogenesis of hemorrhagic colitis which develops as the result of the damaging action of EHEC and the above-mentioned toxins . This leads to the increased level of blood systemic bacterial lipopolysaccharides, which may play, in combination with the action of SLT, an important role in the development of multi-organ pathology in HUS patients.

Cell Tissue Res, 2001 Jan, 303(1), 129 - 36
Bacterial expression of the shrimp molt-inhibiting hormone (MIH): antibody production, immunocytochemical study and biological assay; Gu PL et al.; Molting in shrimp is controlled by the molt-inhibiting hormone (MIH) and ecdysone . MIH inhibits the synthesis of ecdysone in the Y-organ, resulting in molt suppression; it is a neuropeptide member belonging to the eyestalk CHH/MIH/GIH family . The cloning of MIH (formerly MIH-like) of the shrimp Metapenaeus ensis has been reported in a previous study . To obtain a large quantity of fusion protein for antibody production and biological assay, the cDNA encoding the shrimp MIH was inserted into the pRSET bacterial expression vector . His-tagged fusion protein was produced and purified by an Ni2+-charged affinity column . Polyclonal antibody to rMIH was subsequently obtained by immunizing rabbits with purified recombinant proteins . Results from Western blot analysis indicated that the antibody was specific . Furthermore, results from immunocytochemical analysis showed that specific cells in three different clusters of the X-organ, the sinus gland and the axonal tract of the eyestalk contain MIH . To test for the molt-inhibiting activity of rMIH, shrimp at intermolt stage were injected with rMIH and the molt cycle duration of the injected shrimp was monitored . A significant increase in molt cycle duration was recorded for the shrimp injected with the recombinant protein.

Vopr Med Khim, 2000 Nov-Dec, 46(6), 531 - 48
{Bacterial L-asparaginase and glutamin(asparagin)ase: some properties, structure and anti-tumor activity}; Sokolov NN et al.; Experimental material on structurally and functional organization, regulation of biosynthesis and activity, mechanism of action, genetic determinants, heterologous expression of bacterial L-asparaginases is accumulated . The modern approaches to isolation and purification of these enzymes, some questions of practical using in oncology in the schedules combined chemotherapy of leukemia the native and modified forms of L-asparaginases are discussed . The some results before carried out in the IBMC RAMS and number institutes of the Russia on study bacterial L-asparaginases and glutamine(asparagine)ases are summarized.

Acta Microbiol Immunol Hung, 2001, 48(1), 107 - 13
Interaction of immunosuppressive drugs in mouse experiments with special regards to bacterial translocation; Anderlik P et al.; Following intraperitoneally (i.p.) applied treatment with 12.5 mg/mouse prednisolonum (PRD) no bacterial translocation (BT) was observed in mice . The PRD treatment applied in combination with lymphotropic cytostatics as dianhydrogalactitol (30 mg/kg i.p.) or chlorpromazine (75 mg/kg i.p.) both causing BT, did not increase the mice's drug sensitivity to the used agents . According to our results, PRD can be suitable for combined application with other immunosuppressive agents as it can increase immunosuppression without increase of side-effects such as those induced by bacterial translocation.

Vopr Virusol, 2001 Jan-Feb, 46(1), 22 - 4
{Protective activity of a bacterial plasmid, bearing the gene for the tick-borne encephalitis virus NS1 nonstructural protein}; Timofeev AV et al.; Three intramuscular injections (50 micrograms each) with bacterial plasmid pMV45 carrying nonstructural gene of NS1 protein of tick-borne encephalitis (TBE) virus protected 88% Balb/c mice from lethal challenge with the virus . Antibodies to NS1 nonstructural protein were detected in the sera of vaccinated mice after the challenge . Absence of antibodies to E structural protein indicated absence of manifest infectious process in mice vaccinated with plasmid and challenged with a lethal dose of TBE virus.

Nat Biotechnol, 2001 Mar, 19(3), 268 - 72
Acceleration of potato tuber sprouting by the expression of a bacterial pyrophosphatase; Farre EM et al.; Potato is a globally important crop . Unfortunately, potato farming is plagued with problems associated with the sprouting behavior of seed tubers . The data presented here demonstrate that using transgenic technology can influence this behavior . Transgenic tubers cytosolically expressing an inorganic pyrophosphatase gene derived from Escherichia coli under the control of the tuber-specific patatin promoter display significantly accelerated sprouting . The period of presprouting dormancy for transgenic tubers planted immediately after harvest is reduced by six to seven weeks when compared to wild-type tubers . This study demonstrates a method with which to regulate dormancy, an important aspect of potato crop management.

Curr Biol, 2001 Feb 6, 11(3), R103 - 5
Bacterial conjugation: running rings around DNA; Egelman EH; Helicases are active in many aspects of DNA replication, recombination, repair and transcription . An integral membrane bacterial protein assembly involved in the transfer of DNA between cells has been shown to resemble a ring helicase, suggesting that it hydrolyzes ATP to pump DNA through a central channel.

FEBS Lett, 2001 Jan 26, 489(1), 29 - 33
Protein aggregation as bacterial inclusion bodies is reversible; Carrio MM et al.; Inclusion bodies are refractile, intracellular protein aggregates usually observed in bacteria upon targeted gene overexpression . Since their occurrence has a major economical impact in protein production bio-processes, in vitro refolding strategies are under continuous exploration . In this work, we prove spontaneous in vivo release of both beta-galactosidase and P22 tailspike polypeptides from inclusion bodies resulting in their almost complete disintegration and in the concomitant appearance of soluble, properly folded native proteins with full biological activity . Since, in particular, the tailspike protein exhibits an unusually slow and complex folding pathway involving deep interdigitation of beta-sheet structures, its in vivo refolding indicates that bacterial inclusion body proteins are not collapsed into an irreversible unfolded state . Then, inclusion bodies can be observed as transient deposits of folding-prone polypeptides, resulting from an unbalanced equilibrium between in vivo protein precipitation and refolding that can be actively displaced by arresting protein synthesis . The observation that the formation of big inclusion bodies is reversible in vivo can be also relevant in the context of amyloid diseases, in which deposition of important amounts of aggregated protein initiates the pathogenic process.

Genome Res, 2001 Mar, 11(3), 483 - 96
A bacterial artificial chromosome library for sequencing the complete human genome; Osoegawa K et al.; A 30-fold redundant human bacterial artificial chromosome (BAC) library with a large average insert size (178 kb) has been constructed to provide the intermediate substrate for the international genome sequencing effort . The DNA was obtained from a single anonymous volunteer, whose identity was protected through a double-blind donor selection protocol . DNA fragments were generated by partial digestion with EcoRI (library segments 1--4: 24-fold) and MboI (segment 5: sixfold) and cloned into the pBACe3.6 and pTARBAC1 vectors, respectively . The quality of the library was assessed by extensive analysis of 169 clones for rearrangements and artifacts . Eighteen BACs (11%) revealed minor insert rearrangements, and none was chimeric . This BAC library, designated as "RPCI-11," has been used widely as the central resource for insert-end sequencing, clone fingerprinting, high-throughput sequence analysis and as a source of mapped clones for diagnostic and functional studies.

FEBS Lett, 2001 Feb 23, 491(1-2), 94 - 8
Guide DNA technique reveals that the protein component of bacterial ribonuclease P is a modifier for substrate recognition; Tanaka T et al.; We developed a guide DNA technique with which the cleavage efficiency of pre-tRNA substrate raised in the RNase P reaction . The 20-mer guide DNAs hybridizing to the upstream region of the cleaving site enhanced the cleavage reactions of RNA substrates by Escherichia coli RNase P . This guide DNA technique was also applicable to cleavage site selection by choosing the DNA-hybridizing site . Results showed that RNase P accepts DNA/RNA double-stranded 5'-leader region with high catalytic efficiency as well as single-stranded RNA region in pre-tRNAs as substrates, which suggests that the protein component of bacterial RNase P prefers bulky nucleotides . The protein component did not affect the normal 5'-processing reaction of pre-tRNAs, but enhanced the mis-cleaving (hyperprocessing) reactions of tRNA in non-cloverleaf folding . Our results suggested that the protein component of RNase P is a modifier for substrate recognition.

Med Parazitol (Mosk), 1999 Oct-Dec, (4), 46 - 50
{Use of the recombinant bacterial strain to control blood-sucking mosquito larvae}; Ganushkina LA et al.; The recombinant strain of Methylobacillus flagellatum with the cloned synthesis gene Cry 4B of the toxic Bac . thuringiensis var . israelensis protein proved to be effective against larvae of the Anopheles stephensi, An . atroparvus, An . pulcherrimus, An . superpictus, and An . sacharovi cultured in the laboratory . The use of M . flagellatum in combination with T . pyriformis may greatly expand the scope of use of the recombinant strain to control malaria mosquito larvae . Their combined use shows a 6-fold increase in the rate of strain action and a 4-fold decrease in the concentration of the agent . The optimum effects are shown following 24-hour combined intubation of M . flagellatum and Tetrahymena pyriformis.

Pediatr Infect Dis J, 2001 Feb, 20(2), 207 - 13
Bacterial infections and inflammation in the lungs of cystic fibrosis patients; Conese M et al.; The aim of this review is to describe the role of respiratory epithelial cells in processes that contribute to the pathogenesis of lung disease in patients with cystic fibrosis.

Biomol Eng, 2001 Mar, 17(3), 75 - 82
Elicitation of predictable immune responses by using live bacterial vectors; Drabner B et al.; There is an increasing need for novel vaccines able to stimulate efficient and long-lasting responses, which have also low production costs . To confer protective immunity following vaccination, the adequate type of response should be elicited . Vaccines based on attenuated bacterial carriers have contained production and delivery costs, and are able to stimulate more potent immune responses than non-replicating formulations . The improved knowledge on carrier physiology and host response, the availability of different mutants and highly sophisticated expression tools, and the possibility of co-administering modulators enable to trigger predictable responses according to the specific needs . Recent studies support the use of attenuated bacteria not only as conventional carriers, but also as a delivery system for DNA vaccines against infectious agents and tumors . In this review we discuss the most widely used bacterial carrier systems for either antigens or nucleic acid vaccines, and the strategies which have been successfully exploited to modulate the immune responses elicited.

Biophys J, 2001 Mar, 80(3), 1395 - 405
Is there a conserved interaction between cardiolipin and the type II bacterial reaction center?
Wakeham MC, Sessions RB, Jones MR, Fyfe PK.
In a recent publication, the structural details of an interaction between the Rhodobacter sphaeroides reaction center and the anionic phospholipid diphosphatidyl glycerol (cardiolipin) were described (K . E . McAuley, P . K . Fyfe, J . P . Ridge, N . W . Isaacs, R . J . Cogdell, and M . R . Jones, 1999, Proc . Natl . Acad . Sci . U.S.A . 96:14706-14711) . This was the first crystallographic description of an interaction between this biologically important lipid and an integral membrane protein and was also the first piece of evidence that the reaction center has a specific interaction with cardiolipin . We have examined the extent to which the residues that interact with the cardiolipin are conserved in other species of photosynthetic bacteria with this type of reaction center and discuss the possibility that this cardiolipin binding site is a conserved feature of these reaction centers . We look at how sequence variations that would affect the shape of the cardiolipin binding site might affect the protein-cardiolipin interaction, by modeling the binding of cardiolipin to the reaction center from Rhodopseudomonas viridis.

Indian J Exp Biol, 2000 Feb, 38(2), 167 - 76
Development of a computer software for analysis of SDS-PAGE protein fingerprints of bacterial isolates; Saxena SK et al.; Protein fingerprinting is a widely used technique in epidemiological studies for typing bacterial strains . This study reports the development of a computer based gel analysis system . The system has the capability to analyse SDS-PAGE whole-cell protein profiles using digital image processing techniques . The software incorporates spatial and frequency domain operators for image enhancement, support for geometric correction of images and new algorithms for identification of strain tracks and protein bands . The system also provides facilities for correcting imaging defects for inter-gel comparison, similarity analysis, clustering and pictorial representation of results as a dendrogram . The software is highly interactive, user-friendly and can produce accurate results for differentiation of bacterial strains with minimal overhead of time.

Rev Med Interne, 2001 Jan, 22(1), 20 - 9
{Small intestine bacterial overgrowth: six case reports and literature review}; Karsenti D et al.; INTRODUCTION: Small intestinal bacterial overgrowth syndrome (SIBOS) has various clinical and biological presentations . Six observations are described in this review which is aimed at reporting recent data on SIBOS and proposing diagnosis and therapeutic attitudes . CURRENT KNOWLEDGE AND KEY POINTS: Chronic diarrhea, malabsorption syndrome and exsudative enteropathy are the main criteria of diagnosis . Breath hydrogen testing is commonly performed to confirm diagnosis, with a 78% sensitivity and a 89% specificity . The aim of therapy is reparation of malabsorption consequences, reduction of intestinal bacterial overgrowth, and surgical correction of intestinal stasis . In the absence of consensus, norfloxacin or amoxicillin-clavulinic acid (administered for a mean of 7 to 15 days) seem the more appropriate antibiotics . When possible, surgery represents the primary treatment of SIBOS recurrences . FUTURE PROSPECTS AND PROJECTS: Diagnosis of small intestinal bacterial overgrowth syndrome must be evoked on the basis of either surgical or medical context, i.e., the existence of chronic diarrhea, malabsorption syndrome (complete or not), and exsudative enteropathy . This review reports essential factors for diagnosis and treatment.

Anal Chem, 2001 Feb 1, 73(3), 492 - 6
Lysing bacterial spores by sonication through a flexible interface in a microfluidic system; Taylor MT et al.; Cell disruptions using ultrasonic energy transmitted through a flexible interface into a liquid region has limitations because the motion of the vibrating tip is not completely transferred into the liquid . To ensure that some degree of contact will be maintained between the ultrasonic horn tip and the flexible interface, the liquid must be pressurized . The pressure conditions that yield consistent coupling between the ultrasonic horn tip and the liquid region were explored in this study by using an analytical model of the system and test fixture experiments . The nature of the interaction between the horn tip and the flexible interface creates pulses of positive pressure rises, increase in temperature, streaming flow, and almost no cavitation in the liquid . There was sufficient energy to create a cloud of microspheres, or beads, that maintain a consistent pattern of ballistic motion in the liquid . The sonication was found to be repeatable by studying video recordings of bead motion and was shown to be statistically consistent using measurements of temperature rise . Sonication of bacterial spores to obtain measurements of released nucleic acid and SEM images of damaged spores were used to verify the effects of liquid pressure on the horn-interface-liquid coupling.

J Vector Ecol, 2000 Dec, 25(2), 229 - 39
Bacterial abundance in larval habitats of four species of Anopheles (Diptera: Culicidae) in Belize, Central America; Rejmankova E et al.; Numbers of free-living and attached bacteria were counted in surface waters from larval habitats of four species of Anopheles mosquitoes: Anopheles albimanus Wiedemann, An . darlingi Root, An . vestitipennis Dyar and Knab, and An . pseudopunctiopennis Theobald, using a direct count method and DAPI staining technique . Bacterial counts from larval habitats were compared to those from adjacent open water . Several additional variables such as total suspended solids (TSS), particulate organic carbon (POC), and dissolved organic carbon (DOC) were also recorded in order to establish possible relationships with bacterial characteristics . Our results showed that the waters from larval habitats were enriched with bacteria as well as POC and DOC compared to open water . The major component of all samples consisted of cocci, the proportion of rods was similar and there were significantly more attached rods in habitat samples than in open water samples . Anopheles vestitipennis habitats had the highest values of each of the categories of bacteria as well as of POC and DOC.

Rev Prat, 2000 Dec 15, 50(20), 2223 - 30
{Fungal and bacterial nail infections}; Feuilhade de Chauvin M; Fungal and bacterial infections are very common causes of nail deformity . The majority of fungal nail infections are caused by dermatophytes . Dermatophytosis result from a human contamination . A dermatophyte is always a pathogenic agent . Very effective drugs are available to treat dermatophyte nail infections . Yeasts of the genus Candida, notably C . albicans are the second most common cause of nail infection . The infections of nails due to Candida and bacteria are related with Candida sp and bacteria which are common commensals of the gastrointestinal tract, vagina or skin . Except C . albicans which is always a pathogen of skin, the other species of Candida and the bacteria could be a commensal of the skin, a colonizer of a dermatological disease or a true pathogen of nail . It is important to consider these different situations . More rarely, environmental moulds, most often known as saprophytic agents, can affect nails . They do not respond to conventional antifungal drugs . So, their diagnosis do not suffer any error . In order to evaluate properly the patient with possible fungal or bacterial infection of nails and also to choice an accurate therapy, the laboratory confirmation is essential . Clinical diagnosis is not sufficient to distinguish an infection of nails from a dermatological disease (psoriasis, traumatism) and to identify the responsible agent.

Cell Mol Life Sci, 2000 Dec, 57(13-14), 1880 - 93
Biosynthesis of cobalamin (vitamin B12): a bacterial conundrum; Raux E et al.; The biosynthesis of cobalamin (vitamin B12) is described, revealing how the concerted action of around 30 enzyme-mediated steps results in the synthesis of one of Nature's most structurally complex 'small molecules' . The plethora of genome sequences has meant that bacteria capable of cobalamin synthesis can be easily identified and their biosynthetic genes compared . Whereas only a few years ago cobalamin synthesis was thought to occur by one of two routes, there are apparently a number of variations on these two pathways, where the major differences seem to be concerned with the process of ring contraction . A comparison of what is currently known about these pathways is presented . Finally, the process of cobalt chelation is discussed and the structure/function of the cobalt chelatase associated with the oxygen-independent pathway (CbiK) is described.

Environ Microbiol, 2000 Dec, 2(6), 654 - 65
Bacterial chromosomal painting for in situ monitoring of cultured marine bacteria; Lanoil BD et al.; We previously described a new method, bacterial chromosomal painting (BCP), for the in situ identification of bacterial cells . Here, we describe the application of this technique to study the ecology and physiology of cultured marine pelagic bacteria from the western Sargasso Sea (WSS) . A total of 86 bacteria were isolated from seawater collected from near the surface, at a depth of 250 m and from nutrient-amended seawater incubations . The 10 bacterial isolates that were best represented in environmental genomic DNA from the WSS were selected using reverse genome probing . BCP hybridization cell counts were used to determine the depth-specific distribution of one of the alpha proteobacterial isolates, B5-6, in the WSS during two thermal stratification regimes: stratified and partially mixed . The maximum cell count measured for B5-6 at the summer deep chlorophyll maximum was approximately 4% of the total cell count . This study is the first application of BCP to natural environments.

Clin Cardiol, 2001 Feb, 24(2), 166 - 8
Native aortic valve tissue systemic embolization complicating bacterial endocarditis; Garg M et al.; Systemic embolic events are known complications of bacterial endocarditis . Embolization of prosthetic valves has previously been reported in the literature . We report a case of embolization of native aortic valve tissue to the popliteal artery as the presenting event in a patient with subacute bacterial endocarditis . To our knowledge, this rare complication has not been previously reported.

Acta Neurochir (Wien), 2000, 142(12), 1377 - 83
External ventricular drainage catheters: effect of surface heparinization on bacterial colonization and infection; Lundberg F et al.; Surface heparinization of central venous catheters has earlier been shown to reduce the frequency of bacterial colonization and septicaemia . The present study was undertaken to investigate the benefit of surface heparinization of external ventricular drainage (EVD) catheters in relation to bacterial colonization, as measured by bacterial growth and examination by a 16S-rRNA PCR assay, of catheters and of samples of cerebrospinal fluid (CSF) . Ninety-eight heparinized and one hundred unheparinized EVD catheters from the same batch of catheters were used . Twenty point five percent of the heparinized and 22.8% (p = 0.63) of the unheparinized EVD catheters were colonized with bacteria . Culture of CSF, which is the definition of clinical infection in this study, yielded growth in 10.3% of patients with heparinized and in 6.3% (p = 0.18) of those with unheparinized catheters . PCR examination yielded positive signal in 31.3% of patients with heparinized catheters and in 37.7% (p = 0.061) of patients without (CSF and catheters) . In the subgroup of patients with subarachnoid haemorrhages, there was a tendency, though not statistically significant, towards a lowered frequency of colonization with 23.1% for heparinized and 33.3% (p = 0.31) for unheparinized catheters . PCR examination did not contribute any further to the diagnostic procedure in the patients concerned . The EVD catheters are skin-penetrating devices and contamination from the skin flora is common . Skin cultures, obtained after skin disinfection and insertion of catheters, showed growth of bacteria in 62% of the patients.

Protist, 2000 Dec, 151(4), 317 - 27
Bacterial orthologues indicate the malarial plastid gene ycf24 is essential; Law AE et al.; ycf24 is a well conserved gene found in all major groups of bacteria, as well as on red algal plastid genomes and the vestigal plastid genome of apicomplexan pathogens like the malaria parasite Plasmodium falciparum (ORF470) . Some database annotations describe Ycf24 as an ABC transporter subunit, but we find the level of significance is low . To investigate ycf24's function we disrupted it in the cyanobacterium Synechocystis sp., strain PCC6803 which has a multi-copy genome . This showed ycf24 is essential, partial loss producing a terminal phenotype of chlorosis, reduced cell size, loss of DNA, and a striking arrest in cytokinesis . Attempts to disrupt the single copy of ycf24 in E . coli failed to give stable transformants . When Ycf24 was over-expressed in E . coli as a soluble fusion protein, it localized mostly as a band on either side of the nucleoid and nucleoid partitioning was aberrant . We propose the relict plastid organelle of apicomplexans retains its capacity for protein synthesis because Ycf24 is essential.

Cell Mol Life Sci, 1999 Oct 30, 56(5-6), 378 - 83
Bacterial suicide through stress; Aldsworth TG et al.; Outside of the laboratory, bacterial cells are constantly exposed to stressful conditions, and an ability to resist those stresses is essential to their survival . However, the degree of stress required to bring about cell death varies with growth phase, amongst other parameters . Exponential phase cells are significantly more sensitive to stress than stationary phase ones, and a novel hypothesis has recently been advanced to explain this difference in sensitivity, the suicide response . Essentially, the suicide response predicts that rapidly growing and respiring bacterial cells will suffer growth arrest when subjected to relatively mild stresses, but their metabolism will continue: a burst of free-radical production results from this uncoupling of growth from metabolism, and it is this free-radical burst that is lethal to the cells, rather than the stress per se . The suicide response hypothesis unifies a variety of previously unrelated empirical observations, for instance induction of superoxide dismutase by heat shock, alkyl-hydroperoxide reductase by osmotic shock and catalase by ethanol shock . The suicide response also has major implications for current {food} processing methods.

Mol Cells, 2000 Dec 31, 10(6), 723 - 7
Excision repair of 2,5-diaziridinyl-1,4-benzoquinone (DZQ)-DNA adduct by bacterial and mammalian 3-methyladenine-DNA glycosylases; Lee CS; The mechanisms of anticancer activity of 2,5-diaziridinyl-1,4-benzoquinone (DZQ) are believed to involve the alkylation of guanine and adenine bases . In this study, it has been investigated whether bacterial and mammalian 3-methyladenine-DNA glycosylases are able to excise DZQ-DNA adduct with a differential substrate specificity . DZQ-induced DNA adduct was first formed in the radiolabeled restriction enzyme DNA fragment, and excision of the DNA adduct was analyzed following treatment with homogeneous 3-methyladenine-DNA glycosylase from E . coli, rat, and human, respectively . Abasic sites generated by DNA glycosylases were cleaved by the associated lyase activity of the E . coli formamidopyrimidine-DNA glycosylase . Resolution of cleaved DNA on a sequencing gel with Maxam-Gilbert sequencing reactions showed that DZQ-induced adenine and guanine adducts were very good substrates for bacterial and mammalian enzymes . The E . coli enzyme excises DZQ-induced adenine and guanine adducts with similar efficiency . The rat and human enzymes, however, excise the adenine adduct more efficiently than the guanine adduct . These results suggest that the 3-methyladenine-DNA glycosylases from different origins have differential substrate specificity to release DZQ-DNA lesions . The use of 3-methyladenine-DNA glycosylase incision analysis could possibly be applied to quantify a variety of DNA adducts at the nucleotide level.

Int J Syst Evol Microbiol, 2001 Jan, 51(Pt 1), 157 - 68
Herbaspirillum frisingense sp . nov., a new nitrogen-fixing bacterial species that occurs in C4-fibre plants; Kirchhof G et al.; The enrichment of nitrogen-fixing bacteria from the C4-fibre plants, Spartina pectinata, Miscanthus sinensis, Miscanthus sacchariflorus and Pennisetum purpureum, with nitrogen-free semi-solid media led to the isolation of Herbaspirillum-like strains among other diazotrophic bacteria . On the basis of physiological properties, phylogenetic analysis comparing 16S rDNA sequences and DNA-DNA hybridization experiments of chromosomal DNA the new isolates could be grouped together in a new species with the proposed name Herbaspirillum frisingense sp . nov . Morphological characteristics, such as cell size and shape, colony appearance, motility and flagellation are largely identical to the known species Herbaspirillum rubrisubalbicans and Herbaspirillum seropedicae . On the basis of utilization of adipate (-), N-acetyl-D-glucosamine (+), meso-erythritol (-), L-rhamnose (-) and meso-inositol (-) Herbaspirillum frisingense sp . nov . can be distinguished from other known Herbaspirillum spp . Nitrogen-fixing capability was examined by PCR amplification of the nifD gene and an acetylene reduction assay, and was found with all isolates tested . 16S rDNA sequence similarity to the other Herbaspirillum spp . is 98.5-99.1% . In genomic DNA-DNA hybridization experiments Herbaspirillum frisingense sp . nov . forms a homogeneous group with 70-100+/-10% similarity, clearly distinct from Herbaspirillum seropedicae and Herbaspirillum rubrisubalbicans with 1-34% similarity . 16S rRNA-targeted oligonucleotide probes, specific for the whole genus Herbaspirillum and for three Herbaspirillum species were designed and are suitable for fluorescence in situ hybridization . The DNA G+C content of Herbaspirillum frisingense sp . nov . is 63+/-2 mol%, in agreement with the values of 61-65% for the genus . PCR fingerprinting exhibits a consistent pattern for groups of strains isolated from the same plant, suggesting a low genomic diversity among bacteria inhabiting C4-gramineous plant tissues . Low genetic DNA diversity seems to be common between probable endophytic bacterial isolates of the same taxon . The type strain of Herbaspirillum frisingense sp . nov . is GSF30T (= DSM 13128T).

Med Parazitol (Mosk), 2000 Oct-Dec, (4), 43 - 5
{Sensitivity of malaria mosquito larvae to bacterial insecticides in relation to handling}; Iasiukevich VV et al.; The conditions for cultivation of malaria mosquito larvae were found to exert a great impact on their sensitivity to bactoculicide . Elevating keeping temperatures for An . sacharovi and An . pulcherrimus by 10-11 degrees C causes 2-4-fold increases in LC50 . There are virtually no impacts of larval population during their development or temperature-population density relationships . Ten-fold increases in larval population density under which An . stephensi larvae have been grown result in 3-fold decreases in LC50 . There are no impacts of temperatures and temperature-population density relationships either . There are also great larval species-specific differences in their sensitivity to bactoculicide.

Quintessence Int, 2000 Apr, 31(4), 249 - 56
Design features that may influence bacterial plaque retention: a retrospective analysis of failed implants; O'Mahony A et al.; OBJECTIVE: The purpose of this study was to conduct a postmortem examination of retrieved failed dental implants to identify design characteristics that may have contributed to premature loss . METHOD AND MATERIALS: Forty-five failed implants retrieved from 40 patients with no significant risk factors for implant failure (e.g., history of smoking or diabetes) were examined by scanning electron microscopy . All implants were placed by general dentists or periodontists in private practice and had functioned for an average of 4 years . RESULTS: Several design features of currently used implants present plaque-retentive areas . Plaque accumulation occurred along the implant-transmucosal abutment interfaces, transmucosal abutment-prosthesis interfaces, implant-prosthesis interfaces, and on surfaces of the abutment, the implant, and the prosthesis . The size of the microgap between the various components, the degree of surface roughness of the restorations and abutments, the exposure of plasma-sprayed coatings and threaded surfaces of implants, and overcontouring of implant restorations contributed to plaque accumulation and provided an ideal environment for bacterial colonization . CONCLUSION: These implant features may be key precipitating or exacerbating factors in the development of peri-implant inflammation, predisposing patients to implant failure.

Environ Microbiol, 2000 Jun, 2(3), 343 - 7
Phylogenetic relationship and antifouling activity of bacterial epiphytes from the marine alga Ulva lactuca; Egan S et al.; It is widely accepted that bacterial epiphytes can inhibit the colonization of surfaces by common fouling organisms . However, little information is available regarding the diversity and properties of these antifouling bacteria . This study assessed the antifouling traits of five epiphytes of the common green alga, Ulva lactuca . All isolates were capable of preventing the settlement of invertebrate larvae and germination of algal spores . Three of the isolates also inhibited the growth of a variety of bacteria and fungi . Their phylogenetic positions were determined by 16S ribosomal subunit DNA sequencing . All isolates showed a close affiliation with the genus Pseudoalteromonas and, in particular, with the species P . tunicata . Strains of this bacterial species also display a variety of antifouling activities, suggesting that antifouling ability may be an important trait for members of this genus to be highly successful colonizers of animate surfaces and for such species to protect their host against fouling.

J Mol Microbiol Biotechnol, 2001 Jan, 3(1), 83 - 93
OspA, a lipoprotein antigen of the obligate intracellular bacterial pathogen Piscirickettsia salmonis; Kuzyk MA et al.; No effective recombinant vaccines are currently available for any rickettsial diseases . In this regard the first non-ribosomal DNA sequences from the obligate intracellular pathogen Piscirickettsia salmonis are presented . Genomic DNA isolated from Percoll density gradient purified P . salmonis, was used to construct an expression library in lambda ZAP II . In the absence of preexisting DNA sequence, rabbit polyclonal antiserum raised against P . salmonis, with a bias toward P . salmonis surface antigens, was used to identify immunoreactive clones . Catabolite repression of the lac promoter was required to obtain a stable clone of a 4,983 bp insert in Escherichia coli due to insert toxicity exerted by the accompanying radA open reading frame (ORF) . DNA sequence analysis of the insert revealed 1 partial and 4 intact predicted ORF's . A 486 bp ORF, ospA, encoded a 17 kDa antigenic outer surface protein (OspA) with 62% amino acid sequence homology to the genus common 17 kDa outer membrane lipoprotein of Rickettsia prowazekii, previously thought confined to members of the genus Rickettsia . Palmitate incorporation demonstrated that OspA is posttranslationally lipidated in E . coli, albeit poorly expressed as a lipoprotein even after replacement of the signal sequence with the signal sequence from lpp (Braun lipoprotein) or the rickettsial 17 kDa homologue . To enhance expression, ospA was optimized for codon usage in E . coli by PCR synthesis . Expression of ospA was ultimately improved (approximately 13% of total protein) with a truncated variant lacking a signal sequence . High level expression (approximately 42% tot . prot.) was attained as an N-terminal fusion protein with the fusion product recovered as inclusion bodies in E . coli BL21 . Expression of OspA in P . salmonis was confirmed by immunoblot analysis using polyclonal antibodies generated against a synthetic peptide of OspA (110-129) and a strong antibody response against OspA was detected in convalescent sera from coho salmon (Oncorhynchus kisutch).

Lupus, 2000, 9(9), 655 - 63
Immunization with a bacterial ATP-binding cassette transporter fragment suppresses autoimmunity and prolongs survival in MRL/lpr lupus-prone mice; Chang M et al.; The present study was undertaken to better define the role of the U1 70 kDa antigen in a spontaneous murine model of systemic lupus erythematosus (SLE) by testing whether immunization with the U1 70 kDa polypeptide could alter the production of antibodies against U1 70 kDa or against other small nuclear ribonucleoproteins (snRNP), modify disease expression or alter survival . We found that, while immunization with a U1 70 kDa derived fusion protein (70 KFP) tended to delay the development of anti-snRNP antibodies in the sera of MRL/lpr mice, it had no effect on autoimmune-mediated renal disease or survival . Unexpectedly, it was found that MRL/lpr mice immunized with a 367 amino acid fragment of a bacterial ATP-binding cassette transporter, MFP, had prolonged survival compared to saline injection or U1 70 kDa immunization and that this was associated with a delay in the onset of SLE-like proliferative glomerulonephritis . This is the first study, to our knowledge, in which a bacterial ATP-binding cassette transporter was shown to be beneficial in treating a murine model of SLE . We report that MFP significantly prolonged longevity in the MRL/lpr murine model of SLE compared to saline injection or 70 KFP immunization and that improved survival was associated with a delay in the onset of SLE-like glomerulonephritis.

Int J Clin Pract, 2000 Oct, 54(8), 486 - 8
The dogma of identifying occult bacterial infections in young febrile children: a survey of primary-care physicians; al-Zamil FA; Management of undifferentiated febrile illness in young children continues to be a controversial issue among primary-care physicians . A self-administered questionnaire was mailed to 600 randomly selected physicians regarding their management of children with high fever and no focus of infection at various ages: 3 weeks, 7 weeks, 4 months and 20 months . Completed questionnaires were returned by 419 (70%) physicians . Nearly 77% of physicians would hospitalize a 3-week old infant with fever and 70% would treat these infants empirically with antibiotics . Sixty-one per cent of physicians would hospitalize a 7-week-old infant with fever and 46% would treat empirically with antibiotics . Approximately 80% and 93% of physicians, respectively, would not hospitalize 4- and 20-month-old infants with high fever and no focus of infection, but 72% and 59%, respectively, would treat such infants with antibiotics . There was considerable variation in the way physicians managed young febrile children with no focus of infection and the clinical approach of some physicians was remarkably different from current knowledge and recommendations.

J Neuroimaging, 2001 Jan, 11(1), 25 - 9
The use of clinical scales in depicting cerebrovascular complications in bacterial meningitis; Merkelbach S et al.; To evaluate the utility of different scales for clinical assessment, over time, in acute bacterial meningitis, the authors investigated 53 consecutive patients (mean age 53 +/- 17 years) . Clinical status on days 1, 3, 5, 8, and 14 after admission was determined by the Scandinavian Stroke Scale (SSS), Glasgow Coma Scale (GCS), and Hunt and Hess Scale (HH), and, on day 21, by the Glasgow Outcome Scale (GOS) . Transcranial Doppler examinations were performed serially to assess for disease-related arterial narrowing . This was observed in 27 patients (51%) within 2 weeks of admission . All scales were observed to correlate with the 21-day GOS . Patients with cerebral arterial narrowing had significantly decreased SSS scores between days 1 and 8 and worse GCS and HH scores between days 3 and 5 . Mean blood flow velocity in the middle cerebral artery on days 5 and 8 significantly correlated with GOS (r = 0.268, P < .008; r = -0.324, P < .003, respectively) . The use of such clinical scales allows standardized assessment of patients with bacterial meningitis and provides prognostic information . Cerebral arterial narrowing was observed to correlate with neurologic impairment.

Tex Heart Inst J, 2000, 27(4), 401 - 4
Right ventricular and septal anomalies complicated by subacute bacterial endocarditis; He QC et al.; We report the case of a 31-year-old woman with no history of heart disease . She came to the hospital with fever, dyspnea, palpitation, and edema of the lower extremities . She was found to have aortic, mitral, and pulmonary valve insufficiency, and the initial diagnosis was subacute bacterial endocarditis . At surgery, we replaced the aortic and mitral valves with mechanical prostheses and the pulmonary valve with a bioprosthesis . The prostheses were soaked intraoperatively with fluconazole and the heart chambers were irrigated with povidone-iodine to prevent infection by bacteria and fungi . We also found 2 previously unsuspected anomalies: 1 was a muscular bundle that divided the right ventricle into 2 chambers, and the other was a ventricular septal defect, 1.0 cm in diameter . We resected the muscular bundle and patched the septal defect . The patient had an uneventful postoperative course and was in New York Heart Association functional class I at the 15-month follow-up visit . We speculate that this patient's congenital anomalies made the heart more susceptible to damage from the endocarditis . Therefore, any patient who has infective endocarditis should also be examined closely for congenital defects.

J Endod, 2000 Jan, 26(1), 1 - 5
In vivo resistance of coronally induced bacterial ingress by an experimental glass ionomer cement root canal sealer; Friedman S et al.; The resistance of an experimental sealer (KT-308) to bacterial ingress was assessed in six beagle dogs . In four mandibular premolars per dog, canals were prepared, filled with condensed gutta-percha and either KT-308 or Roth 801 cement (n = 24 roots), and the pulp chambers inoculated with plaque . Two additional premolars per dog were similarly root-filled, but not inoculated (n = 12 and 11, respectively) . One incisor per dog was inoculated, but not root-filled (n = 6) . Dogs were terminated after 6 months, and jaw blocks were retrieved and processed for light microscopic examination of the periapical tissues . Inflammation about the inoculated roots was significantly lower (p < 0.03) for KT-308 (17%) than Roth 801 cement (46%) . Inflammation about the noninoculated roots did not differ significantly between KT-308 (8%) and Roth 801 cement (36%) . This study demonstrated a better functional efficacy of KT-308 than of Roth 801 cement, and validated this in vivo model for assessment of root filling materials.

Arch Biochem Biophys, 2000 Nov 15, 383(2), 178 - 84
Amorpha-4,11-diene synthase of Artemisia annua: cDNA isolation and bacterial expression of a terpene synthase involved in artemisinin biosynthesis; Chang YJ et al.; Artemisia annua, an indigenous plant to Korea, contains an antimalarial sesquiterpene, artemisinin . The first committed step of artemisinin biosynthesis is the cyclization of farnesyl diphosphate by a sesquiterpene synthase to produce an amorphane-type ring system . The aims of this research were to molecularly clone and express amorpha-4,11-diene synthase for metabolic engineering . PCR amplification of genomic DNA with a pair of primers, designed from the conserved regions of sesquiterpene synthases of several plants, produced a 184-bp DNA fragment . This fragment was used in Northern blot analysis as a probe, showing approximately 2.2 kb of a single band . Its sequence information was used to produce 2106 bp of a full-length cDNA sequence including 1641 bp of open reading frame for 546 amino acids (kcs12) through a rapid amplification of cDNA ends (RACE) . The deduced amino acid sequence displayed 36% identity with 5-epi-aristolochene synthase of Nicotiana tabacum . A soluble fraction of Escherichia coli harboring kcs12 catalyzed the cyclization of farnesyl diphosphate to produce a sesquiterpene, which was identified through GC-MS analysis as amorpha-4,11-diene.

Adv Ther, 2000 May-Jun, 17(3), 159 - 66
Potential patient preference for 3-day treatment of bacterial vaginosis: responses to new suppository form of clindamycin; Broumas AG et al.; Treatment options for bacterial vaginosis (BV) include oral and topical formulations of metronidazole or topical formulations of clindamycin . Opinions of a new vaginal suppository form of clindamycin, administered for 3 days, were obtained in a survey of 96 women who had been generally satisfied with their previous treatment . Following manipulation of the clindamycin ovule and metronidazole vaginal gel, more respondents expressed a preference for the ovule (52% vs 47%), and 54% preferred the ovule to the gel (44%) for possible future use . The gel was rated easier to use, but the ovule was found to be less messy . After manipulating both products, respondents read a profile of the ovule and identified the 3-day course as its most beneficial feature, citing this reason as increasing their likelihood of completing therapy with the ovule than with the gel . Approximately three fourths of the respondents who initially preferred the gel switched their preference to the ovule after reading the product profile . At that point, 86.5% of all respondents preferred the ovule . These results suggest the potential for improved compliance with therapy for BV with the clindamycin ovule.

J Exp Med, 2001 Jan 15, 193(2), 239 - 46
Secreted lymphotoxin-alpha is essential for the control of an intracellular bacterial infection; Roach DR et al.; Although the essential role of tumor necrosis factor (TNF) in the control of intracellular bacterial infection is well established, it is uncertain whether the related cytokines lymphotoxin-alpha (LTalpha3) and lymphotoxin-beta (LTbeta) have independent roles in this process . Using C57Bl/6 mice in which the genes for these cytokines have been disrupted, we have examined the relative contribution of secreted LTalpha3 and membrane-bound LTbeta in the host response to aerosol Mycobacterium tuberculosis infection . To overcome the lack of peripheral lymph nodes in LTalpha-/- and LTbeta-/- mice, bone marrow chimeric mice were constructed . LT-/- chimeras, which lack both secreted LTalpha3 and membrane-bound LTbeta (LT1beta2 and LT2beta1), were highly susceptible and succumbed 5 wk after infection . LTbeta-/- chimeras, which lack only the membrane-bound LTbeta, controlled the infection in a comparable manner to wild-type (WT) chimeric mice . T cell responses to mycobacterial antigens and macrophage responses in LTalpha-/- chimeras were equivalent to those of WT chimeras, but in LTalpha-/- chimeras, granuloma formation was abnormal . LTalpha-/- chimeras recruited normal numbers of T cells into their lungs, but the lymphocytes were restricted to perivascular and peribronchial areas and were not colocated with macrophages in granulomas . Therefore, LTalpha3is essential for the control of pulmonary tuberculosis, and its critical role lies not in the activation of T cells and macrophages per se but in the local organization of the granulomatous response.

Traffic, 2000 Feb, 1(2), 107 - 18
Poisons, ruffles and rockets: bacterial pathogens and the host cell cytoskeleton; Steele-Mortimer O et al.; The cytoskeleton of eukaryotic cells is affected by a number of bacterial and viral pathogens . In this review we consider three recurring themes of cytoskeletal involvement in bacterial pathogenesis: 1) the effect of bacterial toxins on actin-regulating small GTP-binding proteins; 2) the invasion of non-phagocytic cells by the bacterial induction of ruffles at the plasma membrane; 3) the formation of actin tails and pedestals by intracellular and extracellular bacteria, respectively . Considerable progress has been made recently in the characterization of these processes . It is becoming clear that bacterial pathogens have developed a variety of sophisticated mechanisms for utilizing the complex cytoskeletal system of host cells . These bacterially-induced processes are now providing unique insights into the regulation of fundamental eukaryotic mechanisms.

Cell Microbiol, 1999 Sep, 1(2), 85 - 91
Toxin entry: how bacterial proteins get into mammalian cells; Lord JM et al.; Certain bacteria secrete protein toxins that catalytically modify and disrupt essential processes in mammalian cells, often leading to cell death . As the substrates modified by these toxins are located in the mammalian cell cytosol, a catalytically active toxin polypeptide must reach this compartment in order to act . The toxins bind to receptors on the surface of susceptible cells and enter them by endocytic uptake . Endocytosed toxins initially accumulate in endosomes, where some of these proteins take advantage of the acidic environment within these organelles to form, or contribute to the formation of, protein-conducting channels through which the catalytic polypeptide is able to translocate into the cytosol . Other toxins are unable to respond to low pH in this way and must undergo intracellular vesicular transport to reach a compartment where pre-existing protein-conducting channels occur and can be exploited for membrane translocation--the endoplasmic reticulum . In this way, cell entry by this second group of toxins demonstrates that the secretory pathway of mammalian cells is completely reversible.

Pediatr Neurol, 2001 Jan, 24(1), 28 - 31
Prognostic value of EEG in neonatal bacterial meningitis; Klinger G et al.; The contribution of electroencephalogram (EEG) findings early in the course of neonatal bacterial meningitis to the prediction of severe adverse outcome was assessed in a retrospective cohort study . Infants had known outcomes to 1 year of age and an EEG performed during the first week of illness . EEGs were subclassified as follows: overall EEG description, background activity, presence of positive rolandic sharp waves, presence of seizure activity, and presence of focal abnormal activity . EEG patterns predictive of severe adverse outcome were identified by univariate and multivariate analyses . Of 101 infants admitted with bacterial meningitis, 37 had an EEG performed . Of the 37 infants, 21 had adverse outcomes; nine infants died, and 12 infants had moderate or severe disability . EEG background activity and overall EEG description were identified as predictors of adverse outcome; multivariate analysis indicated that the latter was a stronger predictor (sensitivity 88%, specificity 90%) . Infants with normal or mildly abnormal EEGs had good outcomes whereas those with moderate to markedly abnormal EEGs died or survived with adverse outcome . The accuracy of predictions increased when EEGs were repeated . In a high-risk population of infants with bacterial meningitis, moderate-to-markedly abnormal EEG reliably predicts adverse outcome.

Drug Metab Dispos, 2001 Mar, 29(3), 252 - 7
Effects of bacterial lipopolysaccharide on phenobarbital-induced CYP2B expression in mice; Li-Masters T et al.; Models of inflammation and infection, such as bacterial lipopolysaccharide (LPS), cause suppression of cytochrome P450 expression in various species, although the mechanisms involved are poorly understood . The effects of LPS on expression of phenobarbital (PB)-induced CYP2B1/2 in rats have been well characterized, but less is known about the effects of LPS on PB-induced CYP2B in mice . Since genetically manipulated mice represent an attractive model to study the mechanisms involved in the down-regulation of CYP2B expression by LPS, we investigated the effects of LPS on PB-induced CYP2B expression in mouse liver . Female C57BL/6 mice were injected with 100 mg/kg PB once daily for 4 days to induce CYP2B10 expression, and 1 mg/kg LPS was injected i.p . with the last PB dose . LPS inhibited the mRNA expression of CYP2B10 and CYP2B9 at 6 and 12 h of treatment, with the inhibitory effect more profound at 12 h . LPS also suppressed the CYP2B9 mRNA level at 24 h . However, CYP2B10 mRNA levels in mice treated with PB alone had declined markedly by 24 h after the last PB injection; therefore, no effect of LPS could be discerned . Further experiments showed that injections of 33 mg/kg PB every 8 h produced more stable CYP2B10 mRNA and enzymatic activity . Suppression of CYP2B protein level was found in LPS-treated animals at 24 h of treatment, although no significant effects were noticed at 6 and 12 h of treatment . This study suggests that LPS suppresses the expression of phenobarbital-induced CYP2B expression in mice, which resembles its effects in rats.

An Esp Pediatr, 2001 Jan, 54(1), 69 - 73
{Procalcitonin . A new marker for bacterial infection}; Casado Flores J et al.; Procalcitonin is a polypeptide present in the plasma of healthy subjects in minimal levels (< 0.5 ng/ml) . Serum procalcitonin is markedly increased a few hours after the administration of endotoxin to human volunteers and in invasive bacterial infection (sepsis, septic shock, meningitis) . Procalcitonin is moderately increased in local bacterial infection (pneumonia pyelonephritis) and is unchanged in viral infections or bacterial colonization . Procalcitonin is increased in serious bacterial infections in neonates, children and adults and is currently the best diagnostic marker of severe bacterial infection, being better than leukocyte, interleukin or C-reactive protein counts . C-reactive protein levels can be normal in severe sepsis and some viral infections . We studied 54 children with sepsis in whom plasma procalcitonin levels showed a positive correlation with the vasoactive drugs necessary to maintain cardiovascular activity . The semiquantitative procalcitonin test is simple and easy to use at the bedside at any time and in any hospital as no instruments are required . Within 30 minutes, the test identifies the type of infection and whether antibiotics are indicated.

Glia, 2001 Feb, 33(2), 97 - 106
Actions of exogenous and endogenous IL-10 on glial responses to bacterial LPS/cytokines; Molina-Holgado F et al.; The objective of this study was to investigate the actions of exogenous and endogenous IL-10 on inflammatory responses of glia . Studies were conducted in primary, mixed glial cultures from C57BL/6 (wild-type {WT}) and IL-10-deficient C57BL/6 (IL-10 knockout {KO}) neonatal mice . Activation of cultures from WT mice by bacterial lipopolysaccharide (LPS, 10 ng/ml-10 microg/ml, 24 h), caused dose-dependent increases in nitric oxide (NO) and prostaglandin E(2) (PGE(2)) release . In cultures from IL-10 KO mice, LPS elicited markedly attenuated release of NO (approximately 4-fold) and PGE(2) (approximately 17-fold) . In WT cultures, co-incubation with IL-10 (10 or 100 ng/ml, 24 h) inhibited the effects of LPS on release of NO (30%) and PGE(2) (40-50%) . In cultures from IL-10 KO mice, the addition of IL-10 (10 or 100 ng/ml, 24 h) completely abolished LPS-induced NO and PGE(2) release . LPS did, however, release of IL-1beta and TNF-alpha in cultures from all animals . Co-incubation of WT cultures with IL-10 (1, 10, or 100 ng/ml, 24 h) dose-dependently reduced the release of IL-1beta (by 0%, 15%, 75%, respectively) . In cultures from IL-10 KO mice, co-incubation with IL-10 (1, 10, or 100 ng/ml, 24 h) completely abolished LPS induced release of IL-1beta . Co-incubation with IL-10 (1, 10, 100 ng/ml) reduced, LPS-induced TNF-alpha release dose-dependently in WT cultures (by 15%, 50% and 90%) and abolished LPS-induced TNF-alpha release in cells from IL-10 KO mice . These results indicate that in glia from WT mice, exogenous IL-10 attenuates LPS-induces release of NO, PGE(2), TNF-alpha and IL-1beta . In contrast, mixed glial cultures from IL-10 KO mice showed reduced responses to LPS, but increased sensitivity to exogenous IL-10 .

Eur J Biochem, 2001 Feb, 268(4), 958 - 70
Pumping capacity of bacterial reaction centers and backpressure regulation of energy transduction; van Rotterdam BJ et al.; Transduction of free-energy by Rhodobacter sphaeroides reaction-center-light-harvesting-complex-1 (RCLH1) was quantified . RCLH1 complexes were reconstituted into liposomal membranes . The capacity of the RCLH1 complex to build up a proton motive force was examined at a range of incident light intensities, and induced proton permeabilities, in the presence of artificial electron donors and acceptors . Experiments were also performed with RCLH1 complexes in which the midpoint potential of the reaction center primary donor was modified over an 85-mV range by replacement of the tyrosine residue at the M210 position of the reaction center protein by histidine, phenylalanine, leucine or tryptophan . The intrinsic driving force with which the reaction center pumped protons tended to decrease as the midpoint potential of the primary donor was increased . This observation is discussed in terms of the control of the energetics of the first steps in light-driven electron transfer on the thermodynamic efficiency of the bacterial photosynthetic process . The light intensity at which half of the maximal proton motive force was generated, increased with increasing proton permeability of the membrane . This presents the first direct evidence for so-called backpressure control exerted by the proton motive force on steady-state cyclic electron transfer through and coupled proton pumping by the bacterial reaction center.

Cancer Lett, 2001 Mar 26, 164(2), 169 - 76
Production and characterization of a bacterial single-chain Fv fragment specific to human truncated midkine; Dansithong W et al.; The production (and characterization) of a monoclonal antibody against human truncated midkine (tMK), and the detection of tMK in G401 cells, a Wilms' tumor cell line, as well as in Wilms' tumor patient specimens, have been reported (Paul et al., Cancer Lett . 163 (2001) 245-251) . Here we report the molecular cloning and expression of this monoclonal antibody as a single-chain Fv fragment (scFv) in Escherichia coli . The scFv protein, purified by immobilized metal affinity chromatography, showed a specific affinity to recombinant tMK and native tMK in G401 cells as detected by enzyme-linked immunosorbent assay and immunofluorescence microscopy, respectively . The binding of this protein to recombinant tMK was competitive with the parental monoclonal antibody . These results suggest that this scFv can also be used for Wilms' tumor detection.

J Mol Biol, 2001 Feb 23, 306(3), 403 - 16
Anatomy of a preferred target site for the bacterial insertion sequence IS903; Hu WY et al.; Like many transposons the bacterial insertion sequence IS903 was thought to insert randomly . However, using both genetic and statistical approaches, we have derived a target site for IS903 that is used 84% of the time . Computational and genetic analyses of multiple IS903 insertion sites predicted a preferred target consisting of a 21 bp palindromic pattern centered on the 9 bp target duplication generated during transposition . Here we show that targeting can be dissected into four components: the 5 bp flanking sequences, the most important sequences required for site-specific insertion; the 7 bp palindromic core within the target duplication; the dinucleotide pair at the transposon-target junction; and the local DNA context . Finally, using a substrate with multiple target sites we show that a target site is more likely found by a local bind-and-slide model and not by extended DNA tracking.

Surg Endosc, 2001 Jan, 15(1), 80 - 4
The effect of gases in the intraperitoneal space on cytokine response and bacterial translocation in a rat model; Matsumoto T et al.; BACKGROUND: The aim of this study was to examine cytokine response and bacterial translocation after exposure of the intraperitoneal space to carbon dioxide (CO2), helium (He), and air (Air) in a rat model . METHODS: For this study, 120 Sprague-Dawley rats underwent anesthesia only (Control), 10 mmHg pneumoperitoneum (PP), or abdominal wall lift (AWL) . The rats were divided into five groups according to experimental procedure: Control, PP-CO2, AWL-CO2, AWL-He, and AWL-Air . At 0, 3, 6, and 24 h after the procedures, the levels of interleukin 1beta (IL-1beta) and interleukin 6 (IL-6) in both plasma and peritoneal lavage fluid (PLF) were measured, and the translocation of bacteria to the mesenteric lymph nodes was evaluated . RESULTS: The plasma IL-1beta and IL-6 levels in the PP-CO2, AWL-CO2, and AWL-He groups were significantly lower than those in AWL-Air group at 6 h (p < 0.05) . The PLF IL-1beta (at 3, 6, and 24 h) and IL-6 (at 6 h) levels in the AWL-CO2 group were significantly lower than those in the AWL-Air group (p < 0.05) . There were no significant differences in IL-1beta and IL-6 responses among the PP-CO2, AWL-CO2, and AWL-He groups . The AWL-CO2 and PP-CO2 groups had lower incidences of bacterial translocation than did the AWL-Air group (p < 0.05) . CONCLUSIONS: The results from this study suggest that the gas in the intraperitoneal space, but not the increased intraabdominal pressure, causes the alterations in host cytokine response and bacterial translocation . Carbon dioxide may play a primary role in the reduced immune response associated with laparoscopic surgery.

Arthritis Res, 2001, 3(1), 48 - 53 Epub 2000 Oct 27.
Synovial cytokine mRNA expression during arthritis triggered by CpG motifs of bacterial DNA; Deng GM et al.; Our results show that cytokines derived from macrophages play an important role in pathogenesis of arthritis triggered by CpG oligodinucleotide (CpG ODN) . IL-12 is in this respect an important immunomodulator during the development of joint inflammation.

Curr Treat Options Gastroenterol, 2001 Feb, 4(1), 7 - 14
Small Intestinal Bacterial Overgrowth Syndrome; Meyers JS et al.; Small intestinal bacterial overgrowth is found in many conditions and may present with malabsorption, diarrhea, and malnutrition . Whereas dietary modifications and supplements might help, the primary treatment strategy is the judicious use of antibiotics . The most effective antibiotics, shown either empirically or by clinical trials, are the quinolones, tetracycline, amoxicillin with clavulanic acid, clindamycin, and metronidazole . In an unpredictable fashion, some patients fail to respond to one of these antibiotics, but often will respond to a second . These conditions are often chronic and require periodic or cyclical treatment . In some conditions seen in the elderly or in hypochlorhydric patients the small intestinal bacterial overgrowth is inconsequential and does not require therapy . Surgical management is reserved for the select situations in which there is a clear-cut structural defect.

Echocardiography, 1998 May, 15(4), 381 - 384
Mobilization of a Bacterial Vegetation Visualized During Transesophageal Echocardiography; Hansen F et al.; In this report, we describe visualization during transesophageal echocardiography of an embolus leaving the heart from the mitral valve of a patient with infective endocarditis.

FEMS Immunol Med Microbiol, 2001 Feb, 30(1), 21 - 9
The influence of curli, a MHC-I-binding bacterial surface structure, on macrophage-T cell interactions; Johansson C et al.; Escherichia coli express thin surface fimbriae called curli which bind soluble matrix proteins and major histocompatibility complex (MHC)-I molecules . The present study addressed the ability of purified curli or curliated E . coli to influence peptide presentation on MHC-I, T cell proliferation and bacterial uptake by macrophages . In vitro studies with curli-proficient E . coli YMel and the isogenic curli-deficient strain YMel-1, both expressing the model antigen Crl-OVA, showed that curli expression by E . coli does not appear to influence the efficiency by which the bacteria are processed by murine macrophages for OVA(257-264) presentation on K(b) . Furthermore, curli expression by E . coli did not influence the binding of exogenously added OVA(257-264) peptide to K(b) on the surface of prefixed macrophages . In addition, neither curliated nor non-curliated heat-killed bacteria influenced proliferation of either murine or human T cells stimulated with anti-CD3 . Finally, curliated E . coli adhered to and were internalized by macrophages from C57BL/6 and MHC-I-deficient TAP1(-/-) mice equally well . Together these studies show that curli expression by E . coli does not appear to influence phagocytic processing of bacteria expressing Crl-OVA for OVA(257-264)/K(b) presentation, the binding of exogenously added OVA(257-264) to K(b) or T cell proliferation . In addition, although curli expression by E . coli enhances bacterial interaction with macrophages, curli interaction with MHC-I does not significantly contribute to this adherence.

Proc Natl Acad Sci U S A, 2001 Feb 13, 98(4), 1555 - 9 Epub 2001 Jan 30.
Electrophysiological characterization of specific interactions between bacterial sensory rhodopsins and their transducers; Schmies G et al.; The halobacterial phototaxis receptors sensory rhodopsin I and II (SRI, SRII) enable the bacteria to seek optimal light conditions for ion pumping by bacteriorhodopsin and/or halorhodopsin . The incoming signal is transferred across the plasma membrane by means of receptor-specific transducer proteins that bind tightly to their corresponding photoreceptors . To investigate the receptor/transducer interaction, advantage is taken of the observation that both SRI and SRII can function as proton pumps . SRI from Halobacterium salinarum, which triggers the positive phototaxis, the photophobic receptor SRII from Natronobacterium pharaonis (pSRII), as well as the mutant pSRII-F86D were expressed in Xenopus oocytes . Voltage-clamp studies confirm that SRI and pSRII function as light-driven, outwardly directed proton pumps with a much stronger voltage dependence than the ion pumps bacteriorhodopsin and halorhodopsin . Coexpression of SRI and pSRII-F86D with their corresponding transducers suppresses the proton transport, revealing a tight binding and specific interaction of the two proteins . These latter results may be exploited to further analyze the binding interaction of the photoreceptors with their downstream effectors.

Biochemistry, 2001 Feb 6, 40(5), 1358 - 66
Site-directed solid-state NMR measurement of a ligand-induced conformational change in the serine bacterial chemoreceptor; Murphy OJ 3rd et al.; The challenging nature of studies of membrane proteins has made it difficult to determine the molecular mechanism of transmembrane signaling . For the bacterial chemoreceptor family, there are crystal structures of the internal and external domains, structural models of the transmembrane domain, and evidence for subtle ligand-induced conformational changes, but the signaling mechanism remains controversial . We have used a novel site-directed solid-state NMR distance measurement approach, using (13)C(19)F REDOR, to measure a ligand-induced change of 1.0 +/- 0.3 A in the distance between helices alpha 1 and alpha 4 of the ligand-binding domain in the intact, membrane-bound serine receptor . This distance change is shown not to be due to motion of the side chain and thus is due to motion of either the alpha 1 or the alpha 4 helix . Additional distance measurements can be used to determine the type of backbone motion and to follow it to the cytoplasm, to test and refine current proposals for the mechanism of transmembrane signaling . This is a promising general method for high-resolution measurements of local structure in intact, membrane-bound proteins.

Scand J Immunol, 2001 Feb, 53(2), 184 - 91
Sirolimus interferes with the innate response to bacterial products in human whole blood by attenuation of IL-10 production; Jorgensen PF et al.; Current immunosuppressive strategies are aimed at abrogating the allospecific T-cell response against donor tissues or organs . However, little information is yet available on the potential influences of these drugs on innate immune responses . In order to address this, we have employed a whole blood model . Human whole blood was pretreated with sirolimus, cyclosporine A or tacrolimus in therapeutic as well as supra therapeutic doses, and subsequently stimulated with lipopolysaccharide (LPS), peptidoglycan (PepG) or lipoteichoic acid (LTA) . Plasma cytokine analyses revealed a potent inhibitory effect of sirolimus on interleukin(IL)-10 production induced by all bacterial products tested . In contrast, cyclosporine A and tacrolimus inhibited the tumour necrosis factor (TNF)-alpha production in response to LPS, but not to PepG and LTA . Using a quantitative mRNA analyses, we also observed that sirolimus significantly decreased the IL-10 mRNA accumulation to sub-basal levels in peripheral blood mononuclear cells (PBMC) . This suggests that the sirolimus inhibits IL-10 production by interfering with the IL-10 gene transcription . However, the molecular mechanism of this inhibition remains unclear . Based on the present study and observations by others, we postulate that the clinical use of the sirolimus may be associated with a dysregulated innate immune response to bacterial infection and thus an increased risk of hyperinflammation and sepsis.

Eur J Biochem, 2001 Feb, 268(3), 865 - 75
Identification of phospholipids as new components that assist in the in vitro trimerization of a bacterial pore protein; de Cock H et al.; The in vitro trimerization of folded monomers of the bacterial pore protein PhoE, into its native-like, heat- and SDS-stable form requires incubations with isolated cell envelopes and Triton X-100 . The possibility that membranes could be isolated that are enriched in assembly factors required for assembly of the pore protein was now investigated . Fractionation of total cell envelopes of Escherichia coli via various techniques indeed revealed the existence of membrane fractions with different capacities to support assembly in vitro . Fractions containing mainly inner membrane vesicles supported the formation of trimers that were associated with these membrane vesicles . However, only a proportion of these trimers were heat- and SDS-stable and these were formed with slow kinetics . In contrast, fractions containing mainly outer membrane vesicles supported formation of high amounts of heat-stable trimers with fast kinetics . We identified phospholipids as active assembly components in these membranes that support trimerization of folded monomers in a process with similar characteristics as observed with inner membrane vesicles . Furthermore, phospholipids strongly stimulate the kinetics of trimerization and increase the final yield of heat-stable trimers in the context of outer membranes . We propose that lipopolysaccharides stabilize the assembly competent state of folded monomers as a lipochaperone . Phospholipids are involved in converting the folded monomer into new assembly competent intermediate with a short half-life that will form heat-stable trimers most efficiently in the context of outer membrane vesicles . These results provide biochemical evidence for the involvement of different lipidic components at distinct stages of the porin assembly process.

Clin Exp Immunol, 2001 Jan, 123(1), 62 - 7
Neutrophil accumulation induced by bacterial lipopolysaccharide: effects of dexamethasone and annexin 1; Allcock GH et al.; Annexin 1 (ANX-1) can reduce leucocyte migration in response to cytokines and chemokines in some rodent models of inflammation . However, its effectiveness against an inflammatory stimulus as strong as bacterial lipopolysaccharide (LPS) is unknown . Thus, we have examined whether ANX-1 can modulate LPS-induced neutrophil accumulation in the rat, as assessed by intravital microscopy and by myeloperoxidase (MPO) assay . The anti-inflammatory glucocorticoid, dexamethasone (DEX) was also studied for comparison . LPS superfusion induced adhesion of leucocytes to the endothelium and a subsequent increase in emigration from rat post-capillary venules over 2 h as assessed by intravital microscopy . Either ANX-1 or DEX was able to attenuate this adhesion and emigration of leucocytes . MPO activity in the lung, kidney and ileum was elevated after a 6-h exposure to LPS (intraperitoneal), indicating accumulation of neutrophils in these tissues . DEX attenuated the LPS-induced increase in MPO in the ileum but had no effect on MPO in the lungs or kidneys . This would suggest that the underlying mechanism by which neutrophils accumulate in the ileum, and more generally in the gastrointestinal compartment, is different from other vascular beds . ANX-1 had no effect on the LPS-induced increase in MPO activity in any of the tissues studied . Thus, from these data, ANX-1 appears to reduce leucocyte adhesion and emigration induced by a short-term (2 h), but not a longer (6 h) exposure to LPS.

Clin Exp Immunol, 2001 Jan, 123(1), 56 - 61
Serial analysis of serum and ascitic fluid levels of soluble adhesion molecules and chemokines in patients with spontaneous bacterial peritonitis; Giron-Gonzalez JA et al.; The aim of this work was the evaluation of serum and ascitic fluid levels of chemokines (IL-8, growth-regulated oncogene (Gro-alpha), and monocyte chemotactic protein-1 (MCP-1)), and of soluble adhesion molecules (P-selectin, E-selectin, L-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1)) in patients with spontaneous bacterial peritonitis (SBP) . These compounds were serially analysed in serum and ascitic fluid by ELISA in patients with SBP (n = 20), non-infected cirrhotic controls (n = 12), and healthy controls (n = 15) . Infected and non-infected cirrhotic patients showed significantly higher serum levels of adhesion molecules . SBP was associated with significantly higher serum and ascitic fluid levels of IL-8, Gro-alpha and ICAM-1 and with ascitic fluid concentrations of MCP-1 . Significantly elevated serum levels of both ICAM-1 and VCAM-1 were detected in patient non-survivors after SBP . Thus, higher ascitic fluid levels of chemokines could be implicated in the peritoneal infiltrate in patients with SBP . Prognostic significance can be attributed to serum levels of ICAM-1 and VCAM-1 in these patients.

Cephalalgia, 2000 Oct, 20(8), 738 - 9
Migraine-like headache in bacterial meningitis; Lampl C et al.; The effect of sumatriptan has not been previously described in the treatment of the headache of meningitis, although this headache has similarities to migraine . This study presents the clinical features of two patients who had fulminant bacterial meningitis with migraine-like headache and who experienced no improvement in headache intensity after administration of sumatriptan 6 mg s.c . On these grounds the lack of response of this type of headache to sumatriptan is discussed.

Vaccine, 2001 Feb 8, 19(13-14), 1573 - 80
Use of live bacterial vaccine vectors for antigen delivery: potential and limitations; Medina E et al.; Most infectious agents are restricted to the mucosal membranes or their transit through the mucosa constitutes a critical step in the infection process . Therefore, the elicitation of an efficient immune response, not only at systemic, but also at mucosal level, after vaccination is highly desirable, representing a significant advantage in order to prevent infection . This goal can be only achieved, when the vaccine formulation is administered by the mucosal route . However, soluble antigens given by this route are usually poorly immunogenic . Among the available approaches to stimulate efficient mucosal responses, the use of bacterial carriers to deliver vaccine antigens, probably, constitutes one of the most successful strategies . The potential and limitations of the most extensively studied bacterial carrier systems will be discussed.

J Virol Methods, 2001 Mar, 92(1), 91 - 7
Rapid method for the isolation of full length adenoviral genomes by bacterial intermolecular homologous recombination; Youil R et al.; Recombinant adenoviruses are used widely in gene therapy research . Much work has been carried out to remove specific components of the wild type adenovirus (e.g . E1 gene) in order to make them safer for human use . In addition to such efforts, it is vitally important to ensure that the production of recombinant adenoviruses meet safety guidelines not only with regard to the absence of replication competent adenoviruses but for other variant species that may be present in a viral preparation . In this report, a time and cost efficient method is described for the isolation of full length adenovirus genomes without resorting to plaque purification . The procedure uses a bacterial homologous recombination system and results in the conversion of the double-stranded linear adenovirus genome into a circularized plasmid form that can be easily analyzed by restriction digestion, PCR, DNA sequencing or used in transient transfection studies . Also, the adenovirus plasmids that are generated may also be rescued back into virus form if needed . The entire procedure takes 4 days or less instead of weeks that plaque purification or dilution cloning requires . Furthermore, the method does not require the use of tissue culture materials or facilities . More importantly, this procedure allows for a more extensive and thorough examination of any viral preparation, since it allows for the detection of variants incapable of propagation without the assistance of co-infecting intact adenoviral genomes . Under standard conditions of plaque purification, these variant genomes are not detected . It is predicted that far more variant genomes will be observed using this rapid method than would otherwise be detected by standard plaque purification methods.

Plant Sci, 2000 Dec 7, 160(1), 165 - 176
Resistance to bacterial wilt in somatic hybrids between Solanum tuberosum and Solanum phureja; Fock I I et al.; Somatic hybrid plants were produced after protoplast electrofusion between a dihaploid potato, cv . BF15, and a wild tuber-bearing relative, Solanum phureja, with a view to transferring bacterial wilt resistance into potato lines . A total of ten putative hybrids were selected . DNA analysis using flow cytometry revealed that six were tetraploids, two mixoploids, one amphiploid and one octoploid . In the greenhouse, the putative hybrids exhibited strong vigor and were morphologically intermediate, including leaf form, flowers and tuber characteristics . The hybrid nature of the ten selected plants was confirmed by examining isoenzyme patterns for esterases and peroxidases, and analysis of RAPD and SSR markers . Analysis of chloroplast genome revealed that eight hybrids possessed chloroplast (ct) DNA of the wild species, S . phureja, and only two contained Solanum tuberosum ct type . Six hybrid clones, including five tetraploids and one amphiploid, were evaluated for resistance to bacterial wilt by using race 1 and race 3 strains of Ralstonia solanacearum, originating from Reunion Island . Inoculations were performed by an in vitro root dipping method . The cultivated potato was susceptible to both bacterial strains tested . All somatic hybrids except two were tolerant to race 1 strain, and susceptible to race 3 strain . Interestingly, the amphiploid hybrid clone showed a good tolerance to both strains.

Protein Expr Purif, 2001 Feb, 21(1), 115 - 20
Bacterial expression of an immunologically reactive PCV2 ORF2 fusion protein; Liu Q et al.; The entire coding region of open reading frame 2 (ORF2) of porcine circovirus 2 (PCV2) was linked to the 3'-end of the maltose-binding protein (MBP)-His(8)-tag gene . The fusion protein was expressed as soluble form after induction by isopropylthio-beta-d-galactoside . MBP-His(8)-ORF2 was purified to homogeneity by immobilized metal affinity chromatography based on the interaction of the polyhistidine-tag with metal ions . Expression could represent 1% of the total protein in Escherichia coli, allowing approximately 1 mg of highly purified protein to be obtained per liter of bacterial culture . The fusion protein was recognized in Western blot by anti-PCV2 polyclonal antibody and swine sera with PCV2 infection . In addition, rabbit polyclonal antibody raised against purified MBP-His(8)-ORF2 fusion protein reacted with the ORF2 protein in immunoprecipitation . The availability of this fusion protein should permit a thorough study of prevalence of PCV2 infection in large-scale serological studies of field samples .

Mol Genet Metab, 2001 Jan, 72(1), 15 - 26
Molecular cloning of the guinea pig cytomegalovirus (GPCMV) genome as an infectious bacterial artificial chromosome (BAC) in Escherichia coli; McGregor A et al.; Since cytomegalovirus (CMV) infection is highly species-specific, it is necessary to study animal cytomegaloviruses to assess viral factors which contribute to pathogenesis . The generation of recombinant viruses carrying reporter genes would provide useful tools for studying the genetics of CMV pathogenicity in vivo . We evaluated whether the guinea pig cytomegalovirus (GPCMV) was amenable to such manipulation . Metabolic selection using the guanosylphosphoribosityl transferase (gpt) gene facilitated recovery of a recombinant virus, vAM403, containing a gpt/green fluorescent protein (eGFP) cassette introduced into the HindIII "N" region of the viral genome . This virus had replication kinetics identical to wild-type virus . We next attempted to clone the GPCMV genome as a bacterial artificial chromosome (BAC) . A BAC plasmid containing a gpt/eGFP cassette and the chloramphenicol resistance marker was introduced into HindIII "N" to generate another GPCMV recombinant, vAMBGPCMV . Circular viral DNA isolated from vAMBGPCMV-infected cells was used to transform Escherichia coli . Restriction profiles revealed that the GPCMV genome had been cloned as a BAC plasmid, and transfection of BAC plasmid DNA confirmed that the BAC clone was infectious . A novel strategy based on a unique PmeI site was devised to quickly modify the BAC GPCMV plasmid . Recombinants retained the capability to replicate and express reporter genes in guinea pigs, suggesting that these viruses will be useful for in vivo pathogenesis studies .

Mol Genet Metab, 2001 Jan, 72(1), 8 - 14
Recent advances in herpesvirus genetics using bacterial artificial chromosomes; McGregor A et al.; Although the members of the Herpesvirus family are responsible for a wide variety of human diseases, advances in the understanding of viral molecular mechanisms of pathogenesis have been hampered by the large size of herpesvirus genomes, rendering the viruses difficult to experimentally manipulate . Better techniques have been needed to facilitate mutagenesis of herpesvirus genomes, allowing for the assessment of the role of specific viral gene products in replication, immunity, and pathogenesis . Homologous recombination with plasmids containing genes of interest flanked by selectable markers has been a successful method for generating viral mutants, as has the generation of recombinant virus from transfection of cosmid clones . Although these efforts to generate recombinant viruses have met with modest success, the protocols have been cumbersome . More recently, a novel technique for the manipulation of herpesvirus genomes has been developed . This technology utilizes bacterial F plasmids, and allows for the stable cloning of herpesvirus genomes as bacterial artificial chromosomes (BACs) in Escherichia coli . Once cloned, such BACs are stable, and DNA purified from E . coli is infectious, fully capable of reproducing replication-competent virus . Manipulation of herpesvirus genomes is now feasible using the powerful techniques of bacterial genetics, and should facilitate a better understanding of the molecular pathogenesis of herpesvirus infections .

Nucleic Acids Res . 2001 Feb 1;29(3):E14.
A new method for generating point mutations in bacterial artificial chromosomes by homologous recombination in Escherichia coli; Lalioti M et al.; Bacterial artificial chromosomes (BACs) and P1 artificial chromosomes (PACs), which contain large fragments of genomic DNA, have been successfully used as transgenes to create mouse models of dose-dependent diseases . They are also potentially valuable as transgenes for dominant diseases given that point mutations and/or small rearrangements can be accurately introduced . Here, we describe a new method to introduce small alterations in BACs, which results in the generation of point mutations with high frequency . The method involves homologous recombination between the original BAC and a shuttle vector providing the mutation . Each recombination step is monitored using positive and negative selection markers, which are the Kanamycin-resistance gene, the sacB gene and temperature-sensitive replication, all conferred by the shuttle plasmid . We have used this method to introduce four different point mutations and the insertion of the ss-galactosidase gene in a BAC, which has subsequently been used for transgenic animal production.

Sex Transm Infect, 2001 Feb, 77(1), 63 - 8
Is bacterial vaginosis a sexually transmitted infection?
Morris MC, Rogers PA, Kinghorn GR.
OBJECTIVES: To determine whether the risk factors associated with bacterial vaginosis (BV) are consistent with it being a sexually transmitted infection (STI) by comparing the characteristics of women with BV with those of women with infections recognised as being sexually and non-sexually transmitted . METHODS: A prospective cross sectional study was conducted among female patients presenting for diagnosis at a genitourinary medicine clinic in Sheffield between January 1996 and September 1998 . Demographic and behavioural characteristics were reported from patient records and a standardised questionnaire was administered . Risk factor models for BV, two STIs, and two non-STIs were compiled using a multivariable logistic regression analysis . RESULTS: Among the 8989 females under 45 years eligible for analysis, the prevalence of BV was 12.9% . Risk factors associated with BV included some in common with gonorrhoea and Chlamydia trachomatis and some that were not associated with these two STIs . Risk factors common to BV and the STIs included having had more than one sexual partner in the past 3 months, having a history of a bacterial STI, being of black Caribbean ethnicity, and living in a deprived area . However, BV had a contrasting age profile, being most prevalent among those over the age of 30 . BV was also more common in those who were divorced . CONCLUSIONS: BV is associated with some factors related to the acquisition of gonorrhoea and Chlamydia trachomatis . However, infection is not only determined by those factors and therefore factors other than sexual activity may be important in the development of the condition.

Proc Natl Acad Sci U S A, 2001 Jan 30, 98(3), 892 - 7
Bacterial RNA polymerase subunit omega and eukaryotic RNA polymerase subunit RPB6 are sequence, structural, and functional homologs and promote RNA polymerase assembly; Minakhin L et al.; Bacterial DNA-dependent RNA polymerase (RNAP) has subunit composition beta'betaalpha(I)alpha(II)omega . The role of omega has been unclear . We show that omega is homologous in sequence and structure to RPB6, an essential subunit shared in eukaryotic RNAP I, II, and III . In Escherichia coli, overproduction of omega suppresses the assembly defect caused by substitution of residue 1362 of the largest subunit of RNAP, beta' . In yeast, overproduction of RPB6 suppresses the assembly defect caused by the equivalent substitution in the largest subunit of RNAP II, RPB1 . High-resolution structural analysis of the omega-beta' interface in bacterial RNAP, and comparison with the RPB6-RPB1 interface in yeast RNAP II, confirms the structural relationship and suggests a "latching" mechanism for the role of omega and RPB6 in promoting RNAP assembly.

J Biol Chem, 2000 Apr 14, 275(15), 11058 - 63
Bacterial lipopolysaccharide activates NF-kappaB through toll-like receptor 4 (TLR-4) in cultured human dermal endothelial cells . Differential expression of TLR-4 and TLR-2 in endothelial cells; Faure E et al.; A missense mutation in the cytoplasmic domain of the Toll-like receptor-4 (TLR-4) has been identified as the defect responsible for lipopolysaccharide (LPS) hyporesponsiveness in C3H/HeJ mice . TLR-4 and TLR-2 have recently been implicated in LPS signaling in studies where these receptors were overexpressed in LPS non-responsive 293 human embryonic kidney cells . However, the signaling role of TLR-4 or TLR-2 in human cells with natural LPS response remains largely undefined . Here we show that human dermal microvessel endothelial cells (HMEC) and human umbilical vein endothelial cells express predominantly TLR-4 but very weak TLR-2 and respond vigorously to LPS but not to Mycobacterium tuberculosis 19-kDa lipoprotein . Transient transfection of non-signaling mutant forms of TLR-4 and anti-TLR-4 monoclonal antibody inhibited LPS-induced NF-kappaB activation in HMEC, while a monoclonal antibody against TLR-2 was ineffective . In contrast to LPS responsiveness, the ability of HMEC to respond to 19-kDa lipoprotein correlated with the expression of TLR-2 . Transfection of TLR-2 into HMEC conferred responsiveness to 19-kDa lipoprotein . These data indicate that TLR-4 is the LPS signaling receptor in HMEC and that human endothelial cells (EC) express predominantly TLR-4 and weak TLR-2, which may explain why they do not respond to 19-kDa lipoprotein . The differential expression of TLRs on human EC may have important implications in the participation of vascular EC in innate immune defense mechanisms against various infectious pathogens, which may use different TLRs to signal.

Crit Care Med, 2000 Mar, 28(3), 788 - 94
Peritoneal cytokine concentrations and survival outcome in an experimental bacterial infusion model of peritonitis; Martineau L et al.; OBJECTIVE: To correlate the dynamics of peritoneal cytokines with systemic concentrations and survival outcome . DESIGN: Randomized, controlled study using a recently developed rat model of peritonitis . SETTING: Government research facility . SUBJECTS: Male Sprague-Dawley rats . INTERVENTIONS: Infected animals (INF) received an intraperitoneal infusion of 6.5 x 10(8) colony-forming units of Escherichia coli over 12 hrs, whereas control rats (CON) received a sterile inoculum . Peritoneal fluid and plasma samples were obtained from all rats at the end of the 12-hr infusion period as well as from all animals that survived the 7-day study (SURV) . MEASUREMENTS AND MAIN RESULTS: Interleukin (IL)-1beta concentration in the peritoneal fluid at 12 hrs tended to be higher in nonsurvivors (NONSURV) than in SURV . Tumor necrosis factor-alpha and IL-6 peritoneal concentrations at 12 hrs were significantly greater in NONSURV than in SURV . There were no significant differences in IL-2 and IL-4 peritoneal concentrations at 12 hrs between SURV and NONSURV . Although the concentrations of IL-1beta and tumor necrosis factor-alpha in the peritoneal fluid of INF decreased gradually during the study, these concentrations remained significantly higher than those of CON at 7 days . In contrast, peritoneal IL-2 concentrations remained lower in INF than in CON for most of the experiment . Peritoneal IL-6 concentrations in INF were transiently elevated above those of CON for 12 hrs . Cytokine concentrations in the peritoneal fluid of INF were always higher than those in plasma, which remained relatively unchanged throughout the study . For most of the variables as . sessed, CON showed no significant changes compared with INF . CONCLUSIONS: This model of peritonitis is associated with a significant and prolonged peritoneal inflammatory response that is adversely correlated with survival outcome . Our data would suggest that to be effective, novel immunotherapies should target mainly the peritoneal compartment.

Arq Neuropsiquiatr, 1999 Sep, 57(3B), 808 - 12
{Audiological assessment and follow-up after bacterial meningitis}; Couto MI et al.; Hearing loss is the more frequent sequel of bacterial meningitis in children . This study describes the audiological profile (peripheric and central) of 89 children admitted to the hospital wards with the diagnosis of bacterial meningitis . Those children attended audiological follow up, after their hospital discharge, and were submitted to audiological tests . The results showed that 85.4% among them presented normal hearing in both ears 10.1% presented bilateral neurosensorial hearing loss and 4.5% presented unilateral neurosensorial hearing loss . The results from the auditory processing skills assessment showed that 10% of those children presented auditory localization and recognition of sentences with competitive messages (Paediatric Sentences Identification-ipsilateral) disorders.

J Biol Chem, 2001 Mar 30, 276(13), 10407 - 12 Epub 2001 Jan 02.
Effect of oxygen on induction of the cystine transporter by bacterial lipopolysaccharide in mouse peritoneal macrophages; Sato H et al.; Amino acid transport in mouse peritoneal macrophages is mediated by several membrane carriers with different substrate specificity and sensitivity to environmental stimuli . We reported previously that transport activities of cystine and arginine in the macrophages were induced markedly by low concentrations of bacterial lipopolysaccharide (LPS) . It is known that a variety of macrophage functions are affected by ambient oxygen tension . In this study, we have investigated the effects of oxygen on the induction of amino acid transport activity by LPS and found that the induction of cystine, but not arginine, transport activity was dependent on the ambient oxygen tension . When the macrophages were cultured with 2% O(2) in the presence of 1 ng/ml LPS, induction of cystine transport activity was reduced by approximately 70% compared with cells cultured under normoxic conditions . In macrophages, transport of cystine is mediated by a Na(+)-independent anionic amino acid transporter named system x(c)(-) . System x(c)(-) is composed of two protein components, xCT and 4F2hc, and the expression of xCT was closely correlated with system x(c)(-) activity . A putative NF-kappaB binding site was found in the 5'-flanking region of the xCT gene, but the enhanced expression of xCT by LPS and oxygen was not mediated by NF-kappaB binding . An increase in intracellular GSH in macrophages paralleled induction of xCT, but not gamma-glutamylcysteine synthetase . These results suggest the importance of system x(c)(-) in antioxidant defense in macrophages exposed to LPS and oxidative stress.

Gut, 2001 Feb, 48(2), 206 - 11
The role of small intestinal bacterial overgrowth, intestinal permeability, endotoxaemia, and tumour necrosis factor alpha in the pathogenesis of non-alcoholic steatohepatitis; Wigg AJ et al.; BACKGROUND: Small intestinal bacterial overgrowth may contribute to the development of non-alcoholic steatohepatitis, perhaps by increasing intestinal permeability and promoting the absorption of endotoxin or other enteric bacterial products . AIMS: To investigate the prevalence of small intestinal bacterial overgrowth, increased intestinal permeability, elevated endotoxin, and tumour necrosis factor alpha (TNF-alpha) levels in patients with non-alcoholic steatohepatitis and in control subjects . PATIENTS AND METHODS: Twenty two patients with non-alcoholic steatohepatitis and 23 control subjects were studied . Small intestinal bacterial overgrowth was assessed by a combined (14)C-D-xylose and lactulose breath test . Intestinal permeability was assessed by a dual lactulose-rhamnose sugar test . Serum endotoxin levels were determined using the limulus amoebocyte lysate assay and TNF-alpha levels using an ELISA . RESULTS: Small intestinal bacterial overgrowth was present in 50% of patients with non-alcoholic steatosis and 22% of control subjects (p=0.048) . Mean TNF-alpha levels in non-alcoholic steatohepatitis patients and control subjects were 14.2 and 7.5 pg/ml, respectively (p=0.001) . Intestinal permeability and serum endotoxin levels were similar in the two groups . CONCLUSIONS: Patients with non-alcoholic steatohepatitis have a higher prevalence of small intestinal bacterial overgrowth, as assessed by the (14)C-D-xylose-lactulose breath test, and higher TNF-alpha levels in comparison with control subjects . This is not accompanied by increased intestinal permeability or elevated endotoxin levels.

Oral Microbiol Immunol, 2000 Feb, 15(1), 48 - 52
Adherence of Porphyromonas gingivalis to gingival epithelial cells: modulation of bacterial protein expression; Agnani G et al.; The protein profiles of Porphyromonas gingivalis (ATCC 33277 and W83) bound to KB gingival epithelial cells were analyzed by SDS-PAGE and immunoblotting . We found that a 51-kDa component was formed in bacteria that adhered to the KB cells, whereas 26- to 29-kDa bands were less intensive, in contrast to the protein profile of free bacteria . P . gingivalis ATCC 33277 incubated with protease-treated KB cells retained the profile of free bacteria . These results demonstrate the specificity of bacterial recognition of eukaryotic membrane components.

Oral Microbiol Immunol, 2000 Aug, 15(4), 232 - 7
Relationships between the onset of pustulosis palmaris et plantaris, periodontitis and bacterial heat shock proteins; Ishihara K et al.; Relationships between the onset of pustulosis palmaris et plantaris, periodontitis and heat shock proteins were studied by using enzyme-linked immunosorbent assay to examine levels of immunoglobulin G (IgG) against Escherichia coli GroEL, a recombinant DnaJ of Actinobacillus actinomycetemcomitans heat shock protein, a synthetic peptide made from the 180th to the 188th amino acids of Mycobacterium bovis BCG Hsp65, and a recombinant human Hsp60, in sera obtained from 43 pustulosis palmaris et plantaris patients judged to have chronic infectious diseases of the oral cavity . We found that the titers of IgG against E . coli GroEL and A . actinomycetemcomitans DnaJ in the sera from pustulosis palmaris et plantaris patients were significantly higher than those in the control group, whereas the titers of IgG against the synthetic M . bovis Hsp65 and the recombinant Hsp60 did not differ significantly . Periodontal therapy and extraction of teeth with periapical infectious resulted in remission of pustulosis palmaris et plantaris and a statistically significant reduction in the levels of IgG against E . coli GroEL in 9 of the 22 patients (41%) examined . We also found that the IgG levels against A . actinomycetemcomitans DnaJ in 6 serum samples of 16 (37%) were reduced, but not significantly, after the treatment . These results suggest that the IgG responses to heat shock proteins partially induced by oral bacteria may be related to the onset of pustulosis palmaris et plantaris in some patients.

Am J Gastroenterol, 2000 Dec, 95(12), 3503 - 6
Eradication of small intestinal bacterial overgrowth reduces symptoms of irritable bowel syndrome; Pimentel M et al.; OBJECTIVES: Irritable bowel syndrome is the most common gastrointestinal diagnosis . The symptoms of irritable bowel syndrome are similar to those of small intestinal bacterial overgrowth . The purpose of this study was to test whether overgrowth is associated with irritable bowel syndrome and whether treatment of overgrowth reduces their intestinal complaints . METHODS: Two hundred two subjects in a prospective database of subjects referred from the community undergoing a lactulose hydrogen breath test for assessment of overgrowth were Rome I criteria positive for irritable bowel syndrome . They were treated with open label antibiotics after positive breath test . Subjects returning for follow-up breath test to confirm eradication of overgrowth were also assessed . Subjects with inflammatory bowel disease, abdominal surgery, or subjects demonstrating rapid transit were excluded . Baseline and after treatment symptoms were rated on visual analog scales for bloating, diarrhea, abdominal pain, defecation relief, mucous, sensation of incomplete evacuation, straining, and urgency . Subjects were blinded to their breath test results until completion of the questionnaire . RESULTS: Of 202 irritable bowel syndrome patients, 157 (78%) had overgrowth . Of these, 47 had follow-up testing . Twenty-five of 47 follow-up subjects had eradication of small intestinal bacterial overgrowth . Comparison of those that eradicated to those that failed to eradicate revealed an improvement in irritable bowel syndrome symptoms with diarrhea and abdominal pain being statistically significant after Bonferroni correction (p < 0.05) . Furthermore, 48% of eradicated subjects no longer met Rome criteria (chi2 = 12.0, p < 0.001) . No difference was seen if eradication was not successful . CONCLUSIONS: Small intestinal bacterial overgrowth is associated with irritable bowel syndrome . Eradication of the overgrowth eliminates irritable bowel syndrome by study criteria in 48% of subjects.

FEMS Microbiol Lett, 2001 Jan 1, 194(1), 111 - 9
Metabolic differences between attached and free-living marine bacteria: inadequacy of liquid cultures for describing in situ bacterial activity; Bonin P et al.; Marinobacter sp . strain CAB was cultivated with or without porous glass beads as solid support . Two substrates were used: the hydrophilic sodium lactate and a hydrophobic C(18)-isoprenoid ketone (6,10,14-trimethylpentadecan-2-one (TMP)) . The substrate adsorption onto the beads was measured . Bacterial adhesion was determined by a direct count technique and amounted to 70% of total cells . In the immobilised cell cultures (ICC), generation times were 1.5 and 1.8 times shorter than in the planktonic cultures (FCC) with sodium lactate and with TMP, respectively . In ICC, the growth yields were lower (15.3(FCC) x 10(9) and 0.8(ICC) x 10(9) bacteria mg(-1) of sodium lactate; 50(FCC) x 10(9) and 35(ICC) x 10(9) bacteria mg(-1) of TMP) . The mineralisation of substrates was estimated after mass spectrometric determination of the CO2 production rates of both free and immobilised cell cultures . The results indicated a higher specific CO2 production rate in the ICC with sodium lactate (3.1(FCC)+/-0.2 and 3.5(ICC)+/-0.3 nmol CO2 mg(-1) protein min(-1)) but not in the ICC with TMP (1.9(FCC)+/-0.7 and 0.5(ICC)+/-0.3 nmol CO2 mg(-1) protein min(-1)) . The affinities for the two substrates were lower in the presence of the solid support (K(m,ICC)=18.2+/-0.2 microM and 37.1+/-2.0 microM, for sodium lactate and TMP, respectively) than without support (K(m,FCC)=8.5+/-1.5 microM and 8.4+/-1.2 microM, for sodium lactate and TMP, respectively) . Moreover, the presence of a solid support showed a lower inhibition by the TMP (K(i,FCC)=3.8+/-1.0 microM and K(i,ICC)=12.2+/-2.5 microM) which may explain why the immobilised cell cultures degraded hydrophobic TMP more efficiently than the planktonic cultures.

Biochim Biophys Acta, 2000 Dec 29, 1543(2), 361 - 382
Bacterial 1,3-1,4-beta-glucanases: structure, function and protein engineering; Planas A; 1,3-1,4-beta-Glucanases (or lichenases, EC 3.2.1.73) hydrolyse linear beta-glucans containing beta-1,3 and beta-1,4 linkages such as cereal beta-glucans and lichenan, with a strict cleavage specificity for beta-1,4 glycosidic bonds on 3-O-substituted glucosyl residues . The bacterial enzymes are retaining glycosyl hydrolases of family 16 with a jellyroll beta-sandwich fold and a substrate binding cleft composed of six subsites . The present paper reviews the structure-function aspects of the enzymatic action including mechanistic enzymology, protein engineering and X-ray crystallographic studies.

Hepatogastroenterology, 2000 Nov-Dec, 47(36), 1649 - 53
Spontaneous bacterial peritonitis in Saudi Arabian patients with non-alcoholic liver cirrhosis; Ageely H et al.; BACKGROUND/AIMS: Spontaneous bacterial peritonitis is a frequent and serious complication of liver cirrhosis . Its prevalence varies from one survey to another . There are only very few reports of its occurrence among Arab patients . METHODOLOGY: We studied 115 Saudi Arabian patients with cirrhotic ascites in the Gizan region, an area of hyperendemic hepatitis B, over a 2-year period . RESULTS: Of these patients 12 (10.4%) had at least 1 episode of culture-positive spontaneous bacterial peritonitis (group A), an additional 34 (29.6%) had culture-negative neutrocytic ascites . The occurrence of spontaneous bacterial peritonitis was more frequent in males but was not influenced by the severity of liver disease or age . The overall mortality was 13.9%, however, only 1 patient died of spontaneous bacterial peritonitis-related cause . The remaining deaths were due to other complications of hepatic failure and portal hypertension . The low clinical threshold for treatment and the use of effective broad-spectrum antibiotics have reduced the mortality due to spontaneous bacterial peritonitis . There were a total of 56 recurrent episodes of infection in the patients . Of these episodes 46 occurred among 29 patients with spontaneous bacterial peritonitis and 10 among 62 patients with no infection during the index admissions . CONCLUSIONS: Prophylactic therapy against spontaneous bacterial peritonitis is a feasible strategy in reducing the frequency of recurrent peritonitis and should be recommended in these patients.

Wiad Lek, 2000, 53(9-10), 570 - 3
{Bacterial pericarditis in a patient with Sezary syndrome}; Hefczyc JZ et al.; Sezary Syndrome-T-cell cutaneous lymphoma is considered to be the leucaemic variant of mycosis fungoides . The case of 48 year old patient with phlegmonic pericarditis, due to immunodeficiency in the course of Sezary Syndrome is described in this paper . The patient was treated with vancomycine and three cycles of chemotherapy . The symptoms of disease were reduced.

Semin Immunol, 2000 Dec, 12(6), 527 - 35
CD1 molecules and CD1-dependent T cells in bacterial infections: a link from innate to acquired immunity?
Schaible UE, Kaufmann SH.
The MHC class I-like, non-polymorphic CD1 molecules represent a novel system for the presentation of glycolipid antigens to T lymphocytes . CD1-mediated T cell responses appear to play distinct roles during bacterial infections such as in tuberculosis . This review deals with two aspects of CD1-mediated immune reactions . First we discuss the role of group II CD1-dependent NK T cells in bacterial infection . Second, we provide an insight into differential intracellular meeting points for antigen processing between group I CD1 molecules, mycobacteria and mycobacterial glycolipid antigens .

J Immunol, 2001 Jan 15, 166(2), 1148 - 55
Bacterial invasion augments epithelial cytokine responses to Escherichia coli through a lipopolysaccharide-dependent mechanism; Schilling JD et al.; One mechanism of initiating innate host defenses against uropathogenic Escherichia coli (UPEC) is the production of cytokines by bladder epithelial cells; however, the means by which these cells recognize bacterial pathogens is poorly understood . Type 1 pili, expressed by the majority of UPEC, have been shown to have a critical role in inducing the expression of IL-6 in bladder epithelial cells after exposure to E . coli . In this study, we demonstrate that type 1 pili are not sufficient to activate IL-6 production by bladder epithelial cells . Instead, it was shown that bacterial invasion mediated by type 1 pili augments bladder epithelial responses to E . coli via an LPS-dependent mechanism, leading to the production of IL-6 . RNA transcripts for the LPSR Toll-like receptor 4 (TLR4) was detected in cultured bladder epithelial cells . The in vivo role of TLR4 was assessed using C3H/HeJ mice, which express a dominant negative form of TLR4 . After infection with UPEC, C3H/HeJ mice have large foci of intracellular bacteria that persist within the bladder epithelium in the absence of any notable inflammatory response . These results indicate that LPS is required for bacterial invasion to enhance host responses to E . coli within the bladder.

Am J Perinatol, 2000, 17(6), 315 - 8
Gram-stain diagnosis of bacterial vaginosis after rupture of membranes; Core La BQ et al.; Correlation of the Gram stains prior to and after rupture of the membranes (ROM) and the efficacy of Gram-stain diagnosis for bacterial vaginosis (BV) after membrane rupture has not previously been evaluated . From April 1997 to May 1998, women presenting in labor or for labor induction were invited to participate . Women with ROM prior to hospital presentation or contraindications for vaginal delivery were excluded . A Gram stain of vaginal secretions was obtained prior to and approximately 2 hours after ROM . BV was diagnosed with a Nugent score of 7-10 . The Gram stain prior to and after membrane rupture was evaluated using the Spearman correlation coefficient . Population characteristics (n = 91) included maternal age of 15 to 38 years, 45 (49%) nulliparas, 59 (65%) African-Americans, 23 (25%) Hispanics, 7 (8%) Caucasian, and 2 (2%) Asians . Gestational age upon study entry was 35 to 44 weeks . Twenty-one percent (19 of 91) of the study population was diagnosed with BV . Using the Gram stain prior to membrane rupture as the standard, the efficacy of the Gram stain after ROM had a sensitivity of 26%, specificity of 97%, positive predictive value of 71%, and negative predicative value of 83% . Good correlation of pre- and post-membrane rupture Gram stains was demonstrated (r = 0.69) . Gram stain may be useful for ruling out BV in the presence of membrane rupture, but is not useful for diagnosing BV due to poor sensitivity.

Int J Immunopharmacol, 2000 Dec, 22(12), 1093 - 102
Immunostimulation by bacterial components: I . Activation Of macrophages and enhancement of genetic immunization by the lipopeptide P3CSK4; vd Esche U et al.; Synthetic lipopeptides derived from the N-terminus of bacterial lipoprotein constitute potent macrophage activators and polyclonal B-lymphocyte stimulators . They are also efficient immunoadjuvants in parenteral, oral and nasal immunization either in combination with or after covalent linkage to an antigen . Here we show how alterations in the molecular structure influence their biological properties indicating P3CSK4 as one of the most active members of a lipopentapeptide fatty acid library . This compound resulted in a most pronounced macrophage stimulation as indicated by NO release, activation of NFkappaB translocation, and enhancement of tyrosine protein phosphorylation . Furthermore, P3CSK4 activates/represses an array of at least 140 genes partly involved in signal transduction and regulation of the immune response . Finally we have evidence that P3CSK4 constitutes an effective adjuvant for DNA immunizations, especially increasing weak humoral immune responses . Our findings are of importance for further optimizing both conventional and genetic immunization, and for the development of novel synthetic vaccines.

Microbes Infect, 2000 Nov, 2(14), 1705 - 19
Hijacking of apoptotic pathwaysby bacterial pathogens; Gao L et al.; Increasing evidence indicates that apoptosis of the host cell may constitute a defense mechanism to confine the infection by bacterial pathogens . Certain pathogens have developed elegant mechanisms to modulate the fate of the host cell, which include induction or blockage of apoptosis . These studies will promote our understanding of the pathogenesis of infectious diseases and aid the development of means for therapeutic intervention.

Genesis, 2001 Jan, 29(1), 14 - 21
Rapid engineering of bacterial artificial chromosomes using oligonucleotides; Swaminathan S et al.; A rapid method obviating the use of selectable markers to genetically manipulate large DNA inserts cloned into bacterial artificial chromosomes is described . Mutations such as single-base changes, deletions, and insertions can be recombined into a BAC by using synthetic single-stranded oligonucleotides as targeting vectors . The oligonucleotides include the mutated sequence flanked by short homology arms of 35-70 bases on either side that recombine with the BAC . In the absence of any selectable marker, modified BACs are identified by specific PCR amplification of the mutated BAC from cultures of pooled bacterial cells . Each pool represents about 10 electroporated cells from the original recombination mixture . Subsequently, individual clones containing the desired alteration are identified from the positive pools . Using this BAC modification method, we have observed a frequency of one recombinant clone per 90-260 electroporated cells . The combination of high targeting frequency and the sensitive yet selective PCR-based screening method makes BAC manipulation using oligonucleotides both rapid and simple.

Biotechnol Bioeng, 2001 Feb 5, 72(3), 255 - 60
Variable specific activity of Escherichia coli beta-galactosidase in bacterial cells; Cazorla D et al.; Escherichia coli lacZ is a frequently employed reporter gene for the monitoring of gene expression and recombinant protein production due the simple determination of beta-galactosidase activity in both qualitative and quantitative assays . In the absence of either total or recombinant protein synthesis, we observed a lack of correlation between protein amount and enzymatic activity in both engineered and native beta-galactosidases in Escherichia coli cells . A delayed fading of beta-galactosidase activity compared with the rapid degradation of intact protein suggests a progressive increase in enzyme-specific activity during the life of the protein . This intriguing event does not involve solubilization from major protein aggregates and it occurs both in vivo and in cell extracts, but not in solutions of purified protein . Possible explanations for this activation are examined in the context of the assisted protein folding network and proteolytic degradation of misfolded proteins .

Blood, 2001 Jan 1, 97(1), 235 - 41
The interaction of human peripheral blood eosinophils with bacterial lipopolysaccharide is CD14 dependent; Plotz SG et al.; Bacterial lipopolysaccharide (LPS, endotoxin) is a ubiquitous component of dust and air pollution and is suspected to contribute after inhalation to an activation of eosinophils in bronchial tissues of asthmatic patients, provoking inflammatory and allergic processes . We were therefore interested in the interaction of eosinophil granulocytes with LPS and have examined the activation of and uptake to human peripheral blood eosinophils by LPS . Eosinophils were stimulated by LPS and the endotoxic component lipid A and the release of tumor necrosis factor alpha (TNF-alpha) and of the eosinophil-specific granule protein eosinophil cationic protein (ECP) was estimated . The results show induction of TNF-alpha and ECP-release by LPS and lipid A in a dose-dependent manner . Anti-CD14 monoclonal antibody (moAb) (clone MEM-18) and the synthetic lipid A partial structure 406 blocked the release of TNF-alpha and ECP by LPS-stimulated eosinophils . Studies with radioactively labeled LPS showed dose-dependent uptake of (3)H-LPS to eosinophils . The (3)H-LPS uptake was found to be specific because preincubation with unlabeled LPS, compound 406 and also anti-CD14 antibodies inhibited uptake of (3)H-LPS to eosinophil granulocytes . By flow cytometry using anti-CD14 moAb and by reverse transcriptase-polymerase chain reaction (RT-PCR) technique, CD14 expression was detectable . Furthermore, messenger RNA (mRNA) expression of Toll-like receptors (TLR) 2 and TLR 4 was detected, indicating the presence of these CD14 coreceptors . The results indicate that eosinophils can take up LPS and can be stimulated by LPS in a CD14-dependent manner . Hence, in addition to allergens, eosinophils interact with endotoxin, a process that possibly exacerbates ongoing inflammatory and allergic processes.

Appl Environ Microbiol, 2001 Jan, 67(1), 481 - 3
Effect of introns and AT-rich sequences on expression of the bacterial hygromycin B resistance gene in the basidiomycete Schizophyllum commune; Scholtmeijer K et al.; Previously, it was shown that introns are required for efficient mRNA accumulation in Schizophyllum commune and that the presence of AT-rich sequences in the coding region of genes can result in truncation of transcripts in this homobasidiomycete . Here we show that intron-dependent mRNA accumulation and truncation of transcripts are two independent events that both affect expression of the bacterial hygromycin B resistance gene in S . commune.

Appl Environ Microbiol, 2001 Jan, 67(1), 464 - 8
Surface interactions between Escherichia coli and hemocytes of the Mediterranean mussel Mytilus galloprovincialis lam . leading to efficient bacterial clearance; Canesi L et al.; The role of type 1 fimbriae in the interactions between Escherichia coli and Mytilus galloprovincialis Lam . hemocytes was evaluated . The association of fimbriated strain MG155 with hemocyte monolayers at 18 degrees C was 1.5- and 3- to 4-fold greater than the association of unfimbriated mutant AAEC072 in artificial seawater and in hemolymph serum, respectively . Such differences were apparently due to different adhesive properties since MG155 adhered more efficiently than AAEC072 when hemocytes were incubated at 4 degrees C to inhibit the internalization process . Hemolymph serum increased both association and adherence of MG155 two- to threefold but did not affect association and adherence of AAEC072 . MG155 was also 1.5- to 1.7-fold more sensitive to killing by hemocytes than AAEC072, as evaluated by the number of culturable bacteria after 60 and 120 min of incubation . The role of type 1 fimbriae in MG155 interactions with hemocytes was confirmed by the inhibitory effect of D-mannose . In in vivo experiments MG155 cells were cleared from circulating hemolymph more rapidly than AAEC072 cells were cleared . These results confirm that surface properties are crucial in influencing bacterial persistence and survival within mussel hemolymph.

Proc Natl Acad Sci U S A, 2001 Jan 16, 98(2), 497 - 501 Epub 2001 Jan 02.
Cleavage of cyclin A at R70/R71 by the bacterial protease OmpT; Yam CH et al.; Previous work has shown that cyclin A can be cleaved at Arg-70/Arg-71 by a proteolytic activity present in an in vitro-coupled transcription/translation system by using rabbit reticulocyte lysate programmed by plasmid DNA encoding p27(KIP1), a cyclin-dependent kinase inhibitor, but not by plasmid DNAs encoding other cyclin-dependent kinases inhibitors . Here we report that cyclin A is also cleaved by translation product programmed by plasmid DNA encoding cyclin B . Several findings indicate that the cleavage activity in this assay is provided by the bacterial protease OmpT, which cofractionates with cyclin B and p27(KIP1) plasmid DNAs and is thus carried over into the coupled in vitro transcription/translation reactions . (i) Cleavage activity appeared even when transcription or translation of the cyclin B or p27(KIP1) was blocked . (ii) Activity resembling OmpT, a serine protease that cleaves between dibasic residues, routinely copurifies with p27(KIP1) and cyclin B plasmid DNAs . (iii) Both cyclin A cleavage activity and OmpT activity are heat stable, resistant to denaturation, and inhibited by Zn(2+), Cu(2+), or benzamidine . (iv) Cyclin A cleavage activity is detected when using lysates or DNAs prepared from Escherichia coli strains that contained OmpT but not with strains lacking OmpT . (v) Purified OmpT enzyme itself cleaves cyclin A at R70/R71 . These data indicate that OmpT can be present in certain DNA preparations obtained by using standard plasmid purification protocols, and its presence can potentially affect the outcome and interpretation of studies carried out using in vitro-translated proteins.

J Biol Chem, 2001 Mar 30, 276(13), 10330 - 7 Epub 2000 Dec 21.
Crystallographic evidence for substrate-assisted catalysis in a bacterial beta-hexosaminidase; Mark BL et al.; beta-Hexosaminidase, a family 20 glycosyl hydrolase, catalyzes the removal of beta-1,4-linked N-acetylhexosamine residues from oligosaccharides and their conjugates . Heritable deficiency of this enzyme results in various forms of GalNAc-beta(1,4)-{N-acetylneuraminic acid (2,3)}-Gal-beta(1,4)-Glc-ceramide gangliosidosis, including Tay-Sachs disease . We have determined the x-ray crystal structure of a beta-hexosaminidase from Streptomyces plicatus to 2.2 A resolution (Protein Data Bank code ) . beta-Hexosaminidases are believed to use a substrate-assisted catalytic mechanism that generates a cyclic oxazolinium ion intermediate . We have solved and refined a complex between the cyclic intermediate analogue N-acetylglucosamine-thiazoline and beta-hexosaminidase from S . plicatus to 2.1 A resolution (Protein Data Bank code ) . Difference Fourier analysis revealed the pyranose ring of N-acetylglucosamine-thiazoline bound in the enzyme active site with a conformation close to that of a (4)C(1) chair . A tryptophan-lined hydrophobic pocket envelopes the thiazoline ring, protecting it from solvolysis at the iminium ion carbon . Within this pocket, Tyr(393) and Asp(313) appear important for positioning the 2-acetamido group of the substrate for nucleophilic attack at the anomeric center and for dispersing the positive charge distributed into the oxazolinium ring upon cyclization . This complex provides decisive structural evidence for substrate-assisted catalysis and the formation of a covalent, cyclic intermediate in family 20 beta-hexosaminidases.

Bioorg Med Chem Lett, 2000 Dec 18, 10(24), 2811 - 3
Synthesis and biological evaluation of analogues of bacterial lipid I; Silva DJ et al.; Bacterial Lipid I analogues containing different anomeric groups at the muramic acid moiety were synthesized and screened in MurG enzyme assays run in the presence and absence of cell wall membranes . The results obtained in this study help elucidate the role of the lipid diphosphate in the recognition of Lipid I by MurG.

Crit Rev Oral Biol Med, 2000, 11(4), 467 - 80
Evidence for bacterial causation of adverse pulpal responses in resin-based dental restorations; Bergenholtz G; The widespread use of resin and resin-monomers for bonding of dental restorations to dentin has occurred because of a fundamental shift in the view that injury to the pulp is induced by restorative procedures . While, for many years, the toxic effects of restorative materials were thought to be of crucial importance in the development of adverse pulpal responses, the key role of bacterial leakage at the restoration-tooth interface is now well-recognized . Consequently, if optimal conditions for the preservation of pulpal health are to be ensured, dental restorations should provide an impervious seal against the surrounding tooth structure . However, polymerization shrinkage and contraction stresses induced during setting, as well as a variety of technical difficulties encountered during the clinical operation, often produce less than perfect results . Therefore, modern restorative procedures involving resin and resin-bonded restoratives must still rely on the ability of the pulp to cope with the injurious elements to which it may be exposed during and after the procedure . This review examines factors that may govern the pulp's response to restorative procedures that involve adhesive technologies . An assessment is made of the risks involved as far as the continued vital function of the pulp is concerned . It is concluded that an intact, although thin, wall of primary dentin often enables the pulp to overcome both toxic material effects and the influences of bacterial leakage . In contrast, the pulp may not do equally well following capping of open exposures with resin composites . A dearth of controlled clinical studies in this area of dentistry calls for confirmation that pulpal health prevails over the long term following the use of total-etch and resin-bonding techniques.

J Inorg Biochem, 2000 Nov, 82(1-4), 19 - 25
Probing molecular structure of dioxygen reduction site of bacterial quinol oxidases through ligand binding to the redox metal centers; Tsubaki M et al.; Cytochromes bo and bd are structurally unrelated terminal ubiquinol oxidases in the aerobic respiratory chain of Escherichia coli . The high-spin heme o-CuB binuclear center serves as the dioxygen reduction site for cytochrome bo, and the heme b595-heme d binuclear center for cytochrome bd . CuB coordinates three histidine ligands and serves as a transient ligand binding site en route to high-spin heme o one-electron donor to the oxy intermediate, and a binding site for bridging ligands like cyanide . In addition, it can protect the dioxygen reduction site through binding of a peroxide ion in the resting state, and connects directly or indirectly Tyr288 and Glu286 to carry out redox-driven proton pumping in the catalytic cycle . Contrary, heme b595 of cytochrome bd participate a similar role to CuB in ligand binding and dioxygen reduction but cannot perform such versatile roles because of its rigid structure.

Ethiop Med J, 2000 Jul, 38(3), 165 - 74
Prevalence of bacterial pathogens in children with acute respiratory infection in Addis Ababa; Mohammed E et al.; A study was conducted in the Ethio-Swedish Children's Hospital and different schools and kindergartens in Addis Ababa to determine the prevalence of bacterial agents that are associated with acute respiratory infection in children from 1998-1999 . A total of 883 subjects were studied, out of which 77% were cases from the Ethio-Swedish Children's Hospital and 23% were controls from different schools and kindergartens . From each case and control throat and nasopharyngeal specimens were collected . Culture and different biochemical tests were used to isolate the potential bacterial pathogens . Clinical findings like cough, difficult breathing and fever were correlated with laboratory findings . S . pneumoniae and H . influenzae type b were the most commonly isolated bacteria in both throat and nasopharyngeal specimens; 74% and 70% in the cases and 2% and 5% in the control groups, respectively . This paper discusses the association between throat and nasopharyngeal carriership of bacteria and acute respiratory infection in children in Addis Ababa.

J Biol Chem, 2001 Mar 23, 276(12), 9478 - 85 Epub 2000 Dec 19.
A 72-base pair AT-rich DNA sequence element functions as a bacterial gene silencer; Chen CC et al.; We have previously demonstrated that sequential activation of the bacterial ilvIH-leuO-leuABCD gene cluster involves a promoter-relay mechanism . In the current study, we show that the final activation of the leuABCD operon is through a transcriptional derepression mechanism . The leuABCD operon is transcriptionally repressed by the presence of a 318-base pair AT-rich upstream element . LeuO is required for derepressing the repressed leuABCD operon . Deletion analysis of the repressive effect of the 318-bp element has led to the identification of a 72-bp AT-rich (78% A+T) DNA sequence element, AT4, which is capable of silencing a number of unrelated promoters in addition to the leuABCD promoter . AT4-mediated gene silencing is orientation-independent and occurs within a distance of 300 base pairs . Furthermore, an increased gene-silencing effect was observed with a tandemly repeated AT4 dimer . The possible mechanism of AT4-mediated gene silencing in bacteria is discussed.

J Biol Chem, 2001 Mar 30, 276(13), 9613 - 9 Epub 2000 Dec 19.
Equilibrium binding of single-stranded DNA to the secondary DNA binding site of the bacterial recombinase RecA; Gourves AS et al.; The bacterial recombinase RecA forms a nucleoprotein filament in vitro with single-stranded DNA (ssDNA) at its primary DNA binding site, site I . This filament has a second site, site II, which binds ssDNA and double-stranded DNA . We have investigated the binding of ssDNA to the RecA protein in the presence of adenosine 5'-O-(thiotriphosphate) cofactor using fluorescence anisotropy . The RecA protein carried out DNA strand exchange with a 5'-fluorescein-labeled 32-mer oligonucleotide . The anisotropy signal was shown to measure oligonucleotide binding to RecA, and the relationship between signal and binding density was determined . Binding of ssDNA to site I of RecA was stable at high NaCl concentrations . Binding to site II could be described by a simple two-state equilibrium, K = 4.5 +/- 1.5 x 10(5) m(-1) (37 degrees C, 150 mm NaCl, pH 7.4) . The reaction was enthalpy-driven and entropy-opposed . It depended on salt concentration and was sensitive to the type of monovalent anion, suggesting that anion-dependent protein conformations contribute to ssDNA binding at site II.

J Biol Chem, 2001 Apr 6, 276(14), 11055 - 61 Epub 2000 Dec 18.
Fructose-6-phosphate aldolase is a novel class I aldolase from Escherichia coli and is related to a novel group of bacterial transaldolases; Schurmann M et al.; We have cloned an open reading frame from the Escherichia coli K-12 chromosome that had been assumed earlier to be a transaldolase or a transaldolase-related protein, termed MipB . Here we show that instead a novel enzyme activity, fructose-6-phosphate aldolase, is encoded by this open reading frame, which is the first report of an enzyme that catalyzes an aldol cleavage of fructose 6-phosphate from any organism . We propose the name FSA (for fructose-six phosphate aldolase; gene name fsa) . The recombinant protein was purified to apparent homogeneity by anion exchange and gel permeation chromatography with a yield of 40 mg of protein from 1 liter of culture . By using electrospray tandem mass spectroscopy, a molecular weight of 22,998 per subunit was determined . From gel filtration a size of 257,000 (+/- 20,000) was calculated . The enzyme most likely forms either a decamer or dodecamer of identical subunits . The purified enzyme displayed a V(max) of 7 units mg(-)1 of protein for fructose 6-phosphate cleavage (at 30 degrees C, pH 8.5 in 50 mm glycylglycine buffer) . For the aldolization reaction a V(max) of 45 units mg(-)1 of protein was found; K(m) values for the substrates were 9 mm for fructose 6-phosphate, 35 mm for dihydroxyacetone, and 0.8 mm for glyceraldehyde 3-phosphate . FSA did not utilize fructose, fructose 1-phosphate, fructose 1,6-bisphosphate, or dihydroxyacetone phosphate . FSA is not inhibited by EDTA which points to a metal-independent mode of action . The lysine 85 residue is essential for its action as its exchange to arginine (K85R) resulted in complete loss of activity in line with the assumption that the reaction mechanism involves a Schiff base formation through this lysine residue (class I aldolase) . Another fsa-related gene, talC of Escherichia coli, was shown to also encode fructose-6-phosphate aldolase activity and not a transaldolase as proposed earlier.

Nature, 2000 Dec 7, 408(6813), 740 - 5
A Toll-like receptor recognizes bacterial DNA; Hemmi H et al.; DNA from bacteria has stimulatory effects on mammalian immune cells, which depend on the presence of unmethylated CpG dinucleotides in the bacterial DNA . In contrast, mammalian DNA has a low frequency of CpG dinucleotides, and these are mostly methylated; therefore, mammalian DNA does not have immuno-stimulatory activity . CpG DNA induces a strong T-helper-1-like inflammatory response . Accumulating evidence has revealed the therapeutic potential of CpG DNA as adjuvants for vaccination strategies for cancer, allergy and infectious diseases . Despite its promising clinical use, the molecular mechanism by which CpG DNA activates immune cells remains unclear . Here we show that cellular response to CpG DNA is mediated by a Toll-like receptor, TLR9 . TLR9-deficient (TLR9-/-) mice did not show any response to CpG DNA, including proliferation of splenocytes, inflammatory cytokine production from macrophages and maturation of dendritic cells . TLR9-/- mice showed resistance to the lethal effect of CpG DNA without any elevation of serum pro-inflammatory cytokine levels . The in vivo CpG-DNA-mediated T-helper type-1 response was also abolished in TLR9-/- mice . Thus, vertebrate immune systems appear to have evolved a specific Toll-like receptor that distinguishes bacterial DNA from self-DNA.

J Leukoc Biol, 2000 Dec, 68(6), 909 - 15
TAK1 regulates multiple protein kinase cascades activated by bacterial lipopolysaccharide; Lee J et al.; During inflammation the balance between cell activation and cell death is determined by the tight regulation of multiple intracellular enzyme cascades . Key regulatory steps often involve protein kinases . We show that the prototypical pro-inflammatory molecule, bacterial lipopolysaccharide, activates multiple protein kinases such as p38, JNK, IKK-beta, and PKB/Akt via transforming growth factor beta-activated kinase-1 (TAK1) . We also show that TAK1 plays an important role in similar activation pathways triggered by interleukin-1 . Thus TAK1 must be considered as an important component of intracellular pathways in cells involved in host responses to physiological and/or environmental stress signals during inflammation.

Philos Trans R Soc Lond B Biol Sci, 2000 Oct 29, 355(1402), 1345 - 9
How carotenoids protect bacterial photosynthesis; Cogdell RJ et al.; The essential function of carotenoids in photosynthesis is to act as photoprotective agents, preventing chlorophylls and bacteriochlorophylls from sensitizing harmful photodestructive reactions in the presence of oxygen . Based upon recent structural studies on reaction centres and antenna complexes from purple photosynthetic bacteria, the detailed organization of the carotenoids is described . Then with specific reference to bacterial antenna complexes the details of the photoprotective role, triplet triplet energy transfer, are presented.

J Reprod Med, 2000 Nov, 45(11), 889 - 96
Metronidazole for bacterial vaginosis . A comparison of vaginal gel vs . oral therapy; Hanson JM et al.; OBJECTIVE: To compare the efficacy and safety of 0.75% metronidazole vaginal gel with oral metronidazole for the treatment of bacterial vaginosis (BV) . STUDY DESIGN: Nonpregnant women with BV were enrolled in a multicenter, randomized, investigator-blind treatment trial . Patients were randomly assigned to either 0.75% metronidazole vaginal gel (5 g twice daily for five days) or oral metronidazole (500 mg twice daily for seven days) . Follow-up visits occurred approximately two and five weeks after initiation of therapy . RESULTS: BV was clinically eliminated at the first follow-up visit in 83.7% (36/43, 95% CI 72.3-95.1%) of the intravaginal group and 85.1% (40/47, 95% CI 74.6-95.6%) of the oral group . At the final visit, BV was eliminated in 70.7% (29/41, 95% CI 56.3-85.1%) of the intravaginal group and 71.1% (32/45, 95% CI 57.4-84.8%) of the oral group . Significantly more patients in the oral treatment group (51.8%) reported gastrointestinal complaints as compared to the intravaginal treatment group (32.7%, P = .04) . CONCLUSION: The efficacy of 0.75% metronidazole vaginal gel twice daily for five days in treating BV was similar to that of standard oral metronidazole treatment and was associated with fewer gastrointestinal complaints.

Hepatology, 2001 Jan, 33(1), 66 - 73
Bacterial lipopolysaccharide enhances aflatoxin B1 hepatotoxicity in rats by a mechanism that depends on tumor necrosis factor alpha; Barton CC et al.; Exposure to a nontoxic dose of bacterial endotoxin (lipopolysaccharide {LPS}) potentiates the hepatotoxicity of aflatoxin B(1) (AFB(1)) . Because some of the pathophysiologic effects associated with LPS are mediated through tumor necrosis factor alpha (TNF-alpha), this study was conducted to explore the role of TNF-alpha in the AFB(1)/LPS model . Male Sprague-Dawley rats (250-300 g) were treated with either 1 mg AFB(1)/kg, intraperitoneally, or its vehicle (0.5% dimethyl sulfoxide {DMSO}/water), and 4 hours later with either Escherichia coli lipopolysaccharide (7.4 x 10(6)EU/kg, intravenously) or its saline vehicle . LPS administration resulted in a marked rise in TNF-alpha levels at 6 hours, which preceded the onset of liver injury . TNF-alpha messenger RNA (mRNA) in liver was increased by LPS treatment . The mRNA of receptors (R1 and R2) for TNF-alpha was also examined . R1 mRNA levels were not altered; however, R2 mRNA levels were increased by either AFB(1) or LPS administration . To determine if TNF-alpha plays a causal role in the development of liver injury, the increase in TNF-alpha was attenuated by administration of either pentoxifylline or anti-TNF-alpha serum, and liver injury was assessed . Administration of either of these agents resulted in protection . LPS treatment resulted in the upregulation of gene transcription for cyclooxygenase-2 (COX-2) . However, administration of the selective COX-2 inhibitor NS-398 did not decrease injury . TNF-alpha and COX-2 inhibitors did not affect hepatic sequestration of neutrophils . Furthermore, it did not appear that TNF-alpha contributed to injury through inhibition of tissue repair . These data support the hypothesis that LPS-induced expression of TNF-alpha underlies the potentiation of AFB(1)-induced hepatotoxicity.

J Mol Biol, 2000 Dec 15, 304(5), 687 - 98
RNA polymerase: structural similarities between bacterial RNA polymerase and eukaryotic RNA polymerase II; Ebright RH; Bacterial RNA polymerase and eukaryotic RNA polymerase II exhibit striking structural similarities, including similarities in overall structure, relative positions of subunits, relative positions of functional determinants, and structures and folding topologies of subunits . These structural similarities are paralleled by similarities in mechanisms of interaction with DNA .

Biochemistry, 2000 Dec 26, 39(51), 16252 - 62
Interactions between the donor and acceptor sides in bacterial reaction centers; Ginet N et al.; The apparent equilibrium constant K'(2) for electron transfer between the primary (Q(A)) and secondary (Q(B)) quinone acceptors of the reaction center was measured in chromatophores of Rhodobacter capsulatus . In the presence of the oxidized primary donor P(+), we obtained a value of K'(2)(P(+)) approximately 100 at pH 7.2, based on the rates of recombination from P(+)Q(A-) and P(+)Q(B-) . K'(2) was also measured in the presence of reduced P, from the damping of semiquinone oscillations during a series of single turnover flashes . A 5-fold smaller value, K'(2)(P) approximately 20, was found . Additional information on the interactions between the donor and acceptor sides was obtained by measuring the shift of the midpoint potential of P caused by the presence of Q(B-) or Q(A-)S (where S indicates the presence of the inhibitor stigmatellin) . A stabilization of the oxidized state P(+) was observed in both instances, by 10 mV for Q(B-) and 30 mV for Q(A-)S . The larger stabilization of P(+)Q(A-)S with respect to P(+)Q(B-) does not account for the effect of P(+)/P on K'(2) . Analysis of these results indicates that the interactions between P(+)/P and Q(A)/Q(A)(-) are markedly modified depending on the occupancy of the Q(B) pocket by ubiquinone or by stigmatellin . We propose that the large value of K'(2)(P(+)) results essentially from a conformational destabilization of the P(+)Q(A-) state, that is relieved when the proximal site of the Q(B) pocket is occupied by stigmatellin.

Biochemistry, 2000 Dec 26, 39(51), 16185 - 9
Modeling the bacterial photosynthetic reaction center 3: interpretation of effects of site-directed mutagenesis on the special-pair midpoint potential; Reimers JR et al.; Interpretation of changes in midpoint potential of the "special pair" in bacterial photosynthetic reaction centers caused by site-directed mutagenesis is discussed in terms of a simple tight-binding model which relates them to concomitant variations in spin distribution between the two bacteriochlorophyll molecules of the special pair . Our analysis improves on previous similar ones by Allen and co-workers {Artz, K., Williams, J . C., Allen, J . P., Lendzian, F., Rautter, J., and Lubitz, W . (1997) Proc . Natl . Acad . Sci . U.S.A . 94, 13582; Ivancich, A., Artz, K., Williams, J . C., Allen, J . P., and Mattioli, T . A . (1998) Biochemistry 37, 11812} in that it is both more complete, including electron-phonon coupling, and more accurate . It is applied to analyze data for a series of M160 mutants of Rhodobacter sphaeroides, yielding a value of 0.18+/-0.03 eV for the electronic coupling energy between the highest occupied levels of the two bacteriochlorophylls in the wild-type and a value of the energy offset E(o) between the highest occupied molecular orbitals of the L and M bacteriochlorophylls of 0.14+/-0.03 eV . For a mutant in which the electron hole in the special pair cation is located entirely on the reactive (L) side, a potential of 641+/-30 mV with respect to the normal hydrogen electrode is predicted . This agrees well with the average value ca . 650 mV observed for the heterodimer mutant HL(M202) in which the bacteriochlorophyll on the unreactive M side has been replaced by a bacteriopheophytin, causing extensive charge localization . However, the deduced coupling is found to be very sensitive to small changes in the assumptions used in the model, and various important chemical effects remain to be included.

Mol Microbiol, 2000 Dec, 38(5), 927 - 39
New prospects in studying the bacterial signal recognition particle pathway; Herskovits AA et al.; In vivo and in vitro studies have suggested that the bacterial version of the mammalian signal recognition particle (SRP) system plays an essential and selective role in protein biogenesis . The bacterial SRP system consists of at least two proteins and an RNA molecule (termed Ffh, FtsY and 4.5S RNA, respectively, in Escherichia coli) . Recent evidence suggests that other putative bacterial-specific SRP components may also exist . In vitro experiments confirmed the expected basic features of the bacterial SRP system by demonstrating interactions among the SRP components themselves, between them and ribosomes, ribosome-linked hydrophobic nascent polypeptides or inner membranes . The availability of a conserved (and essential) bacterial SRP version has facilitated the implementation of powerful genetic and biochemical approaches for studying the cascade of events during the SRP-mediated targeting process in vivo and in vitro as well as the three-dimensional structures and the properties of each SRP component and complex.

Trends Microbiol, 2000 Nov, 8(11), 521 - 6
Transposon-based approaches to identify essential bacterial genes; Judson N et al.; Transposons are a powerful tool for identifying genes essential for bacterial viability . The availability of many bacterial genome sequences and the large number of genes of unknown function therein have inspired the generation of a variety of different approaches . These methods are described and their advantages and disadvantages are discussed.

Am J Obstet Gynecol, 2000 Dec, 183(6), 1434 - 9
Asymptomatic bacterial vaginosis: response to therapy; Schwebke JR; OBJECTIVE: Bacterial vaginosis causes symptomatic vaginal discharge and has been associated with preterm birth and with the acquisition of human immunodeficiency virus . Half of all women with bacterial vaginosis are free of symptoms, and treatment of these women is controversial . The objective of this study was to determine the extent of poor symptom recognition in this group of women . STUDY DESIGN: Seventy-five women attending a sexually transmitted disease clinic who had asymptomatic bacterial vaginosis were entered into a randomized, double-blind, placebo-controlled trial comparing metronidazole gel with placebo . Subjects' perceptions about changes in vaginal discharge and odor were determined, and treatment and placebo groups were compared by means of standard statistical analysis . RESULTS: When subjects were stratified by treatment group, there were no differences in their retrospective assessments of vaginal discharge and odor . A subset of women who had normalization of clinical parameters or of Gram stain scores did admit retrospectively to improvement; however, the difference between this group and the group without normalization was not statistically significant . Twenty-one percent of treated women subsequently had vaginal candidiasis . CONCLUSIONS: A greater percentage of women with resolution of bacterial vaginosis did retrospectively notice improvement in vaginal discharge and odor in comparison with those women without resolution; however, this was not statistically significant . These findings do not support routine treatment of women with asymptomatic bacterial vaginosis.

Infect Immun, 2001 Jan, 69(1), 472 - 8
Actinobacillus pleuropneumoniae iron transport and urease activity: effects on bacterial virulence and host immune response; Baltes N et al.; Actinobacillus pleuropneumoniae, a porcine respiratory tract pathogen, has been shown to express transferrin-binding proteins and urease during infection . Both activities have been associated with virulence; however, their functional role for infection has not yet been elucidated . We used two isogenic A . pleuropneumoniae single mutants (DeltaexbB and DeltaureC) and a newly constructed A . pleuropneumoniae double (DeltaureC DeltaexbB) mutant in aerosol infection experiments . Neither the A . pleuropneumoniae DeltaexbB mutant nor the double DeltaureC DeltaexbB mutant was able to colonize sufficiently long to initiate a detectable humoral immune response . These results imply that the ability to utilize transferrin-bound iron is required for multiplication and persistence of A . pleuropneumoniae in the porcine respiratory tract . The A . pleuropneumoniae DeltaureC mutant and the parent strain both caused infections that were indistinguishable from one another in the acute phase of disease; however, 3 weeks postinfection the A . pleuropneumoniae DeltaureC mutant, in contrast to the parent strain, could not be isolated from healthy lung tissue . In addition, the local immune response-as assessed by fluorescence-activated cell sorter and enzyme-linked immunosorbent spot analyses-revealed a significantly higher number of A . pleuropneumoniae-specific B cells in the bronchoalveolar lavage fluid (BALF) of pigs infected with the A . pleuropneumoniae DeltaureC mutant than in the BALF of those infected with the parent strain . These results imply that A . pleuropneumoniae urease activity may cause sufficient impairment of the local immune response to slightly improve the persistence of the urease-positive A . pleuropneumoniae parent strain.

Heart, 2001 Jan, 85(1), 11 - 2
Bacterial endocarditis following repeated tattooing; Satchithananda DK et al.; Body decoration in the form of tattooing is becoming increasingly popular, especially among younger age groups . Although serious infections following tattooing are rare they are well documented . The first reported case of endocarditis caused by repeated tattooing in an individual with known valvar heart disease is presented.

J Microbiol Methods, 2001 Jan, 43(3), 223 - 32
Infection of Vero cells with Coxiella burnetii phase II: relative intracellular bacterial load and distribution estimated by confocal laser scanning microscopy and morphometry; Zamboni DS et al.; Coxiella burnetii, the agent of Q fever in man and of coxiellosis in other species, is an intracellular pathogen not yet grown axenically . Confocal laser fluorescence microscopy and morphometry were used to measure relative C . burnetii phase II loads and their intracellular distribution in aldehyde fixed and DAPI stained Vero cell monolayers . The fluorescence of single horizontal optical sections provided useful information on relative loads of bacteria in cells and vacuoles . The relative density of the bacteria in the vacuoles was inferred from ratios of fluorescence to vacuolar section areas . Relative bacterial loads, bacterial densities and section areas of large vacuoles increased exponentially between days 2 and 4 of the infection of gamma-irradiated host cells, stabilized between days 4 and 6, and decreased thereafter . Estimated minimum doubling times were higher for the overall complement of the intracellular organisms (about 12 h) than for bacteria that were confined to larger vacuoles (about 10 h).

Science, 2000 Dec 15, 290(5499), 2148 - 52
The bacterial flagellar cap as the rotary promoter of flagellin self-assembly; Yonekura K et al.; The growth of the bacterial flagellar filament occurs at its distal end by self-assembly of flagellin transported from the cytoplasm through the narrow central channel . The cap at the growing end is essential for its growth, remaining stably attached while permitting the flagellin insertion . In order to understand the assembly mechanism, we used electron microscopy to study the structures of the cap-filament complex and isolated cap dimer . Five leg-like anchor domains of the pentameric cap flexibly adjusted their conformations to keep just one flagellin binding site open, indicating a cap rotation mechanism to promote the flagellin self-assembly . This represents one of the most dynamic movements in protein structures.

Science, 2000 Dec 15, 290(5499), 2144 - 8
Global analysis of the genetic network controlling a bacterial cell cycle; Laub MT et al.; This report presents full-genome evidence that bacterial cells use discrete transcription patterns to control cell cycle progression . Global transcription analysis of synchronized Caulobacter crescentus cells was used to identify 553 genes (19% of the genome) whose messenger RNA levels varied as a function of the cell cycle . We conclude that in bacteria, as in yeast, (i) genes involved in a given cell function are activated at the time of execution of that function, (ii) genes encoding proteins that function in complexes are coexpressed, and (iii) temporal cascades of gene expression control multiprotein structure biogenesis . A single regulatory factor, the CtrA member of the two-component signal transduction family, is directly or indirectly involved in the control of 26% of the cell cycle-regulated genes.

BJOG, 2000 Nov, 107(11), 1427 - 32
A randomised controlled trial of vaginal clindamycin for early pregnancy bacterial vaginosis; Kurkinen-Raty M et al.; OBJECTIVE: To determine whether treatment of bacterial vaginosis (BV) with vaginal clindamycin affects pregnancy outcome . MATERIALS AND METHODS: Mothers with singleton pregnancies and without previous preterm delivery in 17 health centres in Oulu from March 1996 Until March 1998, in whom BV was diagnosed by Gram stain of a vaginal swab at the first antenatal visit (at the 12th gestational week) were randomised at Oulu University Hospital to have a one-week course of vaginal clindamycin, or placebo . A follow up sample of Gram stain was taken two weeks after randomisation and at the 30th gestational weeks . Pregnancy outcome data was obtained from hospital records . Primary outcome was preterm birth, and puerperal infectious morbidity the other outcome measure . RESULTS: During the study period 1956 women were screened, of whom 143 (7.3%) were BV- positive . One hundred and one were randomised . The total preterm birth rate of BV+ women randomised was 9.9% (10/101) . Preterm birth occurred in 20.7% (6/29) vs 0% (0/26) according to whether BV persisted or not (P < 0.01) . The preterm birth rate was 13.7% (7/51) in the clindamycin group vs 6.0% (3/50) in the placebo group (OR 2.5, 95% CI 0.6-10) . BV was cured just after treatment in 17 out of 51 (33%) of the clindamycin- treated patients vs 17 out of 50 (34%) of the placebo- treated patients (OR 1.0, 95% CI 0.4-2.2) . There was a difference in puerperal infectious morbidity in patients where BV persisted (31%, 9/29) compared with those in which BV did not persist (7.7%, 1/26) (OR 5.4, 95% CI 1.04-28) . Infections were seen in 4/51 (8%) of the clindamycin treated vs 10/50 (20%) of the placebo treated cases, (OR 0.3, 95% CI 0.1-1.2) . CONCLUSION: The prevalence of BV was lower than expected in this low risk population, but nevertheless it increased the risk of preterm birth and puerperal infectious morbidity, the risk being highest in cases where BV persisted during pregnancy . Vaginal clindamycin treatment for BV in the first trimester of pregnancy did not appear to reduce the risk of preterm birth or puerperal infections.

Oncol Rep, 2001 Jan-Feb, 8(1), 161 - 4
The bacterial polysaccharide tecogalan blocks growth of breast cancer cells in vivo; Yunmbam MK et al.; The growth of supportive tissue during the progression of solid tumors is an absolute requirement for the nourishment of the tumor . The blockade of this proliferative response of normal tissues to the growing tumor should hence inhibit tumor progression . We have shown earlier, that the heparinoid pentosan polysulfate (PPS) can block tumor growth and neoangiogenesis induced by Kaposi's FGF as well as by other heparin-binding growth factors (HBGFs) . We now report on the effects of a bacterial polysaccharide, tecogalan, on tumor xenografts of human breast cancer cells . Tecogalan inhibited FGF-dependent SW-13 cells in vitro very similarly to PPS . Growth factor-independent MDA-MB 231 cells were used in animal studies to assess the in vivo potential of tecogalan . Subcutaneous growth of tumors was inhibited by once weekly i.v . administration of tecogalan . PPS single weekly administration showed a similar effect . No gross side effects were observed . Based on our previous studies with these models, we conclude, that tecogalan acts by blocking HBGFs released from tumor cells . Interestingly, single weekly dosing of either PPS or tecogalan appears to be strikingly more efficacious than spreading the dose over several administrations . These findings with a novel compound, tecogalan, and a novel treatment regimen, PPS, suggests a different approach to planning of therapies with these types of drugs.

Genes Dev, 2000 Dec 1, 14(23), 2976 - 88
FtsK functions in the processing of a Holliday junction intermediate during bacterial chromosome segregation; Barre FX et al.; In bacteria with circular chromosomes, homologous recombination can generate chromosome dimers that cannot be segregated to daughter cells at cell division . Xer site-specific recombination at dif, a 28-bp site located in the replication terminus region of the chromosome, converts dimers to monomers through the sequential action of the XerC and XerD recombinases . Chromosome dimer resolution requires that dif is positioned correctly in the chromosome, and the activity of FtsK, a septum-located protein that coordinates cell division with chromosome segregation . Here, we show that cycles of XerC-mediated strand exchanges form and resolve Holliday junction intermediates back to substrate irrespective of whether conditions support a complete recombination reaction . The C-terminal domain of FtsK is sufficient to activate the exchange of the second pair of strands by XerD, allowing both intra- and intermolecular recombination reactions to go to completion . Proper positioning of dif in the chromosome and of FtsK at the septum is required to sense the multimeric state of newly replicated chromosomes and restrict complete Xer reactions to dimeric chromosomes.

J Pathol, 2000 Dec, 192(4), 554 - 9
An assessment of the long-term preservation of the DNA of a bacterial pathogen in ethanol-preserved archival material; Barnes I et al.; To examine the potential for DNA recovery from spirit-preserved medical material, a set of specimens from the Hunterian Collection of the Royal College of Surgeons was investigated . Using a range of DNA extraction techniques and the PCR, no replicable positive amplifications were made from this material of either human or Helicobacter DNA . Experiments with modern stomach biopsies of H . pylori-positive patients suggest that the bacterial DNA is typically present in a much lower concentration (10(3)-fold) than that of the host . The potential for recovery of this organism from spirit specimens is therefore low . The absence of DNA in this material is probably due to several factors, chiefly the incomplete fixation of the specimen by the ethanol storage fluid . Studies such as this demonstrate the need for a good understanding of specimen history when working with archival material .

Biochim Biophys Acta, 2000 Dec 15, 1535(1), 50 - 9
CINC blockade prevents neutrophil Ca(2+) signaling upregulation and gut bacterial translocation in thermal injury; Fazal N et al.; In this study, we have evaluated the role of cytokine-induced neutrophil chemoattractant (CINC), in the upregulation of neutrophil Ca(2+) signaling in neutrophils from thermally injured rats treated with anti-CINC antibody . Additionally, we have determined the effect of the treatment with CINC antibody on the accumulation of activated neutrophils in the intestinal wall, and the effect of such accumulation on gut bacterial translocation . Measurements of myeloperoxidase (MPO) activity and immunohistochemical localization of neutrophils determined neutrophil sequestration in the rat intestine . Agar culture analyses and a specific Escherichia coli beta-galactosidase gene polymerase chain reaction was carried out to detect gut indigenous bacterial invasion into intestinal wall and extraintestinal mesenteric lymph nodes (MLN) . The results showed that pretreatment of rats with anti-CINC antibody attenuated the thermal injury-induced enhancement in {Ca(2+)}(i) responses in neutrophils both in the basal and Formyl-Met-Leu-Phe stimulated conditions . Moreover, treatment with the CINC antibody decreased neutrophil infiltration into the gut and attenuated thermal injury-caused translocation of bacteria into the MLN.

Am J Respir Crit Care Med, 2000 Dec, 162(6), 2063 - 8
Community-acquired bacterial pneumonia in human immunodeficiency virus-infected patients: validation of severity criteria . The Grupo Andaluz para el Estudio de las Enfermedades Infecciosas; Cordero E et al.; Severity criteria for community-acquired pneumonia (CAP) have always excluded patients with human immunodeficiency virus (HIV) infection . A 1-yr, multicenter, prospective observational study of HIV-infected patients with bacterial CAP was done to validate the criteria used in the American Thoracic Society (ATS) guidelines for CAP, and to determine the prognosis-associated factors in the HIV-infected population with bacterial CAP . Overall, 355 cases were included, with an attributable mortality of 9.3% . Patients who met the ATS criteria had a longer hospital stay (p = 0.01), longer duration of fever (p < 0.001), and higher attributable mortality (13.1% versus 3.5%, p = 0.02) than those who did not . Three factors were independently related to mortality: CD4(+) cell count < 100/microl, radiologic progression of disease, and shock . Pleural effusion, cavities, and/or multilobar infiltrates at admission were independently associated with radiologic progression . A prognostic rule based on the five criteria of shock, CD4(+) cell count < 100/microl, pleural effusion, cavities, and multilobar infiltrates had a high negative predictive value for mortality (97.1%) . The attributable mortality for severe pneumonia was 11.3%, as compared with 1.3% for nonsevere disease (p = 0.008) . The ATS severity criteria are valid in HIV-infected patients with bacterial CAP . Our study provides the basis for identification of patients who may require hospitalization determined by clinical judgment and the five clinical criteria of shock, a CD4(+) cell count < 100/microl, pleural effusion, cavities, and multilobar involvement . These prognostic factors should be validated in independent cohort studies.

Int J Med Microbiol, 2000 Oct, 290(4-5), 441 - 5
Bordetella pertussis adenylate cyclase toxin as a tool to analyze molecular interactions in a bacterial two-hybrid system; Karimova G et al.; Bordetella pertussis secretes a calmodulin-activated adenylate cyclase toxin (CyaA) that is able to enter into eukaryotic cells . We took advantage of the modular structure of the catalytic domain of CyaA to design a genetic system that can detect protein-protein interactions in Escherichia coli . This bacterial two-hybrid system is based on the functional complementation between two complementary fragments, T25 and T18, of the catalytic domain of CyaA, in an E . coli cya strain . This bacterial two-hybrid system could find applications in the studies of structure/function relationships of proteins, in functional analysis of genomes, in high-throughput screening of interacting ligands and in design of new therapeutic agents.

J Obstet Gynecol Neonatal Nurs, 2000 Nov-Dec, 29(6), 606 - 12
Bacterial vaginosis and Chlamydia trachomatis among pregnant abused and nonabused Hispanic women; King EA et al.; OBJECTIVE: To compare the prevalence of bacterial vaginosis (BV) and Chlamydia trachomatis (CT) among abused and nonabused pregnant Hispanic women . DESIGN: Retrospective audit of charts of 233 pregnant, abused Hispanic women and 468 pregnant, nonabused Hispanic women . SETTING: Three urban prenatal clinics of a public health department in the southwestern United States . SAMPLE: The medical records of 701 pregnant Hispanic women . MAIN OUTCOME MEASURE: Diagnosis of BV and/or CT among abused and nonabused pregnant women . RESULTS: Combined prevalence of BV and CT was significantly higher for abused women (z score = 2.55; df = 138; p < .05) . There was no significant difference between abused and nonabused women for CT alone (z score = .96; df = 33; p < .05); however, prevalence of BV was significantly higher for abused women (z score = 1.99; df = 104; p < .05) . CONCLUSION: In this sample of pregnant Hispanic women prevalence of BV was significantly higher in those who had been abused, indicating the need for targeted screening of all abused pregnant women for BV.

Kansenshogaku Zasshi, 2000 Oct, 74(10), 811 - 5
{Serial measurement of anti-interleukin-8 IgG autoantibody in cerebrospinal fluid of infants with bacterial meningitis}; Takasaki J et al.; We serially measured concentrations of interleukin (IL)-8 and anti-IL-8 IgG autoantibody in cerebrospinal fluid of infants with bacterial meningitis, and also measured these concentrations in cerebrospinal fluid obtained from infants without meningitis on admission . We have reported that the IL-8 concentration in cerebrospinal fluid of infants with purulent meningitis rapidly decreases after the initiation of therapy . Thus, in the present study, the IL-8 concentration in infants with purulent meningitis only before the initiation of therapy was significantly higher compared with that in infants without meningitis . However, the concentration of anti-IL-8 IgG autoantibody was still high after the initiation of therapy . The concentration of anti-IL-8 IgG autoantibody was significantly higher compared with that in infants without meningitis until the 15th day after the initiation of therapy . The time lag between the decrease of IL-8 and anti-IL-8 IgG autoantibody demonstrated in the present study could be used to indicate the past presence of a large amount of IL-8, even if the IL-8 concentration was already low.

Int Microbiol, 1999 Jun, 2(2), 81 - 6
Bacterial symbioses . Predation and mutually beneficial associations; Esteve I et al.; The endosymbiotic theory, which has proved to explain the origin of mitochondria and chloroplasts, also posits the origin of nucleus and other cellular organelles that could have derived from ancient relationships among bacteria . It seems that predation might have been a prerequisite to the establishment of symbiosis as a source of evolutionary novelty . This review describes current different examples of bacteria able not only to attack and degrade other bacteria, but also to establish stable symbiotic relationships with different eukaryotic organisms.

Int Microbiol, 1999 Jun, 2(2), 71 - 80
Synthesis of the bacterial magnetosome: the making of a magnetic personality; Bazylinski DA; Magnetotactic bacteria synthesize intracellular, enveloped, single magnetic domain crystals of magnetite (Fe3O4, Fe2+Fe2(3+)O4) and/or greigite (Fe3S4) called magnetosomes . The magnetosomes contain well-ordered crystals that have narrow size distributions and consistent species- and/or strain-specific morphologies . These characteristics are features of a process called biologically-controlled mineralization in which an organism exerts a great degree of crystallochemical control over the nucleation and growth of the mineral particle . Because of these features, the mineral particles have been used as biomarkers although not without controversy . These unique structures impart a permanent magnetic dipole moment to the cell causing it to align and swim along geomagnetic field lines, a behavior known as magnetotaxis . The apparent biological advantage of magnetotaxis is that it aids cells in more efficiently locating and maintaining position in vertical chemical gradients common in many natural aquatic environments.

Int Microbiol, 1999 Mar, 2(1), 11 - 4
Accelerated evolution in bacterial endosymbionts of aphids; Buades C et al.; When compared with free living bacteria, it is proposed that there are at least two endosymbiotic processes in aphids based on the A + T content as well as the increased evolutionary rate of the beta-subunit of the F-ATPase complex in different endosymbiotic bacteria . The first well established process corresponds to the integration of Buchnera aphidicola more than 150 million years ago . The other is postulated to correspond to new endosymbiotic processes in which the bacteria involved contain less A + T and show a lower increase of evolutionary rates when compared with B . aphidicola . It is proposed, therefore, that endosymbioses are active processes in aphid evolution.

Immunology, 2000 Nov, 101(3), 333 - 41
Antigen-specific proliferation of porcine CD8alphaalpha cells to an extracellular bacterial pathogen; Waters WR et al.; A vaccine inducing protective immunity to a spirochaete-induced colitis of pigs predominantly stimulates expansion of CD8+ cells in vivo and in antigen-stimulated lymphocyte cultures . CD8+ cells, however, are rarely considered necessary for protection against extracellular bacterial pathogens . In the present study, pigs recovering from colitis resulting from experimental infection with Brachyspira (Serpulina) hyodysenteriae had increased percentages of peripheral blood CD4- CD8+ (alphaalpha-expressing) cells compared with non-infected pigs . CD8alphaalpha+ cells proliferated in antigen-stimulated cultures of peripheral blood mononuclear cells from B . hyodysenteriae-vaccinated pigs . Proliferating CD8alphaalpha+ cells consisted of CD4-, CD4+ and gammadelta T-cell receptor-positive cells . CD4- CD8alphabeta+ cells from vaccinated or infected pigs did not proliferate upon in vitro antigen stimulation . Of the CD8alphaalpha cells that had proliferated, flow cytometric analysis indicated that the majority of the CD4+ CD8+ cells were large (i.e . lymphoblasts) whereas the CD4- CD8+ cells were predominantly small . Addition of monoclonal antibodies (mAb) specific for either porcine major histocompatibility complex (MHC) class I or class II antigens diminished B . hyodysenteriae-specific proliferative responses whereas addition of mAb to porcine MHC II, but not porcine MHC I, reduced the CD8alphaalpha response . In vitro depletion of CD4+ cells by flow cytometric cell sorting diminished, but did not completely abrogate, the proliferative response of cells from vaccinated pigs to B . hyodysenteriae antigen stimulation . These results suggest that CD8alphaalpha cells are involved in recovery and possibly protection from a spirochaete-induced colitis of pigs; yet, this response appears to be partially dependent upon CD4+ cells.

J Infect Dis, 2001 Jan 1, 183(1), 149 - 53 Epub 2000 Nov 30.
Vascular endothelial growth factor in bacterial meningitis: detection in cerebrospinal fluid and localization in postmortem brain; van der Flier M et al.; Vascular endothelial growth factor (VEGF) is a potent vascular permeability factor and a mediator of brain edema . To assess the role of VEGF during bacterial meningitis, VEGF was measured in cerebrospinal fluid (CSF) and blood of 37 patients with bacterial meningitis and 51 control patients, including 16 patients with viral meningitis . Circulating VEGF levels were similar in bacterial meningitis patients and control patients . VEGF(CSF) was detected in 11 (30%) of 37 of bacterial meningitis patients (range, <25-633 pg/mL) but in none of the control patients . The median VEGF index was 6.2 (range, 0.6-42), indicating intrathecal production . Median CSF cell counts, protein levels, and CSF: serum albumin ratios were higher for patients with detectable VEGF(CSF), although the difference was not statistically significant . VEGF immunoreactivity in autopsy brain specimens was found in the inflammatory infiltrate of patients with bacterial meningitis . These results indicate that inflammatory cells secrete VEGF during bacterial meningitis and that VEGF may contribute to blood-brain barrier disruption.

Biochemistry, 2000 Dec 12, 39(49), 15032 - 43
Ubiquinone binding, ubiquinone exclusion, and detailed cofactor conformation in a mutant bacterial reaction center; McAuley KE et al.; The X-ray crystal structure of a Rhodobacter sphaeroides reaction center with the mutation Ala M260 to Trp (AM260W) has been determined . Diffraction data were collected that were 97.6% complete between 30.0 and 2.1 A resolution . The electron density maps confirm the conclusions of a previous spectroscopic study, that the Q(A) ubiquinone is absent from the AM260W reaction center (Ridge, J . P., van Brederode, M . E., Goodwin, M . G., van Grondelle, R., and Jones, M . R . (1999) Photosynthesis Res . 59, 9-26) . Exclusion of the Q(A) ubiquinone caused by the AM260W mutation is accompanied by a change in the packing of amino acids in the vicinity of the Q(A) site that form part of a loop that connects the DE and E helices of the M subunit . This repacking minimizes the volume of the cavity that results from the exclusion of the Q(A) ubiquinone, and further space is taken up by a feature in the electron density maps that has been modeled as a chloride ion . An unexpected finding is that the occupancy of the Q(B) site by ubiquinone appears to be high in the AM260W crystals, and as a result the position of the Q(B) ubiquinone is well-defined . The high quality of the electron density maps also reveals more precise information on the detailed conformation of the reaction center carotenoid, and we discuss the possibility of a bonding interaction between the methoxy group of the carotenoid and residue Trp M75 . The conformation of the 2-acetyl carbonyl group in each of the reaction center bacteriochlorins is also discussed.

Genetics, 2000 Dec, 156(4), 1493 - 501
Experimental analysis of molecular events during mutational periodic selections in bacterial evolution; Notley-McRobb L et al.; A fundamental feature of bacterial evolution is a succession of adaptive mutational sweeps when fitter mutants take over a population . To understand the processes involved in mutational successions, Escherichia coli continuous cultures were analyzed for changes at two loci where mutations provide strong transport advantages to fitness under steady-state glucose limitation . Three separate sweeps, observed as classic periodic selection events causing a change in the frequency of neutral mutations (in fhuA causing phage T5 resistance), were identified with changes at particular loci . Two of the sweeps were associated with a reduction in the frequency of neutral mutations and the concurrent appearance of at least 13 alleles at the mgl or mlc loci, respectively . These mgl and mlc polymorphisms were of many mutational types, so were not the result of a mutator or directed mutation event . The third sweep observed was altogether distinct and involved hitchhiking between T5 resistance and advantageous mgl mutations . Moreover, the hitchhiking event coincided with an increase in mutation rates, due to the transient appearance of a strong mutator in the population . The spectrum of mgl mutations among mutator isolates was distinct and due to mutS . The mutator-associated periodic selection also resulted in mgl and fhuA polymorphism in the sweeping population . These examples of periodic selections maintained significant genotypic diversity even in a rapidly evolving culture, with no individual "winner clone" or genotype purging the population.

J Pediatr Surg, 2000 Dec, 35(12), 1701 - 5
A model of bacterial translocation in neuroblastoma-bearing mice; Kanai M et al.; PURPOSE: The aim of this study was to establish a model of bacterial translocation (BT) in neuroblastoma-bearing mice . METHODS: A suspension of 1 x 10(6) cells of the murine neuroblastoma cell line C1300 was injected subcutaneously into the thighs of 8-week-old female A/J mice, which were then killed after 7, 14, and 21 days . Some of the mice were given 1-microm or 2-microm fluorescein-labeled latex beads in their drinking water for 7 days before being killed . Mesenteric lymph nodes (MLNs) were aseptically removed and cultured for 72 hours at 37 degrees C . Segments of distal ileum were obtained for histologic examination . Samples of venous blood were obtained for laboratory tests . RESULTS: Tumors were found at the injection sites on days 14 and 21 after C1300 injection . Although tumors were not found in 7 days, significantly high number of 1-microm latex beads were detected in MLNs compared with the control, and the number increased with tumor growth . The number of 2-microm latex beads was significantly higher on days 14 and 21 . The percentage of mice with MLN cultures positive were significantly higher on day 14, and the percentage increased along with tumor growth . On day 21 after C1300 injection, body weight loss and anemia were observed, and histologic findings of the terminal ileum showed mucosal edema and villous thinning . Serum levels of interleukin (IL)-6 were significantly higher in mice killed 14 and 21 days after injection . CONCLUSIONS: The results suggest that BT from the gut to MLNs may occur in neuroblastoma C1300-bearing mice, and it increases along with tumor growth . Even in the early stage of malignancy, particles as small as 1 microm may translocate from the gut to MLNs.

EMBO J, 2000 Dec 1, 19(23), 6419 - 26
SRP-dependent co-translational targeting and SecA-dependent translocation analyzed as individual steps in the export of a bacterial protein; Neumann-Haefelin C et al.; Recently it has been recognized that the signal recognition particle (SRP) of Escherichia coli represents a specific targeting device for hydrophobic inner membrane proteins . It has remained unclear, however, whether the bacterial SRP functions in concert with SecA, which is required for the translocation of secretory proteins across the inner membrane . Here, we have analyzed a hybrid protein constructed by fusing the signal anchor sequence of an SRP-dependent inner membrane protein (MtlA) to the mature part of an exclusively SecA-requiring secretory protein (OmpA) . We show that the signal anchor sequence of MtlA confers the novel properties onto nascent chains of OmpA of being co-translationally recognized and targeted to SecY by SRP . Once targeted to SecY, ribosome-associated nascent chains of the hybrid protein, however, remain untranslocated unless SecA is present . These results indicate that SRP and SecA cooperate in a sequential, non-overlapping manner in the topogenesis of those membrane proteins which, in addition to a signal anchor sequence, harbor a substantial hydrophilic domain to be translocated into the periplasm.

Org Lett, 2000 Nov 30, 2(24), 3857 - 60
Vinylogous amide analogues of diaminopimelic acid (DAP) as inhibitors of enzymes involved in bacterial lysine biosynthesis; Caplan JF et al.; {reaction: see text} Vinylogous amides 5 and 6 have been synthesized from L-propargyl glycine and tested against diaminopimelate (DAP) enzymes involved in bacterial lysine biosynthesis . Both are reversible inhibitors of DAP D-dehydrogenase and DAP epimerase with IC(50) values in the 500 microM range . Compound 5 shows competitive inhibition against the L-dihydrodipicolinate (DHDP) reductase with a K(i) value of 32 microM, which is comparable to the planar dipicolinate 16 (K(i) = 26 microM), the best known inhibitor of the enzyme.

Biochemistry, 2000 Dec 5, 39(48), 14822 - 30
Preferential binding of equine ferricytochrome c to the bacterial photosynthetic reaction center from Rhodobacter sphaeroides; Larson JW et al.; Redox titration of horse heart cytochrome c (cyt c), in the presence of varying concentrations of detergent-solubilized photosynthetic reaction center (RC) from Rhodobacter sphaeroides, revealed an RC concentration-dependent decrease in the measured cyt c midpoint potential that is indicative of a 3.6 +/- 0.2-fold stronger binding affinity of oxidized cytochrome to a single binding site . This effect was correlated with preferential binding in the functional complex by redox titration of the fraction of RCs exhibiting microsecond, first-order, special pair reduction by cytochrome . A binding affinity ratio of 3.1 +/- 0.4 was determined by this second technique, confirming the result . Redox titration of flash-induced intracomplex electron transfer also showed the association in the electron transfer-active complex to be strong, with a dissociation constant of 0.17 +/- 0.03 microM . The tight binding is associated with a slow off-rate which, in the case of the oxidized form, can influence the kinetics of P(+) reduction . The pitfalls of the common use of xenon flashlamps to photoexcite fast electron-transfer reactions are discussed with relation to the first electron transfer from primary to secondary RC quinone acceptors . The results shed some light on the diversity of kinetic behavior reported for the cytochrome to RC electron-transfer reaction.

Microbes Infect, 2000 Oct, 2(12), 1407 - 17
Bacterial and host-derived cationic proteins bind alpha2-laminins and enhance Mycobacterium leprae attachment to human Schwann cells; de Melo Marques MA et al.; It has recently been demonstrated that laminin alpha2 chains present on the surface of Schwann cells are involved in the process of attachment of Mycobacterium leprae to these cells . In this study, a protein in the M . leprae cell wall that was found to be capable of binding alpha2-containing laminins (merosin) was isolated and characterized . The M . leprae laminin-binding protein was identified as a 21-kDa histone-like protein (Hlp), a highly conserved cationic protein present in other species of mycobacteria . The gene that encodes this protein was PCR amplified, cloned, and expressed, and the recombinant protein was shown to bind alpha2-laminins . More significantly, when added exogenously, Hlp was able to greatly enhance the attachment of mycobacteria to ST88-14 human Schwann cells . The capacity to bind alpha2-laminins and to enhance mycobacterial adherence to Schwann cells was also found in other cationic proteins such as host-derived histones . Moreover, mutation in the hlp gene was shown not to affect the capacity of mycobacteria to bind to ST88-14 cells, suggesting that alternative adhesins and/or pathways might be used by mycobacteria during the process of adherence to Schwann cells . The potential role of Hlp as a fortuitous virulence factor contributing to the pathogenesis of M . leprae-mediated nerve damage is discussed.

Crit Care Med, 2000 Nov, 28(11), 3692 - 6
Inducible nitric oxide synthase gene knockout mice have increased resistance to gut injury and bacterial translocation after an intestinal ischemia-reperfusion injury; Suzuki Y et al.; OBJECTIVE: Intestinal ischemia-reperfusion after severe shock states is often associated with bacterial translocation and intestinal barrier dysfunction . Our previous studies showed that inducible nitric oxide synthase (iNOS) gene knockout mice were resistant to endotoxin-induced bacterial translocation and ileal mucosal damage . The goal of this study was to test whether iNOS mediates bacterial translocation after intestinal ischemia-reperfusion, using iNOS knockout mice (iNOS-/-) and their wild-type littermates (iNOS+/+) . DESIGN: Prospective animal study with concurrent controls . SETTING: Small animal laboratory . SUBJECTS: Thirty-eight iNOS knockout mice and 51 wild-type littermates . INTERVENTIONS: iNOS+/+ mice or iNOS-/- mice were subjected to a sham operation or 30 mins of superior mesenteric artery occlusion followed by reperfusion . Twenty-four hours after reperfusion, bacterial translocation to mesenteric lymph nodes, ileal villous damage, and cecal bacterial population were evaluated . MEASUREMENTS AND MAIN RESULTS: Sham operation did not induce bacterial translocation, change cecal bacterial population levels, or cause ileal villous damage . Intestinal ischemia-reperfusion caused bacterial translocation in 72% of the iNOS+/+ mice but only 28% of the iNOS-/- mice . Both iNOS+/+ and iNOS-/- mice subjected to superior mesenteric artery occlusion (SMAO) in which bacterial translocation occurred had cecal bacterial population levels that were three logs higher than mice subjected to sham SMAO or mice subjected to SMAO in which bacterial translocation did not occur . The magnitude of villous injury was less in the iNOS-/- mice than the iNOS+/+ mice after SMAO, although the incidence of ileal villous damage was significantly higher in both the iNOS+/+ and iNOS-/- mice in which bacterial translocation occurred after SMAO than in the mice in which bacterial translocation did not occur after SMAO . iNOS+/+ mice subjected to SMAO had increased plasma concentrations of nitrite (NO2-) and nitrate (NO3-), and the plasma concentrations of NO2- and NO3- were highest in the mice in which bacterial translocation had occurred . CONCLUSION: iNOS knockout mice were more resistant to intestinal ischemia-reperfusion-induced bacterial translocation and mucosal injury than wild-type mice, suggesting that iNOS might play a role in intestinal ischemia-reperfusion-induced loss of gut barrier function.

Braz J Infect Dis, 1999 Feb, 3(1), 15 - 22
C-reactive Protein Follow-up of Children With Acute Bacterial Meningitis; Dias Leite R et al.; Twenty-two children with bacterial meningitis were prospectively studied to follow C-reactive protein (CRP) in serum at admission, 2nd, 5th and 7th days of treatment, and in the cerebrospinal fluid (CSF) at admission, to investigate if there is any relationship of its levels with the clinical evolution . CRP was measured by latex agglutination and/or ELISA techniques with detection limits of 0.15mg/L and 0.9mg/L, respectively . Patients were classified according to clinical evolution in two groups: uneventful recovery (n=12) and complicated evolution (n=10) . Clinical complications observed were: relapse of fever (8), persistent fever (4), arthritis (4), ventricle enlargement (2), subdural effusion (1), subdural empyema (1), ataxia (1), cervical hypotonia (1), deafness (1), endophthalmitis (1), acute otitis media (1), secondary skin infection (1) and treatment change due to poor clinical response (1) . A significant fall in CRP serum levels was observed among the uneventful recovery group after admission . Distinctly, in the group with a complicated evolution CRP levels showed either secondary elevation or remained high continuously . Mean serum CRP levels were significantly lower in the uneventful recovery group than in the complicated evolution group on 5th day and on 7th day . CRP levels below 20mg/L on 5th and 7th days were associated with an uneventful recovery, and CRP levels higher than 20mg/L on those same days were associated with a complicated clinical evolution (p=0.01* and p=0.0015*, respectively) . We conclude that serum CRP levels monitoring in children with bacterial meningitis represents a useful and objective information about the clinical evolution . This procedure is inexpensive and suitable for use in endemic areas lacking sophisticated laboratories.

Curr Infect Dis Rep, 2000 Dec, 2(6), 506 - 512
Recurrent Bacterial Vaginosis; Hay P; Bacterial vaginosis (BV) is a common cause of vaginal discharge in women of childbearing age . In some individuals, it recurs frequently after treatment, frustrating both the patient and the physician . Standard BV treatment--metronidazole or clindamycin, administered either intravaginally or orally--is followed by relapse in approximately 30% of cases, within one month . Our inability to prevent relapse reflects our lack of understanding of how BV originates . BV has been associated with infectious morbidity in obstetrics and gynecology . Recent studies have found it to be a risk factor for HIV spread . These findings increase the need for us to be able to control recurrent BV and reduce its prevalence in the general population.

Curr Infect Dis Rep, 2000 Aug, 2(4), 332 - 336
New Advances in the Pathogenesis and Pathophysiology of Bacterial Meningitis; Nathan BR et al.; Acute bacterial meningitis continues to be a significant health concern, with a fatality rate of more than 30% in some studies . Although the face of bacterial meningitis has changed substantially over the past 15 years, this disease still causes significant mortality (particularly in underdeveloped countries) and neurological sequelae . Our understanding of the pathophysiology of bacterial meningitis continues to develop . Our understanding of the mechanisms of neuronal injury now includes the concept that many of the pathological changes are only secondary to the infection and that the human immune system contributes to the majority of the neuronal death . A complicated series of interactions among immune, vascular and central nervous system cells, cytokines and chemokines, matrix metalloproteinases and free radical molecules are ultimately responsible for many bacterial meningitis changes . We hope that a complete understanding of these processes will ultimately lead to better diagnostic techniques and improved treatments.

Curr Infect Dis Rep, 2000 Apr, 2(2), 154 - 159
Viral-Bacterial Synergy in Otitis Media: Implications for Management; Heikkinen T et al.; Despite the extensive use of broad-spectrum antibiotics, poor clinical response to the treatment of acute otitis media is common . Evidence derived from numerous studies during the past two decades supports a crucial role for respiratory viruses in the etiology and pathogenesis of acute otitis media, and recent studies indicate that viruses may also have a profound adverse effect on the resolution of this disease . Viruses seem to interact with bacteria and enhance the local inflammatory process in the middle ear . Effective adjuvant therapies are needed to improve outcome in children with otitis media . Increasing knowledge of the role of viruses, viral-bacterial interaction, and host inflammatory mechanisms in otitis media may lead to major improvements in the management of this disease.

Curr Infect Dis Rep, 1999 Oct, 1(4), 393 - 397
Gynecologic Complications of Bacterial Vaginosis: Fact or Fiction?
Soper DE.
Bacterial vaginosis is a complex alteration of vaginal flora causing mild symptoms in women characterized by a milky vaginal discharge associated with a fishy odor worsened after unprotected vaginal coitus . It is the most common cause of vaginitis . It is now clear that this seemingly harmless disorder causing nuisance symptoms is linked to a number of important adverse reproductive tract sequelae, including the obstetrical sequelae of preterm labor and delivery . Literature published over the past year continue to support the association of bacterial vaginosis with adverse gynecologic sequelae in women.

J Bacteriol, 2000 Dec, 182(24), 7092 - 6
The truncated form of the bacterial heat shock protein ClpB/HSP100 contributes to development of thermotolerance in the cyanobacterium Synechococcus sp . strain PCC 7942; Clarke AK et al.; ClpB is a highly conserved heat shock protein that is essential for thermotolerance in bacteria and eukaryotes . One distinctive feature of all bacterial clpB genes is the dual translation of a truncated 79-kDa form (ClpB-79) in addition to the full-length 93-kDa protein (ClpB-93) . To investigate the currently unknown function of ClpB-79, we have examined the ability of the two different-sized ClpB homologues from the cyanobacterium Synechococcus sp . strain PCC 7942 to confer thermotolerance . We show that the ClpB-79 form has the same capacity as ClpB-93 to confer thermotolerance and that the ClpB-79 protein contributes ca . one-third of the total thermotolerance developed in wild-type Synechococcus, the first in vivo demonstration of a functional role for ClpB-79 in bacteria.

Hosp Med, 2000 Jul, 61(7), 475 - 7
Adverse obstetric sequelae of bacterial vaginosis; Adinkra P et al.; Bacterial vaginosis is increasingly associated with adverse sequelae in obstetrics such as preterm prelabour rupture of membranes, preterm labour and preterm birth . It is important to diagnose the condition early in pregnancy where appropriate treatment can be administered to women who are symptomatic or who are at high risk of adverse sequelae.

Int J Antimicrob Agents, 2000 Nov, 16(3), 357 - 60
Effect of highly active antiretroviral therapy on the incidence of bacterial pneumonia in HIV-infected subjects; de Gaetano Donati K et al.; The aim of the study was to assess the incidence of hospital and community acquired bacterial pneumonia in HIV-infected subjects prior to and after the introduction of highly active antiretroviral therapy (HAART) . We studied 266 patients with bacterial pneumonia over two separate periods, 154 in the first period and 112 in the second period . A statistically significant difference in the incidence of bacterial pneumonia in the two study periods was observed ranging from 13.1 to 8.5 episodes per 100 persons . The incidence of community-acquired bacterial pneumonia decreased from 10.7 to 7.7 (P=0.01), while that of nosocomial episodes decreased from 2.4 to 0.8 episodes (P=0.003) . Low levels of peripheral CD(4+) cells (<100/mm(3)) and intravenous drug abuse (IVDA) were significantly associated with the development of community-acquired bacterial pneumonia, while an increasing value of APACHE III score and prolonged hospitalisation increased the risk of nosocomial bacterial pneumonia in both study periods.

Int J Immunopharmacol, 2000 Nov, 22(11), 989 - 99
Radioprotection of haemopoietic stem cells by a single injection of bacterial lysate - IRS-19 administered to mice before or after irradiation; Fedorocko P et al.; Data in this report describes the effect of a single injection of bacterial lysate IRS-19 prior to irradiation of C57Bl/6 mice on recovery of colony-forming cells (CFC) after sublethal and lethal doses of radiation . The injection of IRS-19 promoted an earlier recovery of colony-forming cells in the bone marrow and spleen . For example, 5 and 9 days after 7.5 Gy irradiation, the number of CFU-S per femur was approximately 1.7-2.3-fold higher in IRS-19-injected mice than in saline-injected mice . Also, pretreatment of mice with IRS-19 induced an increase in the number of endogenous haemopoietic stem cells (endoCFU-S) . In the postradiation period (5-21 days) significantly increased bone marrow and spleen cellularity and accelerated myelopoietic regeneration (committed progenitor granulocyte-macrophage-colony-forming cells, GM-CFC) in the bone marrow and spleen compared with saline-treated controls . At the time of presumed irradiation, (i.e . 24 h after administration of the drug to the non-irradiated mice), there was no significant difference between the control mice and mice treated with IRS-19 in numbers of femoral and spleen GM-CFC . In contrast, the number of nucleated femoral cells decreased significantly in the group treated with IRS-19 . Moreover, treatment with IRS-19 caused a sustained increase in serum colony-stimulating activity which was followed by an enhanced repopulation of GM-CFC in the femoral marrow and spleen . Administration of the agent 24 h prior to irradiation rather than postirradiation appeared most effective with respect to radioprotection . Intravenous rather than i.p . and p.o . was the most effective route of administration in the mouse . Furthermore, single, high-dose injection appeared to be more effective than repeated, lower dose injections . Results suggest that the radioprotective properties associated with the administration of IRS-19 are largely a consequence of the induction of haemopoietic colony-stimulating activities and potentially the activation and/or enhancement of cytokine cascades in the recipient animals . These changes may ultimately impact the cell cycle profile of the haemopoietic cells and therefore their ability to withstand and/or recover from radiation insult.

Breast Cancer, 1994 Dec 30, 1(2), 89 - 94
Complete Nucleotide Sequence of Mouse Mammary Tumor Virus from JYG Chinese Wild Mice: Absence of Bacterial Insertion Sequences in the Cloned Viral gag Gene; Nishio M et al.; Mammary tumors of a newly isolated strain of Chinese wild mouse (JYG mouse) harbor exogenous mouse mammary tumor virus (MMTV) . The complete nucleotide sequence of exogenous JYG-MMTV was determined on the proviral 5' long terminal repeat (LTR)(partial)-gag-pol-env-3' LTR (partial) fragment cloned into a plasmid vector and the cDNA sequence from JYG-MMTV producing cells . Similarly to the other MMTV species the LTR of JYG-MMTV contains an open reading frame (ORF) . The amino acid sequence of the JYG-MMTV ORF resembles that of SW-MMTV (92% identity) and endogenous Mtv-7 (93% identity) especially at the C-terminal region . Thus, a functional similarity in T-cell receptor V beta recognition as a superantigen is implicated among these MMTV species . Analysis of the viral gag nucleotide sequence revealed that this gene is not disrupted by the bacterial insertion sequence IS1 or IS2, which have been reported to be present in the majority of the plasmids containing the gag region . Comparison of amino acid sequences of JYG-MMTV with those of BR6-MMTV showed that over 96% of the amino acids of gag, pol, protease and env products are identical . These results suggest the intact nature of the nucleotide sequence of the near full-length MMTV genome cloned in the plasmid.

Endocrinology, 2000 Nov, 141(11), 4000 - 12
Diametric effects of bacterial endotoxin lipopolysaccharide on adrenal and Leydig cell steroidogenic acute regulatory protein; Hales KH et al.; Immune activation results in the activation of adrenal steroidogenesis and inhibition of gonadal steroidogenesis . Previous studies indicated that these effects were caused primarily by activation and suppression of the secretion of ACTH and LH, respectively . However, other evidence indicated a direct effect of the immune system on the gonads . In this study, serum testosterone, quantitated by RIA after lipopolysaccharide injection, showed a significant decrease within 2 h . Parallel measurement of serum LH showed no change . There were no differences in LH receptor or cAMP produced in Leydig cells between vehicle- and lipopolysaccharide-injected mice . The 30-kDa form of the steroidogenic acute regulatory (StAR) protein was quantitated, by Western blot, in Leydig cells and was found to decrease in a time-dependent manner . No change in StAR protein messenger RNA (mRNA) was detected by Northern analysis during this time, nor were any changes found in the levels of mRNA for the steroidogenic enzymes P450scc, 3beta-hydroxysteroid dehydrogenase delta4-delta5-isomerase, or P450c17 . In the adrenal, StAR protein was increased, as was StAR protein mRNA . No changes were observed in the levels of mRNA for P450scc, 3beta-hydroxysteroid dehydrogenase delta4-delta5-isomerase, or P450c21 . Thus, although the mechanisms of regulation differ, changes in the levels of StAR protein are a sensitive indicator of the steroidogenic capacity of these two tissues.

Phys Rev E Stat Phys Plasmas Fluids Relat Interdiscip Topics, 2000 Jul, 62(1 Pt B), 1444 - 7
Nutrient chemotaxis suppression of a diffusive instability in bacterial colony dynamics; Arouh S et al.; Bacteria grown on a semisolid agar surface have been observed to form branching patterns as the colony envelope propagates outward . The fundamental cause of this instability relates to the need for limited nutrient to diffuse towards the colony . Here, we investigate the effect on this instability of allowing the bacteria to move chemotactically in response to the nutrient gradient . Our results show that this additional effect has a tendency to suppress the instability . Our calculations are done within the context of a simple "cutoff" model of colony dynamics, but presumably remain valid for more complex and hence more realistic approaches.

Proc Natl Acad Sci U S A, 2000 Dec 5, 97(25), 13567 - 72
Zinc plays a key role in human and bacterial GTP cyclohydrolase I; Auerbach G et al.; The crystal structure of recombinant human GTP cyclohydrolase I was solved by Patterson search methods by using the coordinates of the Escherichia coli enzyme as a model . The human as well as bacterial enzyme were shown to contain an essential zinc ion coordinated to a His side chain and two thiol groups in each active site of the homodecameric enzymes that had escaped detection during earlier studies of the E . coli enzyme . The zinc ion is proposed to generate a hydroxyl nucleophile for attack of imidazole ring carbon atom eight of the substrate, GTP . It may also be involved in the hydrolytic release of formate from the intermediate, 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5'-triphosphate, and in the consecutive Amadori rearrangement of the ribosyl moiety.

Mol Gen Genet, 2000 Oct, 264(3), 241 - 50
The broad bean nodulin VfENOD18 is a member of a novel family of plant proteins with homologies to the bacterial MJ0577 superfamily; Hohnjec N et al.; Full-length transcript sequences were isolated from broad bean root nodules, which encode a novel nodulin designated VfENOD18 . The corresponding transcripts were detected in early and in late stages of nodule development and were localized exclusively in the nitrogen-fixing zone III . The VfENOD18 sequence is not only homologous to a number of ESTs from various mono- and dicotyledonous plants, but also to the ATP-binding protein MJ0577 from Methanococcus jannaschii and to a range of bacterial proteins that belong to the MJ0577 superfamily . Hence, VfENOD18 is a member of a ubiquitous family of plant proteins that might function as ATP-binding proteins or ATPases . On the genomic level, VfENOD18 genes can be divided into two groups on the basis of differences in their 5' UTRs . One group lacks the 5' UTR region including the ATG initiation codon, whereas the second group contained the complete 5' UTR region . Further upstream of this VfENOD18 gene, a retrotransposon sequence was identified . The -14/-964 VfENOD18 promoter fragment was devoid of complete organ-specific elements known from other nodulin gene promoters . Nevertheless, this region was able to mediate full promoter activity in the central region of transgenic Vicia hirsuta root nodules.

Curr Biol, 2000 Nov 2, 10(21), R777 - 80
Bacterial motility: how do pili pull?
Kaiser D.
Forceful retraction of a bacterial pilus has been directly observed for the first time . As retraction clarifies the basic mechanochemistry of single cell twitching and gliding movements, so cell-to-cell signalling by contact clarifies the coordination of multicellular gliding movements.

AIDS Read, 1999 Nov, 9(8), 580 - 3
Journal Club . HIV-associated bacterial pneumonia; Sanders A; Bacterial pneumonia is the most common cause of death from pneumonia in patients with HIV disease, causing greater mortality than Pneumocystis carinii pneumonia . The challenge for the clinician evaluating the HIV-infected patient with pneumonia is to quickly distinguish clinically among all possible causes and to initiate therapy based on the most likely diagnosis . While an understanding of typical clinical and radiographic presentations is essential, bronchoscopy is the preferred test for reliably identifying the causative organism.

Eur J Biochem, 2000 Dec, 267(23), 6849 - 57
Cloning and bacterial expression of monomeric short-chain dehydrogenase/reductase (carbonyl reductase) from CHO-K1 cells; Terada T et al.; Mammalian carbonyl reductase (EC 1.1.1.184) is an enzyme that can catalyze the reduction of many carbonyl compounds, using NAD(P)H . We isolated a cDNA of carbonyl reductase (CHO-CR) from CHO-K1 cells which was 1208 bp long, including a poly(A) tail, and contained an 831-bp ORF . The deduced amino-acid sequence of 277 residues contained a typical motif for NADP+-binding (TGxxxGxG) and an SDR active site motif (S-Y-K) . CHO-CR closely resembles mammalian carbonyl reductases with 71-73% identity . CHO-CR cDNA had the highest similarity to human CBR3 with 86% identity . Using the pET-28a expression vector, recombinant CHO-CR (rCHO-CR) was expressed in Escherichia coli BL21 (DE3) cells and purified with a Ni2+-affinity resin to homogeneity with a 35% yield . rCHO-CR had broad substrate specificity towards xenobiotic carbonyl compounds . RT-PCR of Chinese hamster tissues suggest that CHO-CR is highly expressed in kidney, testis, brain, heart, liver, uterus and ovary . Southern blotting analysis indicated the complexity of the Chinese hamster carbonyl reductase gene.

Genome, 2000 Oct, 43(5), 820 - 6
Construction of a binary bacterial artificial chromosome library of Petunia inflata and the isolation of large genomic fragments linked to the self-incompatibility (S-) locus; McCubbin AG et al.; The Solanaceae family of flowering plants possesses a type of self-incompatibility mechanism that enables the pistil to reject self pollen but accept non-self pollen for fertilization . The pistil function in this system has been shown to be controlled by a polymorphic gene at the S-locus, termed the S-RNase gene . The pollen function is believed to be controlled by another as yet unidentified polymorphic gene at the S-locus, termed the pollen S-gene . As a first step in using a functional genomic approach to identify the pollen S-gene, a genomic BAC (bacterial artificial chromosome) library of the S2S2 genotype of Petunia inflata, a self-incompatible solanaceous species, was constructed using a Ti-plasmid based BAC vector, BIBAC2 . The average insert size was 136.4 kb and the entire library represented a 7.5-fold genome coverage . Screening of the library using cDNAs for the S2-RNase gene and 13 pollen-expressed genes that are linked to the S-locus yielded 51 positive clones, with at least one positive clone for each gene . Collectively, at least 2 Mb of the chromosomal region was spanned by these clones . Together, three clones that contained the S2-RNase gene spanned approximately 263 kb . How this BAC library and the clones identified could be used to identify the pollen S-gene and to study other aspects of self-incompatibility is discussed.

Laryngoscope, 2000 Nov, 110(11), 1954 - 6
Absence of bacterial infection in the mucosal secretion in chronic laryngitis; Ebenfelt A et al.; OBJECTIVES: To examine whether an infectious process is present in the laryngeal secretion in patients with chronic laryngitis . STUDY DESIGN: Mucosal secretion from vocal cords and ventricular folds from 14 patients with chronic laryngitis was examined . Twelve patients with healthy larynxes served as control subjects . METHODS: Secretion from the laryngeal mucosa was sampled with an imprint technique during general anesthetic . The samples were stained and examined by light and fluorescence microscopy . The numbers of leukocytes and bacteria and the extension of phagocytosis were estimated . RESULTS: In the secretion from eight of the patients with chronic laryngitis we could observe huge numbers of bacteria, whereas only few bacteria were seen in the secretion from the control subjects . However, in both groups, only a few neutrophils were observed and phagocytosis was not present, indicating that the bacteria were present as colonizers . CONCLUSIONS: No infectious process is present in the secretion in chronic laryngitis . Further studies concerning the pathogenesis should focus on the pathological processes or conditions of the mucosa.

J Mol Evol, 2000 Nov, 51(5), 459 - 63
Replication orientation affects the rate and direction of bacterial gene evolution; Tillier ER et al.; In many bacterial genomes, the leading and lagging strands have different skews in base composition; for example, an excess of guanosine compared to cytosine on the leading strand . We find that Chlamydia genes that have switched their orientation relative to the direction of replication, for example by inversion, acquire the skew of their new "host" strand . In contrast to most evolutionary processes, which have unpredictable effects on the sequence of a gene, replication-related skews reflect a directional evolutionary force that causes predictable changes in the base composition of switched genes, resulting in increased DNA and amino acid sequence divergence.

FEBS Lett, 2000 Nov 10, 484(3), 280 - 4
Quenching of singlet oxygen by carotenoids produced in escherichia coli - attenuation of singlet oxygen-mediated bacterial killing by carotenoids; Tatsuzawa H et al.; We examined the viability of Escherichia coli transformants harboring various carotenoids synthesizing genes in a medium containing an enzymatic singlet oxygen generating system, which contained myeloperoxidase, hydrogen peroxide and Br(-) at pH 4.5 . Singlet oxygen quenching activities of various carotenoids in phosphatidyl choline micelles in aqueous media were also studied using the same enzymatic singlet oxygen generating system . Viability of the transformants producing carotenoids was higher than that of the wild type E . coli in the singlet oxygen generation mixture . Of the transformants tested, the viability of zeaxanthin-diglucoside producing transformant was the highest . Carotenoids in increasing order of k(q) values were beta-carotene, a cyclic carotene<zeaxanthin with hydroxy groups < or =lycopene, an acyclic carotene<canthaxanthin and astaxanthin with keto groups <<zeaxanthin-diglucoside . The k(q) value of zeaxanthin-diglucoside was 3.5 times higher than that of beta-carotene . These results suggest that orientation of the carotenoids in lipid layers of micelles and also in phospholipid membrane of bacteria is important for quenching of singlet oxygen . Furthermore, the viability of transformants producing lycopene and phytoene was almost as high as that of the transformant producing zeaxanthin-glucoside.

J Biol Chem, 2001 Mar 16, 276(11), 8225 - 30 Epub 2000 Nov 16.
Identification of the human mitochondrial oxodicarboxylate carrier . Bacterial expression, reconstitution, functional characterization, tissue distribution, and chromosomal location; Fiermonte G et al.; In Saccharomyces cerevisiae, the genes ODC1 and ODC2 encode isoforms of the oxodicarboxylate carrier . They both transport C5-C7 oxodicarboxylates across the inner membranes of mitochondria and are members of the family of mitochondrial carrier proteins . Orthologs are encoded in the genomes of Caenorhabditis elegans and Drosophila melanogaster, and a human expressed sequence tag (EST) encodes part of a closely related protein . Information from the EST has been used to complete the human cDNA sequence . This sequence has been used to map the gene to chromosome 14q11.2 and to show that the gene is expressed in all tissues that were examined . The human protein was produced by overexpression in Escherichia coli, purified, and reconstituted into phospholipid vesicles . It has similar transport characteristics to the yeast oxodicarboxylate carrier proteins (ODCs) . Both the human and yeast ODCs catalyzed the transport of the oxodicarboxylates 2-oxoadipate and 2-oxoglutarate by a counter-exchange mechanism . Adipate, glutarate, and to a lesser extent, pimelate, 2-oxopimelate, 2-aminoadipate, oxaloacetate, and citrate were also transported by the human ODC . The main differences between the human and yeast ODCs are that 2-aminoadipate is transported by the former but not by the latter, whereas malate is transported by the yeast ODCs but not by the human ortholog . In mammals, 2-oxoadipate is a common intermediate in the catabolism of lysine, tryptophan, and hydroxylysine . It is transported from the cytoplasm into mitochondria where it is converted into acetyl-CoA . Defects in human ODC are likely to be a cause of 2-oxoadipate acidemia, an inborn error of metabolism of lysine, tryptophan, and hydroxylysine.

Microb Ecol, 2000 Aug, 40(3), 200 - 208
Bacterial Growth on Photochemically Transformed Leachates from Aquatic and Terrestrial Primary Producers; Anesio AM et al.; We measured bacterial growth on phototransformed dissolved organic matter (DOM) leached from eight different primary producers . Leachates (10 mg C liter(-1)) were exposed to artificial UVA + UVB radiation, or kept in darkness, for 20 h . DOM solutions were subsequently inoculated with lake water bacteria . Photoproduction of dissolved inorganic carbon (DIC), ranging from 3 to 16 microg C liter(-1) h(-1), and changes in the absorptive characteristics of the DOM were observed for all leachates upon UV irradiation . The effects of irradiation exposure on DOM bioavailability varied greatly, depending on leachate and type of bacterial growth criterion . Bacterial carbon utilization (biomass production plus respiration) over the entire incubation period (120 h) was enhanced by UV radiation of leachate from the terrestrial leaves, relative to carbon utilization in non-irradiated leachates . Conversely, carbon utilization was reduced by radiation of the leachates from aquatic macrophytes . In a separate experiment, the stable C and N isotope composition of bacteria grown on irradiated and non-irradiated DOM was estimated . Bacterial growth on UV-irradiated DOM was enriched in (13)C relative to the bacteria in the non-irradiated treatments; this result may be explained by selective assimilation of photochemically produced, isotopically enriched labile compounds.

Proc Natl Acad Sci U S A, 2000 Nov 21, 97(24), 13086 - 91
Identification of the proton pathway in bacterial reaction centers: both protons associated with reduction of QB to QBH2 share a common entry point; Adelroth P et al.; The reaction center from Rhodobacter sphaeroides uses light energy for the reduction and protonation of a quinone molecule, Q(B) . This process involves the transfer of two protons from the aqueous solution to the protein-bound Q(B) molecule . The second proton, H(+)(2), is supplied to Q(B) by Glu-L212, an internal residue protonated in response to formation of Q(A)(-) and Q(B)(-) . In this work, the pathway for H(+)(2) to Glu-L212 was studied by measuring the effects of divalent metal ion binding on the protonation of Glu-L212, which was assayed by two types of processes . One was proton uptake from solution after the one-electron reduction of Q(A) (DQ(A)-->D(+)Q(A)(-)) and Q(B) (DQ(B)-->D(+)Q(B)(-)), studied by using pH-sensitive dyes . The other was the electron transfer k(AB)((1)) (Q(A)(-)Q(B)-->Q(A)Q(B)(-)) . At pH 8.5, binding of Zn(2+), Cd(2+), or Ni(2+) reduced the rates of proton uptake upon Q(A)(-) and Q(B)(-) formation as well as k(AB)((1)) by approximately an order of magnitude, resulting in similar final values, indicating that there is a common rate-limiting step . Because D(+)Q(A)(-) is formed 10(5)-fold faster than the induced proton uptake, the observed rate decrease must be caused by an inhibition of the proton transfer . The Glu-L212-->Gln mutant reaction centers displayed greatly reduced amplitudes of proton uptake and exhibited no changes in rates of proton uptake or electron transfer upon Zn(2+) binding . Therefore, metal binding specifically decreased the rate of proton transfer to Glu-L212, because the observed rates were decreased only when proton uptake by Glu-L212 was required . The entry point for the second proton H(+)(2) was thus identified to be the same as for the first proton H(+)(1), close to the metal binding region Asp-H124, His-H126, and His-H128.

Probl Tuberk, 2000, (5), 47 - 9
{The problems of the PCR analysis of low-bacterial-count tissue samples in extrapulmonary tuberculosis}; Vishnevskaia EB; PCR was used to study clinical materials from patients with extrapulmonary tuberculosis or diseases of nontuberculous etiology (renal tissue, scrapes of the endometrium and uterine cavity, bony fragments) . It was shown that when the oligobacillary material analyzed, a concurrent study of some fragments of the sample largely enhanced the efficiency of PCR detection of Mycobacterium tuberculosis DNA . For the biopsy and surgery tissue specimens, PCR provided the best results in isolating DNA for the alkaline lysis test.

FEMS Microbiol Rev, 2000 Dec, 24(5), 647 - 60
Ecology and evolution of bacterial microdiversity; Schloter M et al.; Using high resolution molecular fingerprinting techniques like random amplification of polymorphic DNA, repetitive extragenic palindromic PCR and multilocus enzyme electrophoresis, a high bacterial diversity below the species and subspecies level (microdiversity) is revealed . It became apparent that bacteria of a certain species living in close association with different plants either as associated rhizosphere bacteria or as plant pathogens or symbiotic organisms, typically reflect this relationship in their genetic relatedness . The strain composition within a population of soil bacterial species at a given field site, which can be identified by these high resolution fingerprinting techniques, was markedly influenced by soil management and soil features . The observed bacterial microdiversity reflected the conditions of the habitat, which select for better adapted forms . In addition, influences of spatial separation on specific groupings of bacteria were found, which argue for the occurrence of isolated microevolution . In this review, examples are presented of bacterial microdiversity as influenced by different ecological factors, with the main emphasis on bacteria from the natural environment . In addition, information available from some of the first complete genome sequences of bacteria (Helicobacter pylori and Escherichia coli) was used to highlight possible mechanisms of molecular evolution through which mutations are created; these include mutator enzymes . Definitions of bacterial species and subspecies ranks are discussed in the light of detailed information from whole genome typing approaches.

Plant Sci, 2000 Nov 6, 159(2), 233 - 242
Enhanced tolerance of transgenic Brassica juncea to choline confirms successful expression of the bacterial codA gene; Prasad KV et al.; Brassica juncea cv . Pusa Jaikisan was transformed with the codA gene for choline oxidase from Arthrobacter globiformis with an aim to introduce glycine betaine biosynthetic pathway, as it lacks any means to synthesize glycine betaine . Western blot analysis revealed the presence of choline oxidase in the protein extract from the codA transgenic lines, demonstrating that the bacterial codA gene had been successfully transcribed and translated in transgenic lines . Good activity of choline oxidase indicated its presence in fully functional form in the transformed lines . This was further confirmed by the presence of glycine betaine only in the transformed lines of B . juncea . The shoots of both wild type and transformed lines were exposed to various concentrations of choline in order to evaluate if the introduction of the codA gene in any way enhances the potential of B . juncea to tolerate high levels of choline . The growth (in terms of fresh weight and dry weight) of the shoots of transformed lines exposed to high levels of choline was significantly superior to those of wild type . Moreover, the loss in chlorophyll content and the activity of photosystem II in shoots of the transformed lines exposed to high concentration of choline were significantly lower than that observed in wild type . These results showed that shoots of B . juncea transformed with the codA gene, most probably had the potential to readily convert choline to glycine betaine . Therefore, choline tolerance can be used as an efficient marker for the identification of the lines transformed with the codA gene.

J Virol, 2000 Dec, 74(23), 11088 - 98
Reconstitution of Marek's disease virus serotype 1 (MDV-1) from DNA cloned as a bacterial artificial chromosome and characterization of a glycoprotein B-negative MDV-1 mutant; Schumacher D et al.; The complete genome of Marek's disease virus serotype 1 (MDV-1) strain 584Ap80C was cloned in Escherichia coli as a bacterial artificial chromosome (BAC) . BAC vector sequences were introduced into the U(S)2 locus of the MDV-1 genome by homologous recombination . Viral DNA containing the BAC vector was used to transform Escherichia coli strain DH10B, and several colonies harboring the complete MDV-1 genome as an F plasmid (MDV-1 BACs) were identified . DNA from various MDV-1 BACs was transfected into chicken embryo fibroblasts, and from 3 days after transfection, infectious MDV-1 was obtained . Growth of MDV-1 recovered from BACs was indistinguishable from that of the parental virus, as assessed by plaque formation and determination of growth curves . In one of the MDV-1 BAC clones, sequences encoding glycoprotein B (gB) were deleted by one-step mutagenesis using a linear DNA fragment amplified by PCR . Mutant MDV-1 recovered after transfection of BAC DNA that harbored a 2.0-kbp deletion of the 2.6-kbp gB gene were able to grow and induce MDV-1-specific plaques only on cells providing MDV-1 gB in trans . The gB-negative virus reported here represents the first MDV-1 mutant with a deletion of an essential gene and demonstrates the power and usefulness of BACs to analyze genes and gene products in slowly growing and strictly cell-associated herpesviruses.

Microbiology, 2000 Nov, 146 ( Pt 11), 2845 - 54
Comparative sequence analyses reveal frequent occurrence of short segments containing an abnormally high number of non-random base variations in bacterial rRNA genes; Wang Y et al.; rRNA genes are thought unlikely to be laterally transferred, because rRNA must coevolve with a large number of cellular components to form the highly sophisticated translation apparatus and perform protein synthesis . In this paper, the authors first hypothesized that lateral gene transfer (LGT) might occur to rRNA genes via replacement of gene segments encoding individual domains of rRNA: the 'simplified complexity hypothesis' . Comparative sequence analyses of the 16S and 23S rRNA genes from a large number of actinomycete species frequently identified rRNA genes containing short segments with an abnormally high number of non-random base variations . These variations were nearly always characterized by complementing covariations of several paired bases within the stem of a hairpin . The nature of these base variations is not consistent with random mutations but satisfies well the predictions of the 'simplified complexity hypothesis' . The most parsimonious explanation for this phenomenon is the lateral transfer of rRNA gene segments between different bacterial species . This mode of LGT may create mosaic rRNA genes and occur repeatedly in different regions of a gene, gradually destroying the evolutionary history recorded in the nucleotide sequence.

Nucleic Acids Res, 2000 Nov 1, 28(21), 4397 - 402
RecA-independent ectopic transposition in vivo of a bacterial group II intron; Martinez-Abarca F et al.; RmInt1 is a group II intron of Sinorhizobium meliloti which was initially found within the insertion sequence ISRm2011-2 . Although the RmInt1 intron-encoded protein lacks a recognizable endonuclease domain, it is able to mediate insertion of RmInt1 at an intron-specific location in intronless ISRm2011-2 recipient DNA, a phenomenon termed homing . Here we have characterized three additional insertion sites of RmInt1 in the genome of S.meliloti . Two of these sites are within IS elements closely related to ISRm2011-2, which appear to form a characteristic group within the IS630-Tc1 family . The third site is in the oxi1 gene, which encodes a putative oxide reductase . The newly identified integration sites contain conserved intron-binding site (IBS1 and IBS2) and delta' sequences (14 bp) . The RNA of the intron-containing oxi1 gene is able to splice and the oxi1 site is a DNA target for RmInt1 transposition in vivo . Ectopic transposition of RmInt1 into the oxi1 gene occurs at 20-fold lower efficiency than into the homing site (ISRm2011-2) and is independent of the major RecA recombination pathway . The possibility that transposition of RmInt1 to the oxi1 site occurs by reverse splicing into DNA is discussed.

Nat Cell Biol, 2000 Nov, 2(11), 848 - 51
Glycine 384 is required for presenilin-1 function and is conserved in bacterial polytopic aspartyl proteases; Steiner H et al.; Endoproteolysis of beta-amyloid precursor protein (betaAPP) and Notch requires conserved aspartate residues in presenilins 1 and 2 (PS1 and PS2) . Although PS1 and PS2 have therefore been proposed to be aspartyl proteases, no homology to other aspartyl proteases has been found . Here we identify homology between the presenilin active site and polytopic aspartyl proteases of bacterial origin, thus supporting the hypothesis that presenilins are novel aspartyl proteases.

Nat Cell Biol, 2000 Nov, 2(11), 792 - 6
Molecular model of a lattice of signalling proteins involved in bacterial chemotaxis; Shimizu TS et al.; Coliform bacteria detect chemical attractants by means of a membrane-associated cluster of receptors and signalling molecules . We have used recently determined molecular structures, in conjunction with plastic models generated by three-dimensional printer technology, to predict how the proteins of the complex are arranged in relation to the plasma membrane . The proposed structure is a regular two-dimensional lattice in which the cytoplasmic ends of chemotactic-receptor dimers are inserted into a hexagonal array of CheA and CheW molecules . This structure creates separate compartments for adaptation and downstream signalling, and indicates a possible basis for the spread of activity within the cluster.

Biosci Biotechnol Biochem, 2000 Sep, 64(9), 2008 - 11
Application of a metal switch to aqualysin I, a subtilisin-type bacterial serine protease, to the S3 site residues, ser102 and gly131; Tanaka T et al.; We applied 'metal switch' experiments to the S3 site residues, Ser102 and Gly131, of aqualysin I, a subtilisin-type serine protease . We showed that two histidines introduced at these positions did take part in histidine-metal-histidine bridge formation, and metal ions inhibited the protease activities . These results indicate that two histidines are near each other, and both side chains are metal-accessible . This is the first report on application of the metal-switch technique to a subtilisin-related enzyme.

Acta Crystallogr D Biol Crystallogr, 2000 Nov, 56 ( Pt 11), 1462 - 3
Crystallization and preliminary X-ray analysis of a bacterial lysozyme produced by Streptomyces globisporus; Shiba T et al.; The extracellular bacteriolytic enzyme produced by Streptomyces globisporus shows a beta-1,4-N,6-O-diacetylmuramidase activity as well as a beta-1,4-N-acetylmuramidase activity . Crystals of this enzyme have been obtained by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant . They belong to the tetragonal space group P4(1)2(1)2, with unit-cell parameters a = 63.11 (4), c = 121.1 (1) A, diffract to at least 2.0 A resolution and are suitable for high-resolution structure analysis . The crystal structure was solved by molecular replacement using lysozyme produced by S . erythraeus as a search model . The structure refinement is now in progress.

Klin Med (Mosk), 2000, 78(9), 35 - 40
{Some pathogenetic mechanisms of bacterial bronchial asthma in paecilomycosis}; Akhunova AM; 2-week to 18 year follow-up data are analysed for 190 patients aged 3 to 70 years (91 females and 99 males) with bronchial asthma in Paecilomyses infection . Paecilomyces fungi play the role of a specific agent responsible for development of bacterial asthma . The bronchial spasm was provoked, on the one side, by hypersensitivity with participation of IgE defending the organism from the fulgi persisting in the blood and pulmonary tissue, on the other side, by biologically active substances produced by fungal cells, exogenic phospholipase A2, in particular . Clinical characteristics, course and outcome of bacterial asthma in Paecilomyces infection depend on the resistance of immune system to hematogenic fungal infection and immune reactions running in elimination of the agent.

Rev Med Chil, 2000 Jul, 128(7), 767 - 71
{Evaluation of Nugent and Amsel criteria for the diagnosis of bacterial vaginosis}; Navarrete P et al.; BACKGROUND: Bacterial vaginosis (BV) is a common disease in reproductive-age women and is associated to important gynecologic and obstetric complications . AIM: To study the occurrence of BV in apparently healthy women attending family planning clinics, using Amsel and Nugent diagnostic criteria . MATERIAL AND METHODS: Two hundred thirty nine women consulting for symptoms associated to cervicovaginitis, were studied . A sample from the lateral walls of the vagina was obtained with a sterile swab for microscopic analysis, Gram stain and amine test . RESULTS: According to Amsel and Nugent criteria a 31.1% and 31.8% BV prevalence was observed . The sensitivity and specificity of Nugent criteria, compared with Amsel criteria were 83.3% and 92.1%, respectively . CONCLUSIONS: The high prevalence of BV found in this study suggests that this vaginal infection should be diagnosed with standardized methods . Nugent criteria are economic easy to perform and sensitive and we propose that they should be used in local health centers.

Biochemistry (Mosc), 2000 Sep, 65(9), 1082 - 90
Interaction of bacterial endotoxins with chitosan . Effect of endotoxin structure, chitosan molecular mass, and ionic strength of the solution on the formation of the complex; Davydova VN et al.; The interaction of endotoxins of different structure (lipopolysaccharides (LPS) and lipopolysaccharide-protein complexes (LPPC)) with chitosan has been studied . It was shown that the mechanism of interaction is rather complicated and depends on the macromolecular organization of endotoxin as well as on the degree of polymerization of the chitosan . Chitosan with molecular mass of 20 kD reveals higher affinity to LPS than chitosan with molecular mass of 140 kD . Endotoxins with long O-specific chains can bind completely with chitosan with the formation of LPS-chitosan and LPPC-chitosan complexes with weight ratios between the original components of 1:1 and 1:5 . When endotoxins with higher degree of hydrophobicity and short O-specific chains were mixed with chitosan, a part of the LPS remained unbound . The stability of the complexes formed depends on ionic strength . It was shown that, in addition to electrostatic forces, other types of forces take part in the formation of the complexes . A decrease in acute toxicity of various LPSs is observed on their binding with chitosans.

Biofizika, 2000 Jul-Aug, 45(4), 660 - 5
{Comparative effectiveness of antioxidants in protecting the bacterial plasmatic membrane from the active froms of oxygen}; Ivanov AIu et al.; The effect of hydroxyl radicals OH . generated by the decomposition of H2O2 by Fe2+ ions (Fenton reaction) on the barrier properties of plasma membranes of Escherichia coli cells K-12 was studied by electroorientation spectroscopy . It was found that the administration of hydrogen peroxide led to the disturbance of the barrier properties of plasma membranes only when the cells were preincubated with Fe2+ ions and their constant concentration in the system was maintained by ascorbate or dithiotreitol (150-500 microM) . The extent of the toxic action on plasma membranes depended on the concentration of reacting elements and the substance used as a reducer Fe2+ . The efficiency of protection of antioxidants of different classes (enzymic, SH-containing, and phenolic compounds) against the toxic action of hydroxyl radicals on plasmatic membranes was shown.

Biochim Biophys Acta, 2000 Oct 18, 1502(2), 247 - 56
DMSO and glycerol reduce bacterial death induced by expression of truncated N-terminal huntingtin with expanded polyglutamine tracts; Nagao Y et al.; Huntington's disease (HD) is caused by CAG repeat expansion in exon 1 of a large gene, IT15, possessing 67 exons . Transgenic mice expressing a truncated N-terminal peptide of huntingtin with an expanded polyglutamine tract translated only from exon 1 develop symptoms similar to Huntington's disease . In the present study, a bacterial system (Escherichia coli) was used to express truncated peptides of huntingtin translated from exon 1 of the HD gene . Bacterial death was observed after the induction of peptides with expanded polyglutamine tracts, and both sodium dodecyl sulfate (SDS)-soluble peptides and insoluble aggregated material were detected by immunoblotting in the homogenates of such E . coli . E . coli death was partially reduced by the addition of dimethylsulfoxide (DMSO) or glycerol to the medium, with a consequent decrease in aggregated material and an increase in SDS-soluble peptide in the homogenate . These results suggest that DMSO and glycerol may decrease the toxicity of huntingtin with expanded polyglutamine tracts by acting as chemical chaperones.

Arthritis Rheum, 2000 Oct, 43(10), 2283 - 9
The major role of macrophages and their product tumor necrosis factor alpha in the induction of arthritis triggered by bacterial DNA containing CpG motifs; Deng GM et al.; OBJECTIVE: To understand the mechanisms of arthritis triggered by CpG-containing oligonucleotides (ODN) . METHODS: Following the induction of CpG ODN-triggered arthritis in mice, we analyzed the impact of depletion of immune cells, including neutrophils, natural killer (NK) cells, and monocyte/macrophages, on the arthritis, as well as the impact in SCID mice lacking T and B cells . In addition, tumor necrosis factor alpha (TNFalpha) knockout mice were studied, and intraarticular administration of p65 antisense to nuclear factor kappaB (NF-kappaB) was used to examine effects in CpG ODN-triggered arthritis . Cytokine messenger RNA expression in synovial tissue was evaluated by in situ hybridization . RESULTS: The presence of macrophages was mandatory for the mediation of arthritis triggered by CpG ODN, whereas the absence of neutrophils, NK cells, T cells, and B cells was of minor importance in this regard . The proinflammatory cytokines TNFalpha, interleukin-1beta, and interleukin-12, which originate from macrophages, were frequently found in the inflamed joints, and TNFalpha was confirmed to be an important mediator in the development of arthritis, since the incidence and severity of joint inflammation were markedly reduced in TNFalpha knockout mice . NF-kappaB exerted an important regulatory role in the development of CpG ODN-mediated arthritis, since local administration of antisense to the p65 subunit of NF-kappaB diminished the incidence of inflammation by 50% . CONCLUSION: Macrophages and their products play an important role in the development of arthritis triggered by bacterial DNA containing CpG motifs.

Protein Expr Purif, 2000 Oct, 20(1), 98 - 104
Development of a recombinant bacterial expression system for the active form of a human transforming growth factor beta type II receptor ligand binding domain; Boesen CC et al.; Expression systems have been designed to test the suitability of expressing the high cysteine containing extracellular domain (residues 1-136) of human transforming growth factor beta type II receptor (TbetaRII) . Receptor expressed using a baculovirus system was functional following both enzymatic deglycosylation and elimination of the N-terminal 22 amino acids by protease degradation . Bacterial expression of a TbetaRII lacking the 26 N-terminal amino acids retained the ability to bind its ligand, TGF-beta1 . Receptor expressed in bacteria was sensitive to proteolytic degradation at residue Lys98 but a K98T mutation eliminated degradation and did not disrupt binding . Although several different forms of TbetaRII were expressed, only a fusion with glutathione S-transferase gave soluble TbetaRII, which was purified at a yield of 0.1 mg/10 L of bacterial growth . N-Terminal truncations of TbetaRII (residues 22-136 or 27-136) could be refolded from inclusion bodies and purified to an active form with an efficiency of 10% .

Curr Microbiol, 2000 Mar, 40(3), 190 - 3
Formation of aniline as a transient metabolite during the metabolism of tetryl by a sulfate-reducing bacterial consortium; Boopathy R; A laboratory study was conducted to determine whether tetryl (2,4,6-trinitrophenylmethylnitramine) can be degraded by an anaerobic process . The results indicated that the metabolic conversion of tetryl to aniline is possible by a sulfate-reducing bacterial (SRB) consortium . This SRB consortium metabolized tetryl by co-metabolism with pyruvate as a growth substrate . For every mole of tetryl metabolized, 1 mole of aniline was produced, and the aniline was further metabolized . This metabolic conversion of tetryl is likely to be of value in the anaerobic treatment of tetryl-contaminated soil and ground water, such as found at many military ammunition sites.

J Acquir Immune Defic Syndr, 2000 Aug 15, 24(5), 483 - 7
HTLV-II and bacterial infections among injection drug users; Safaeian M et al.; OBJECTIVE: To examine whether select bacterial infections are associated with HTLV-II infection among injection drug users, we conducted a nested case control study within an ongoing cohort study . METHOD: HTLV-II status was determined by enzyme-linked immunosorbent assay, immunofluorescent assay, and immunoblot . Diagnosis of bacterial pneumonia, infective endocarditis, and skin abscess was confirmed by standardized chart reviews . Three sets of cases were identified based on diagnosis of bacterial pneumonia, infective endocarditis validated by chart review, or self-reported skin abscess . Each case was matched to a minimum of 5 controls by age, HIV status, and study follow-up duration . Risk factors for each bacterial infection were analyzed separately by conditional logistic regression methods . RESULTS: Prevalence of HTLV-II infection ranged from 7% to 11% in cases and controls . The bivariate association of HTLV-II and bacterial pneumonia revealed an odds ratio (OR) of 1.1 (95% confidence interval {CI}, 0.6-2.0); the association of infective endocarditis and HTLV-II revealed an OR of 1.7 (95% CI, 0 . 7-3.9); and the association between HTLV-II and skin abscess revealed an OR of 1.3 (95% CI, 0.6-2.0) . These ORs were unaltered by adjustment for other factors . CONCLUSION: Our results suggest that these three bacterial infections were not significantly associated with HTLV-II infection within a population of injection drug users . Additional associations between HTLV-II infection and disease outcomes merit further exploration.

Int J Syst Evol Microbiol, 2000 Sep, 50 Pt 5, 1877 - 86
Systematic relationships and cospeciation of bacterial endosymbionts and their carpenter ant host species: proposal of the new taxon Candidatus Blochmannia gen . nov; Sauer C et al.; The systematic relationships of intracellular bacteria of 13 Camponotus species (carpenter ants) from America and Europe were compared to those of their hosts . Phylogenetic trees of the bacteria and the ants were based on 16S rDNA (rrs) gene sequences and mitochondrial cytochrome oxidase subunit I (COI) gene sequences, respectively . The bacterial endosymbionts of Camponotus spp . form a distinct lineage in the y-subclass of the Proteobacteria . The taxa most closely related to these bacteria are endosymbionts of aphids and the tsetse fly . The bacterial and host phylogenies deduced from the sequence data show a high degree of congruence, providing significant evidence for cospeciation of the bacteria and the ants and a maternal transmission route of the symbionts . The cloned rrs genes of the endosymbionts contain putative intervening sequences (IVSs) with a much lower G+C content than the mean of the respective rrs genes . By in situ hybridization specific 16S rDNA oligonucleotide probes verified the presence of the bacteria within tissues of three of the eukaryotic hosts . It is proposed that the endosymbionts of these three carpenter ants be assigned to a new taxon 'Candidatus Blochmannia gen . nov.' with the symbionts of the individual ants being species named according to their host, 'Candidatus Blochmannia floridanus sp . nov.', 'Candidatus Blochmannia herculeanus sp . nov.' and 'Candidatus Blochmannia rufipes sp . nov.'.

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