Vet Microbiol, 2000 Jul 31, 75(2), 167 - 75 A comparison of four serological tests for the diagnosis of caseous lymphadenitis in sheep and goats; Dercksen DP et al.; A double antibody sandwich ELISA (ELISA A) developed for the detection of Corynebacterium pseudotuberculosis infection in sheep and goats was modified to improve its sensitivity . To establish the sensitivity and specificity of this modified ELISA (ELISA B), sera from 183 sheep and 186 goats were tested using ELISAs A and B . Comparison was also made with two further ELISAs (C and D) developed in Australia that, respectively, detect antibodies to cell wall antigens or toxin.ELISA B had the best performance of the four tests . Its specificity was 98+/-1% for goats and 99+/-1% sheep . Its sensitivity was 94+/-3% for goats and 79+/-5% for sheep . ELISA B will now be tested for use in caseous lymphadenitis eradication and control programmes in The Netherlands . It will also be used in experimental studies of CL in Scotland. Commun Dis Rep CDR Rev, 1994 Apr 29, 4(5), R63 - 4 'Corynebacterium ulcerans': a potential cause of diphtheria; Kisely SR et al.; Symptoms similar to those of diphtheria may occasionally be caused by micro-organisms other than Corynebacterium diphtheriae, such as zoonotic corynebacteria . A case of pharyngitis caused by toxigenic 'C . ulcerans' in a 9 year old girl illustrates the importance of including diphtheroids in the differential diagnosis . The presenting symptoms may have been attenuated by previous immunisation against C . diphtheriae . 'C . ulcerans' should be considered in the differential diagnosis of pharyngitis, especially in patients with a history of contact with farm animals or the consumption of raw dairy products. J Med Microbiol, 2000 Jul, 49(7), 621 - 5 Expression of trypsin-like activity by the genera Corynebacterium and Actinomyces in canine periodontitis; Takada K et al.; Trypsin-like activity (TLA), clinical parameters and TLA-positive bacteria were examined in periodontitis and healthy sites in dogs . TLA was markedly higher in periodontitis than at healthy sites . There was good correlation between TLA positivity and severity of periodontal disease . The proportions of TLA-positive bacteria to total isolates in periodontitis and healthy sites were 21.1% and 2.1%, respectively . Among TLA-positive bacteria in periodontitis sites, 4.4% showed strong TLA activity, 35.3% showed moderate and 60.3% showed weak activity . In the healthy sites, all the TLA-positive bacteria showed weak activity . In all, 90% of the total number of TLA-positive bacteria were identified as belonging to the family Actinomycetaceae; 40% of bacteria belonging to the family Actinomycetaceae were identified as genus Corynebacterium with moderate trypsin-like activity and the remaining 60% were identified as genus Actinomyces with weak activity . Obligately anaerobic bacteria accounted for only 5.9% of the total population of TLA-positive bacteria; they were gram-negative coccobacilli, gram-positive rods and gram-positive cocci . These observations suggested that bacteria in the family Actinomycetaceae may play an important role in periodontitis and that measurement of TLA is a clinically reliable marker for the diagnosis of periodontitis in dogs. Syst Appl Microbiol, 2000 Apr, 23(1), 15 - 24 Characterization of the ribosomal rrnD operon of the cephamycin-producer 'Nocardia lactamdurans' shows that this actinomycete belongs to the genus Amycolatopsis; Barreiro C et al.; The cephamycin producer strain 'Nocardia lactamdurans' contains four ribosomal RNA (rrn) operons . One of them (rrnD) was cloned from a DNA library in the bifunctional cosmid pJAR4 . A 2229 bp region of rrnD has been sequenced . The 'N . lactamdurans' rrnD operon maintains the canonical order 5'-16S-23S-5S-3' . Four of the consensus Gurtler-Stanisch sequences were found in the 16S rRNA gene and a fifth one in the sequenced 5' region of the 23S rRNA gene . The anti Shine-Dalgarno sequence of 'N . lactamdurans' (located in the 3'-end of the 16S rRNA gene) was found to be 5'-CCUCCUUUCU-3' and is identical to that of Corynebacterium lactofermentum and Mycobacterium tuberculosis . A phylogenetic analysis of 'N . lactamdurans' by the neighbor-joining method using the entire 16S rRNA nucleotide sequence revealed that this actinomycete is closely related to Amlycolatopsis orientalis subsp orientalis, Amycolatopsis coloradensis, Amycolatopsis alba, Amycolatopsis sulphurea and other Amycolatopsis sp . but only distantly related to species of the genus Nocardia . The cephamycin producer 'N . lactamdurans' NRRL 3802 should be, therefore, classified as Amycolatopsis lactamdurans . The deduced secondary structure of the 16S rRNA is very similar to that of A . colorandensis and A . alba but different from those of species of the Nocardia genus supporting the incorporation of 'N . lactamdurans' into the genus Amycolatopsis. Biosci Biotechnol Biochem, 2000 May, 64(5), 1067 - 70 Purification of alginate oligosaccharides with root growth-promoting activity toward lettuce; Iwasaki K et al.; Sodium alginate was degraded by alginate lyase from Corynebacterium sp., and the product was purified by an ultrafiltration (UF) membrane module . The UF treatment was carried out at a transmembrane pressure of 0.15 MPa and a flow velocity of 0.6 m/s in the cross-flow mode, and non-degraded alginate was almost completely removed . The alginate oligosaccharide obtained was a mixture of di- to octasaccharides and had promoting activity toward lettuce root elongation (about 2-fold compared with the control) in the concentration range of 200-3000 microg/ml . The effect of the degree of polymerization on this activity was examined by using each oligosaccharide fractionated by gel chromatography . The tri-, tetra-, penta- and hexasaccharides were each found to have root growth-promoting activity in a lettuce bioassay. J Clin Microbiol, 2000 Jul, 38(7), 2633 - 7 Characterization of United Kingdom isolates of Corynebacterium pseudotuberculosis using pulsed-field gel electrophoresis; Connor KM et al.; Caseous lymphadenitis is a chronic suppurative disease caused by Corynebacterium pseudotuberculosis and is responsible for serious economic losses to the sheep and goat industry . Caseous lymphadenitis was first reported for goats in the United Kingdom in 1990 and for sheep in 1991 . Recent evidence suggests that the prevalence of the disease within the national flock is increasing . Fifty isolates of C . pseudotuberculosis from the United Kingdom comprising sheep and horse isolates, the original goat outbreak strain, and the type strain were characterized by biotyping, antimicrobial susceptibility, production of phospholipase D, and genotyping by pulsed-field gel electrophoresis using SfiI and SmaI . All of the isolates were confirmed as C . pseudotuberculosis, and all produced phospholipase D but none reduced nitrate . Restriction with SfiI generated 16 to 18 bands between 48.5 and 290 kb and differentiated six pulsotypes . We conclude that 80% of the strains tested were epidemiologically related to the outbreak strain and that the equine profile was distinct both phenotypically and genotypically. Appl Environ Microbiol, 2000 Jul, 66(7), 2981 - 7 Cloning of the malic enzyme gene from Corynebacterium glutamicum and role of the enzyme in lactate metabolism; Gourdon P et al.; Malic enzyme is one of at least five enzymes, known to be present in Corynebacterium glutamicum, capable of carboxylation and decarboxylation reactions coupling glycolysis and the tricarboxylic acid cycle . To date, no information is available concerning the physiological role of the malic enzyme in this bacterium . The malE gene from C . glutamicum has been cloned and sequenced . The protein encoded by this gene has been purified to homogeneity, and the biochemical properties have been established . Biochemical characteristics indicate a decarboxylation role linked to NADPH generation . Strains of C . glutamicum in which the malE gene had been disrupted or overexpressed showed no detectable phenotype during growth on either acetate or glucose, but showed a significant modification of growth behavior during lactate metabolism . The wild type showed a characteristic brief period of exponential growth on lactate followed by a linear growth period . This growth pattern was further accentuated in a malE-disrupted strain (Delta malE) . However, the strain overexpressing malE maintained exponential growth until all lactate had been consumed . This strain accumulated significantly larger amounts of pyruvate in the medium than the other strains. Zh Mikrobiol Epidemiol Immunobiol, 2000 Jan-Feb, (1), 27 - 30 {The effect of the nature of the iron sources on the growth of different Corynebacterium diphtheriae strains}; Gavrilova NA et al.; In the study of the influence of organic and inorganic sources of iron on the growth of 5 C . diphtheriae clinical isolates bacterial growth was found to depended on the nature of the source of iron and its concentration . Differences between the strains in the level of growth, observed when ferric sulfate was used as the only source of iron in the medium, were established . Quantitative differences in the concentrations of inorganic and organic sources of iron, necessary for growth, were determined . The influence of three chemical chelators on the growth of C . diphtheriae under the conditions of iron deficiency in the culture medium was studied . The results of the study are indicative of the possibility of the differentiation of C . diphtheriae isolated according to the level of iron consumption. Res Microbiol, 2000 Apr, 151(3), 201 - 8 Isolation and curing of the Klebsiella pneumoniae large indigenous plasmid using sodium dodecyl sulphate; El-Mansi M et al.; The ability of Klebsiella pneumoniae (NCTC, CL687/80) to produce and, in turn, excrete glutamate has been equated with the presence of a large indigenous plasmid with an apparent molecular mass in the region of 96 +/- 2 kbp (n = 6) . Unlike mitomycin C, novobiocine and ethidium bromide (curing agents), the use of sodium dodecyl sulphate (SDS) proved very effective in curing the plasmid with a relatively high frequency (6.25 x 10(-4)) . Furthermore, the absence of isocitrate dehydrogenase (ICDH) activity in the cured strain strongly suggests that the structural gene encoding ICDH in this organism, in sharp contrast to all known ICDHs, is plasmid-encoded . Moreover, the SDS-based protocol reported for the isolation of the K . pneumoniae indigenous plasmid has proven successful with other organisms including Pseudomonad and Corynebacteria, as well as in recombinant strains of Escherichia coli. J Vet Med B Infect Dis Vet Public Health, 2000 May, 47(4), 287 - 93 Effect of shearing on the incidence of caseous lymphadenitis in Awassi sheep in Jordan; al-Rawashdeh OF et al.; A total of 876 sheep from five flocks in north Jordan were selected to study the effect of shearing on the incidence of caseous lymphadenitis (CLA) . The animals were divided into two age groups, sheep aged 1-2 years and those aged > or = 3 years . Blood samples were collected from the animals at the time of shearing and again 6 months later . A toxin enzyme-linked immunosorbent assay was used to identify sheep that had been infected with Corynebacterium pseudotuberculosis . The point prevalences of CLA were 6.59% and 21.06% in the 1-2-year and > or = 3-year age groups, respectively, and were significantly higher (P < 0.01) in the > or = 3-year age group . The overall prevalence among all ages was 15.3% . In the shorn sheep, the incidence of CLA was 22.46% and 9.47% in the 1-2-year and > or = 3-year age groups, respectively, and was significantly higher (P < 0.05) in the 1-2-year age group . In the control animals, the incidence was 8% and 5.26% in the 1-2-year and > or = 3-year age groups, respectively, and was different (P < 0.01) between the shorn (22.46%) and control (8%) animals of the 1-2-year age group . An epidemiological survey of 35 sheep farms revealed the prevalence of CLA, shearing wounds and unhygienic conditions during shearing in all farms . In conclusion, the prevalence of CLA increases with age and the incidence increases only in young sheep after shearing . Sheep are sheared under unhygienic conditions, which may be a contributing factor in increasing both the prevalence and the incidence of CLA. Clin Lab, 2000, 46(5-6), 255 - 60 A closer look at the insertion within helix 54 of the 23S-rRNA of the genus Corynebacterium (and related taxa); Hugle B et al.; Direct sequencing of the insertion in helix 54 of the 23S-rRNA of different strains of 8 species of the genus Corynebacterium was performed in order to determine the extent of variations of these strains . The results demonstrated that there is considerable variation within different strains of a single species . While this clearly jeopardizes the usefulness of the insertion with regard to species-specific diagnostic probes, the variations were found to concentrate within two clearly defined regions comprising the ascending and descending parts of the distal helix of a putative stemlike secondary structure . The remainder of the insertion sequences seem to be fairly constant within a single species while displaying considerable differences toward sequences of other species of the same genus. Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 2000 Jun, 89(6), 744 - 8 Checkerboard DNA-DNA hybridization analysis of endodontic infections; Siqueira JF Jr et al.; OBJECTIVE: The purpose of this investigation was to examine the microbiota of infected root canals by using a molecular genetic method . STUDY DESIGN: The presence and levels of 42 bacterial species were determined in 28 root canal samples by using whole genomic DNA probes and checkerboard DNA-DNA hybridization . To confirm the presence of bacterial DNA in clinical samples, a polymerase chain reaction with an ubiquitous bacterial primer was undertaken . RESULTS: The results of the checkerboard DNA-DNA hybridization analysis showed that 22 of the 42 DNA probes tested were reactive with 1 or more samples . The number of bacterial species in the root canal samples ranged from 1 to 17 (mean, 4.7) . Seventeen of the 28 root canal samples were positive for at least 1 DNA probe . The most prevalent species found were as follows: Bacteroides forsythus (39 . 3% of the cases); Haemophilus aphrophilus (25%); Corynebacterium matruchotii (21.4%); Porphyromonas gingivalis (17.9%); and Treponema denticola (17.9%) . CONCLUSIONS: The microbiologic data of the present investigation indicated that molecular genetic methods can provide significant additional knowledge regarding the endodontic microbiota by detecting bacterial species that are difficult or impossible to culture . In addition, our findings support the current concept that endodontic infections are mixed infections of polymicrobial etiology. Eur J Biochem, 2000 Jun, 267(12), 3442 - 52 Kinetic properties of the glucose-6-phosphate and 6-phosphogluconate dehydrogenases from Corynebacterium glutamicum and their application for predicting pentose phosphate pathway flux in vivo; Moritz B et al.; The glucose-6-phosphate (Glc6P) and 6-phosphogluconate (6PG) dehydrogenases of the amino-acid-producing bacterium Corynebacterium glutamicum were purified to homogeneity and kinetically characterized . The Glc6P dehydrogenase was a heteromultimeric complex, which consists of Zwf and OpcA subunits . The product inhibition pattern of the Glc6P dehydrogenase was consistent with an ordered bi-bi mechanism . The 6PG dehydrogenase was found to operate according to a Theorell-Chance ordered bi-ter mechanism . Both enzymes were inhibited by NADPH and the 6PG dehydrogenase additionally by ATP, fructose 1,6-bisphosphate (Fru1,6P2), D-glyceraldehyde 3-phosphate (Gra3P), erythrose 4-phosphate and ribulose 5-phosphate (Rib5P) . The inhibition by NADPH was considered to be most important, with inhibition constants of around 25 microM for both enzymes . Intracellular metabolite concentrations were determined in two isogenic strains of C . glutamicum with plasmid-encoded NAD- and NADP-dependent glutamate dehydrogenases . NADP+ and NADPH levels were between 130 microM and 290 microM, which is very much higher than the respective Km and Ki values . The Glc6P concentration was around 500 microM in both strains . The in vivo fluxes through the oxidative part of the pentose phosphate pathway calculated on the basis of intracellular metabolite concentrations and the kinetic constants of the purified enzymes determined in vitro were in agreement with the same fluxes determined by NMR after 13C-labelling . From the derived kinetic model thus validated, it is concluded that the oxidative pentose phosphate pathway in C . glutamicum is mainly regulated by the ratio of NADPH and NADP+ concentrations and the specific enzyme activities of both dehydrogenases. J Nat Prod, 2000 May, 63(5), 596 - 8 Indolyl carboxylic acids by condensation of indoles with alpha-keto acids; Garbe TR et al.; The novel indole derivatives 2,2-bis(3,3'-indolyl)propionic acid (1); 1,1,1,-tris(3,3',3"-indolyl)ethane (2); and 2,2-bis(3, 3'-indolyl)isocaproic acid (3) were isolated from solvent extracts of indole-supplemented supernatants of Escherichia coli and corynebacteria . The compounds were also obtained by chemical synthesis: compounds 1 and 2 from indole and pyruvic acid and compound 3 from indole and alpha-ketoisocaproic acid, following incubation at 37 degrees C in aqueous medium . Tryptophan and pyruvic acid gave the novel 2-(2-tryptophanyl)lactic acid (4) . The condensation reaction between indoles and alpha-keto acids was of general nature, and the mild reaction conditions suggested it may proceed in vivo . Examples for endogenous occurrence may be the neuro-degenerative diseases phenylketonuria and maple syrup urine disease, both characterized by elevated blood levels of alpha-keto acids. J Clin Microbiol, 2000 Jun, 38(6), 2400 - 2 Retrospective diagnosis of diphtheria by detection of the Corynebacterium diphtheriae tox gene in a formaldehyde-fixed throat swab using PCR and sequencing analysis; Komiya T et al.; The tox gene of Corynebacterium diphtheriae was detected in a formaldehyde-fixed throat swab taken from a 68-year-old man who was reported to have died of suffocation due to a pharyngeal tumor . DNA templates prepared from bacterial cells fixed with 10% formaldehyde were subjected to a PCR analysis with tox gene-specific PCR primers . The resultant 112-nucleotide-long PCR product was sequenced using a dye terminator method, and an expected 57-nucleotide-long internal sequence of the tox gene was detected . This method is applicable for retrospective diagnosis in diphtheria cases in which only a formaldehyde-fixed clinical sample is available. J Laryngol Otol, 2000 Mar, 114(3), 184 - 8 Microbiology of the middle meatus: a comparison between normal adults and children; Gordts F et al.; Middle meatal samples were obtained from 52 carefully selected healthy adults . In 75 per cent of the test subjects bacterial organisms were cultured . However, growth was often poor and the predominant species suggest a commensal flora: coagulase-negative staphylococci were retrieved from 35 per cent, Corynebacterium sp . from 23 per cent and Staphyloccus aureus from eight per cent of the adults . These data are very different from those previously obtained among children where--even in the absence of obvious ENT pathology--the most frequently cultured organisms were typical sinusitis pathogens: Haemophilus influenzae present in 40 per cent, Moraxella catarrhalis in 34 per cent and Streptococcus pneumoniae in 50 per cent of children . Furthermore, Streptococcus viridans and Neisseria sp., both organisms that might be able to inhibit colonization by some of the pathogens and found commonly among children, are virtually absent in healthy adults. FEMS Microbiol Lett, 2000 Jun 1, 187(1), 83 - 8 Response to nitrogen starvation in Corynebacterium glutamicum; Schmi R et al.; Proteins strongly synthesized in Corynebacterium glutamicum during nitrogen restriction were examined by two-dimensional gel electrophoresis and microsequencing . Two main groups of enzymes were identified beside miscellaneous proteins, enzymes involved (i) in protein synthesis, and (ii) in carbon metabolism . Biochemical measurements revealed an increase of oxygen consumption during nitrogen starvation, indicating an enhanced energy demand of the cells . By Northern hybridizations, an increased transcription for the gap and fda genes upon nitrogen deprivation was shown. Int J Syst Evol Microbiol, 2000 Jan, 50 Pt 1, 371 - 80 Leifsonia poae gen . nov., sp . nov., isolated from nematode galls on Poa annua, and reclassification of 'Corynebacterium aquaticum' Leifson 1962 as Leifsonia aquatica (ex Leifson 1962) gen . nov., nom . rev., comb . nov . and Clavibacter xyli Davis et al . 1984 with two subspecies as Leifsonia xyli (Davis et al . 1984) gen . nov., comb . nov; Evtushenko LI et al.; The new genus Leifsonia gen . nov . with two new species, Leifsonia poae sp . nov . (type strain VKM Ac-1401T) and Leifsonia aquatica (ex Leifson 1962) nom . rev., comb . nov . (the type species, with VKM Ac-1400T = DSM 20146T = JCM 1368T as type strain), is proposed to accommodate bacteria found in Poa annua root gall, induced by the nematode Subanguina radicicola, and 'Corynebacterium aquaticum' Leifson 1962 . Further, it is proposed to reclassify Clavibacter xyli Davis et al . 1984 with two subspecies in the new genus as Leifsonia xyli (Davis et al . 1984) comb . nov., Leifsonia xyli subsp . xyli (Davis et al . 1984) comb . nov . and Leifsonia xyli subsp . cynodontis (Davis et al . 1984) comb . nov . Members of the proposed genus are characterized by coryneform morphology, peptidoglycans based upon 2,4-diaminobutyric acid, the major menaquinone MK-11, phosphatidylglycerol and diphosphatidylglycerol as principal phospholipids, the high content of anteiso- and iso-branched saturated fatty acids, and a DNA G+C base composition of 66-73 mol% . They form a distinct phylogenetic branch attached to the line of descent of Agromyces spp . The new and reclassified species of the new genus clearly differ from each other phylogenetically and phenetically and can be recognized by their morphologies, the cell wall sugar composition, the requirement of complex media for growth, and numerous physiological characteristics, including the oxidase reaction. Int J Syst Evol Microbiol, 2000 Jan, 50 Pt 1, 347 - 53 Corynebacterium simulans sp . nov., a non-lipophilic, fermentative Corynebacterium; Wattiau P et al.; Three coryneform strains isolated from clinical samples were analysed . These strains fitted the biochemical profile of Corynebacterium striatum by conventional methods . However, according to recently described identification tests for fermenting corynebacteria, the strains behaved rather like Corynebacterium minutissimum . The three isolates could be distinguished from C . minutissimum by a positive nitrate and nitrite reductase test and by not fermenting maltose; from C . striatum by their inability to acidify ethylene glycol and to grow at 20 degrees C . Genetic studies based on 16S rRNA showed that the three strains were in fact different from C . minutissimum and C . striatum (96.9 and 98% similarity, respectively) and from other corynebacteria . They represent a new species for which the name Corynebacterium simulans sp . nov . is proposed . The type strain is DSM 44415T (= UCL 553T = Co 553T). J Am Soc Nephrol, 2000 Jun, 11(6), 1138 - 40 Encrusted pyelitis of native kidneys; Hertig A et al.; This study reports the first four cases of encrusted pyelitis involving native kidneys . The clinical features, management, and outcome of these patients were analyzed . Predisposing factors were underlying urologic disease and/or urologic manipulations, debilitating diseases, hospitalization, and prolonged antibiotic therapies . Presenting symptoms were renal failure in three patients with ureteroileal urinary diversion and manifestations of cystitis in one patient . Computed tomography scan of the urinary tract was critical for diagnosis . Presence of struvite was demonstrated by crystalluria and infrared spectrophotometry analysis of the encrusted material . Corynebacterium urealyticum urinary infection was identified in one case . Surgery (one patient) and palliative ureteral diversion (one patient), respectively, led to death and end-stage renal failure . Successful dissolution of encrusted pyelitis was obtained in two patients treated with intravenous vancomycin and local acidification of the renal collecting system . Clinical observation shows that encrusted pyelitis is a threatening disorder that destroys the native kidneys and may lead to end-stage renal failure . Successful treatment of the disease by chemolysis and antibiotics depends on correct and early diagnosis . Diagnosis required recognition of the predisposing factors, computed tomography imaging of the urinary tract, crystalluria, and identification of urea-splitting bacteria with prolonged culture on selective medium. Pediatr Cardiol, 2000 May-Jun, 21(3), 282 - 3 Atrioventricular block in a toxic child: do not forget diphtheria; Perles Z et al.; We describe a 4.5-year-old girl who presented with severe febrile throat infection and who, after a few days, developed ventricular tachycardia followed by atrioventricular block . Although a pacemaker was inserted, she died of cardiogenic shock . Throat cultures were positive for Corynebacterium diphtheriae. J Bacteriol, 2000 Jun, 182(11), 3204 - 9 Another unusual type of citric acid cycle enzyme in Helicobacter pylori: the malate:quinone oxidoreductase; Kather B et al.; The only enzyme of the citric acid cycle for which no open reading frame (ORF) was found in the Helicobacter pylori genome is the NAD-dependent malate dehydrogenase . Here, it is shown that in this organism the oxidation of malate to oxaloacetate is catalyzed by a malate:quinone oxidoreductase (MQO) . This flavin adenine dinucleotide-dependent membrane-associated enzyme donates electrons to quinones of the electron transfer chain . Similar to succinate dehydrogenase, it is part of both the electron transfer chain and the citric acid cycle . MQO activity was demonstrated in isolated membranes of H . pylori . The enzyme is encoded by the ORF HP0086, which is shown by the fact that expression of the HP0086 sequence from a plasmid induces high MQO activity in mqo deletion mutants of Escherichia coli or Corynebacterium glutamicum . Furthermore, this plasmid was able to complement the phenotype of the C . glutamicum mqo deletion mutant . Interestingly, the protein predicted to be encoded by this ORF is only distantly related to known or postulated MQO sequences from other bacteria . The presence of an MQO shown here and the previously demonstrated presence of a 2-ketoglutarate:ferredoxin oxidoreductase and a succinyl-coenzyme A (CoA):acetoacetyl-CoA transferase indicate that H . pylori possesses a complete citric acid cycle, but one which deviates from the standard textbook example in three steps. J Bacteriol, 2000 Jun, 182(11), 3088 - 96 Quantitative determination of metabolic fluxes during coutilization of two carbon sources: comparative analyses with Corynebacterium glutamicum during growth on acetate and/or glucose; Wendisch VF et al.; Growth of Corynebacterium glutamicum on mixtures of the carbon sources glucose and acetate is shown to be distinct from growth on either substrate alone . The organism showed nondiauxic growth on media containing acetate-glucose mixtures and simultaneously metabolized these substrates . Compared to those for growth on acetate or glucose alone, the consumption rates of the individual substrates were reduced during acetate-glucose cometabolism, resulting in similar total carbon consumption rates for the three conditions . By (13)C-labeling experiments with subsequent nuclear magnetic resonance analyses in combination with metabolite balancing, the in vivo activities for pathways or single enzymes in the central metabolism of C . glutamicum were quantified for growth on acetate, on glucose, and on both carbon sources . The activity of the citric acid cycle was high on acetate, intermediate on acetate plus glucose, and low on glucose, corresponding to in vivo activities of citrate synthase of 413, 219, and 111 nmol . (mg of protein)(-1) . min(-1), respectively . The citric acid cycle was replenished by carboxylation of phosphoenolpyruvate (PEP) and/or pyruvate (30 nmol . {mg of protein}(-1) . min(-1)) during growth on glucose . Although levels of PEP carboxylase and pyruvate carboxylase during growth on acetate were similar to those for growth on glucose, anaplerosis occurred solely by the glyoxylate cycle (99 nmol . {mg of protein}(-1) . min(-1)) . Surprisingly, the anaplerotic function was fulfilled completely by the glyoxylate cycle (50 nmol . {mg of protein}(-1) . min(-1)) on glucose plus acetate also . Consistent with the predictions deduced from the metabolic flux analyses, a glyoxylate cycle-deficient mutant of C . glutamicum, constructed by targeted deletion of the isocitrate lyase and malate synthase genes, exhibited impaired growth on acetate-glucose mixtures. Zh Mikrobiol Epidemiol Immunobiol, 2000 Mar-Apr, (2), 100 - 2 {The correction of dysbiosis of the female reproductive tract by using an estradiol-containing preparation}; Bukharin OV et al.; The influence of Ovestin, an estradiol-containing preparation, on the vaginal microflora in women of the reproductive age in dysbiosis was studied . As the result of the local application of this preparation, the normalization of microflora was noted in 92% of women, which was manifested by an increase in the number of lactobacteria and indigenous corynebacteria, a decrease in the number, or elimination, of opportunistic microorganisms and the suppression of their capacity for persistence. J Biol Chem, 2000 Aug 18, 275(33), 25365 - 71 The active form of the R2F protein of class Ib ribonucleotide reductase from Corynebacterium ammoniagenes is a diferric protein; Huque Y et al.; Corynebacterium ammoniagenes contains a ribonucleotide reductase (RNR) of the class Ib type . The small subunit (R2F) of the enzyme has been proposed to contain a manganese center instead of the dinuclear iron center, which in other class I RNRs is adjacent to the essential tyrosyl radical . The nrdF gene of C . ammoniagenes, coding for the R2F component, was cloned in an inducible Escherichia coli expression vector and overproduced under three different conditions: in manganese-supplemented medium, in iron-supplemented medium, and in medium without addition of metal ions . A prominent typical tyrosyl radical EPR signal was observed in cells grown in rich medium . Iron-supplemented medium enhanced the amount of tyrosyl radical, whereas cells grown in manganese-supplemented medium had no such radical . In highly purified R2F protein, enzyme activity was found to correlate with tyrosyl radical content, which in turn correlated with iron content . Similar results were obtained for the R2F protein of Salmonella typhimurium class Ib RNR . The UV-visible spectrum of the C . ammoniagenes R2F radical has a sharp 408-nm band . Its EPR signal at g = 2.005 is identical to the signal of S . typhimurium R2F and has a doublet with a splitting of 0.9 millitesla (mT), with additional hyperfine splittings of 0.7 mT . According to X-band EPR at 77-95 K, the inactive manganese form of the C . ammoniagenes R2F has a coupled dinuclear Mn(II) center . Different attempts to chemically oxidize Mn-R2F showed no relation between oxidized manganese and tyrosyl radical formation . Collectively, these results demonstrate that enzymatically active C . ammoniagenes RNR is a generic class Ib enzyme, with a tyrosyl radical and a diferric metal cofactor. J Biochem (Tokyo), 2000 May, 127(5), 813 - 9 Rescuing activity of galactoglycerolipids from cellular lesions induced by 5-aminolevulinic acid; Nakata K; An anti-oxygen radical reagent of a bacterial metabolite, M874 monogalactoglycerolipid (di-O-12-methyl-tetradecanoyl-3-O-beta-D-galactopyranosyl-sn-glycerol ), was tested for its ability to protect two organisms against cellular lesions induced by 5-aminolevulinic acid (ALA) and light . In Corynebacterium flavescens ATCC 10340, extracellular uroporphyrin and coproporphyrin were the main porphyrin products . Although less than 2 mM ALA increased porphyrin synthesis, ALA levels above 3 mM inhibited the synthesis . Depending on the light intensity, the amount of porphyrin decreased and ALA-induced cytotoxicity increased . The lesion was more severe in the case of coproporphyrin than uroporphyrin . The porphyrin lesion produced in low intensity light (300 lx) was considerably reduced by 100 microM M874 glycolipid, although the reduction in intense light (3,000 lx) was restricted to a lower level . Similar results were obtained with radish (Raphanus sativus) . The ALA concentration that inhibited porphyrin synthesis and stem growth was similar to that seen with C . flavescens . Although the exogenous addition of M874 glycolipid to the radish did not prevent ALA-induced cellular injury, the co-culture of radish and a glycolipid producing bacterium (Microbacterium sp . M874) resulted in a significant prevention of cellular injury . This was true only under enforced adhesion conditions through the action of a polysaccharide flocculant H12 . Some species of monogalactoglycerolipids were found in Corynebacterium and radish that showed prominent oxygen radical-protecting activities similar to that of M874 glycolipid . These monogalactoglycerolipids might function in vivo as agents to prevent ALA-induced cytological lesions, although the concentrations were low in Corynebacterium and radish. J Bacteriol, 2000 May, 182(10), 2696 - 701 A mutation in the Corynebacterium glutamicum ltsA gene causes susceptibility to lysozyme, temperature-sensitive growth, and L-glutamate production; Hirasawa T et al.; The Corynebacterium glutamicum mutant KY9714, originally isolated as a lysozyme-sensitive mutant, does not grow at 37 degrees C . Complementation tests and DNA sequencing analysis revealed that a mutation in a single gene of 1,920 bp, ltsA (lysozyme and temperature sensitive), was responsible for its lysozyme sensitivity and temperature sensitivity . The ltsA gene encodes a protein homologous to the glutamine-dependent asparagine synthetases of various organisms, but it could not rescue the asparagine auxotrophy of an Escherichia coli asnA asnB double mutant . Replacement of the N-terminal Cys residue (which is conserved in glutamine-dependent amidotransferases and is essential for enzyme activity) by an Ala residue resulted in the loss of complementation in C . glutamicum . The mutant ltsA gene has an amber mutation, and the disruption of the ltsA gene caused lysozyme and temperature sensitivity similar to that in the KY9714 mutant . L-Glutamate production was induced by elevating growth temperature in the disruptant . These results indicate that the ltsA gene encodes a novel glutamine-dependent amidotransferase that is involved in the mechanisms of formation of rigid cell wall structure and in the L-glutamate production of C . glutamicum. J Vet Med B Infect Dis Vet Public Health, 2000 Feb, 47(1), 55 - 62 Abscess disease, caseous lymphadenitis, and pulmonary adenomatosis in imported sheep; Moller K et al.; The occurrence of abscess disease, caseous lymphadenitis, and pulmonary adenomatosis in sheep in Denmark is reported for the first time . Subcutaneous abscesses were observed in imported 4- to 5-month-old lambs of the Lacaune breed 10 days after arrival in Denmark . Abscesses were mostly located in the head, neck and shoulder regions close to the regional lymph nodes . Bacteriological examinations revealed growth of Staphylococcus aureus ssp . anaerobius in all animals with subcutaneously located abscesses containing a viscous white-yellow odourless mass . In addition, Corynebacterium pseudotuberculosis was isolated from abscesses in one animal and lesions consistent with pulmonary adenomatosis were found in four animals. Prikl Biokhim Mikrobiol, 2000 Mar-Apr, 36(2), 221 - 4 {Preparation and immunochemical properties of monoclonal antibodies to potato ring rot pathogen Corynebacterium sepedonicum}; Ambrosova SM et al.; Five stable hybridoma lines producing monoclonal antibodies to Corynebacterium sepedonicum were obtained . The specificity of monoclonal antibodies obtained was characterized . Interactions of the antibodies with native cells and antigenic preparations from bacterial cell extracts were studied . The epitope specificity of these antibodies to their recognized antigens and the use of the antibodies in advanced immunodiagnostic assays are discussed. Prikl Biokhim Mikrobiol, 2000 Mar-Apr, 36(2), 117 - 21 {ATP and polyphosphate-dependent bacterial NAD+-kinases}; Filippovich SIu et al.; Measurable levels of activity of NAD+ kinases of actinomycetes Micrococcus luteus and Corynebacterium ammoniagenes were observed after substituting inorganic tripolyphosphate for ATP, whereas the enzyme from the eubacterium Escherichia coli was not active with this substrate . Gradient PAGE found two molecular isoforms of NAD+ kinase in C . ammoniagenes and E . coli; four forms were found in M . luteus . All isoforms of this enzyme found in C . ammoniagenes and M . luteus displayed a NADP-synthesizing activity in the presence of either ATP or tripolyphosphate . Because of its capability of utilizing inorganic tripolyphosphate, M . luteus is the most promising NADP producer organism. Int J Parasitol, 2000 Apr 24, 30(5), 599 - 607 Identification of a 30 kDa antigen from leishmania (L.) chagasi amastigotes implicated in protective cellular reponses in a murine model; Pinto AR et al.; An antigen of apparent molecular mass of 30 kDa, termed p30, was purified from Leishmania (L.) chagasi amastigotes after separation of parasite extracts by sodium dodecyl sulfate-polyacrylamide gel eletroctrophoresis followed by electroelution . The use of the purified antigen in lymphocyte cultures from BALB/c mice previously immunised with L . (L.) chagasi amastigotes led to high levels of proliferation . Animal immunisation with p30 plus complete Freund's adjuvant either by subcutaneous or intraperitoneal route led to comparable antigenic stimulation . Similar stimulation indices induced by p30 were also obtained when animals were immunised with Corynebacterium parvum as adjuvant by the intraperitoneal route . Detection of IL-2 and IFN-gamma in the supernatants from lymphocytes stimulated by p30 and inhibition of the production of these lymphokines in the presence of anti-CD4 strongly indicated the involvement of the Th1 subset in the responses elicited by p30 antigen . Immunisation of BALB/c mice with p30 provided partial protection against challenge with L . (L.) chagasi amastigotes, indicating a protective role for p30 and that Th1 can be related to accquired resistance to visceral leishmaniasis in a murine model . Further characterisation studies were performed by the use of a monoclonal antibody directed to a cysteine proteinase of 30 kDa from L . (L.) amazonensis amastigotes . Despite the cross-reactivity presented by p30 from both Leishmania species, the p30 from L . (L.) chagasi amastigotes lacks proteolytic activity. Appl Microbiol Biotechnol, 2000 Mar, 53(3), 257 - 61 Large-scale production of CMP-NeuAc and sialylated oligosaccharides through bacterial coupling; Endo T et al.; A large-scale production system of cytidine 5'monophospho-N-acetylneuraminic acid (CMP-NeuAc) and sialyloligosaccharides was established by a whole-cell reaction through the combination of recombinant Escherichia coli strains and Corynebacterium ammonia-genes . For the production of CMP-NeuAc, two recombinant E . coli strains were generated that overexpressed the genes of CMP-NeuAc synthetase and CTP synthetase, respectively . C . ammoniagenes contributed to the formation of UTP from orotic acid . CMP-NeuAc was accumulated at 27 mM (17 g/l) after a 27-h reaction starting with orotic acid and N-acetylneuraminic acid . When E . coli cells that overexpressed the alpha-(2-->3)-sialyltransferase gene of Neisseria gonorrhoeae were put into the CMP-NeuAc production system, 3'-sialyllactose was accumulated at 52 mM (33 g/l) after an 11-h reaction starting with orotic acid, N-acetylneuraminic acid, and lactose . Almost no oligosaccharide byproducts other than 3'-sialyllactose were observed after the reaction . The production of 3'-sialyllactose at a 5-l jar fermenter scale was almost the same as that at a beaker scale, which indicated the high potential of the 3'-sialyllactose production on an industrial scale. J Biol Chem, 2000 Apr 21, 275(16), 11686 - 92 Identification of the proximal ligand His-20 in heme oxygenase (Hmu O) from Corynebacterium diphtheriae . Oxidative cleavage of the heme macrocycle does not require the proximal histidine; Wilks A et al.; The coordination and spin-state of the Corynebacterium diphtheriae heme oxygenase (Hmu O) and the proximal Hmu O H20A mutant have been characterized by UV-visible and resonance Raman (RR) spectrophotometry . At neutral pH the ferric heme-Hmu O complex is a mixture of six-coordinate high spin and six-coordinate low spin species . Changes in the UV-visible and high frequency RR spectra are observed as a function of pH and temperature, with the six-coordinate high spin species being converted to six-coordinate low spin . The low frequency region of the ferrous RR spectrum identified the proximal ligand to the heme as a neutral imidazole with a Fe-His stretching mode at 222 cm(-1) . The RR characterization of the heme-CO complex in wt-Hmu O confirms that the proximal imidazole is neither ionized or strongly hydrogen-bonded . Based on sequence identity with the mammalian enzymes the proximal ligand in HO-1 (His-25) and HO-2 (His-45) is conserved (His-20) in the bacterial enzyme . Site-specific mutagenesis identified His-20 as the proximal mutant based on electronic and resonance Raman spectrophotometric analysis . Titration of the heme-Hmu O complex with imidazole restored full catalytic activity to the enzyme, and the coordination of imidazole to the heme was confirmed by RR . However, in the absence of imidazole, the H20A Hmu O mutant was found to catalyze the initial alpha-meso-hydroxylation of the heme . The product of the aerobic reaction was determined to be ferrous verdoheme . Hydrolytic conversion of the verdoheme product to biliverdin concluded that oxidative cleavage of the porphyrin macrocycle was specific for the alpha-meso-carbon . The present data show that, in marked contrast to the human HO-1, the proximal ligand is not essential for the initial alpha-meso-hydroxylation of heme in the C . diphtheriae heme oxygenase-catalyzed reaction. Biochim Biophys Acta, 2000 Apr 12, 1484(2-3), 87 - 92 The change in leukotrienes and lipoxins in activated mouse peritoneal macrophages; Calorini L et al.; The aim of this study was to investigate to what extent the generation of leukotrienes (LTs) and lipoxins (LXs) was affected by the expression of definite levels of macrophage activation . We used a system of murine peritoneal macrophages at different states of activation consisting in resident macrophages and FCS-, thioglycollate- or Corynebacterium parvum-elicited macrophages . The profile of lipoxygenase metabolites in resident macrophages was characterized by the presence of high levels of 12-HETE, followed by 15-HETE, 5-HETE, LTB(4) and 6-trans-LTB(4), 6-trans-12-epi-LTB(4) . A comparable pattern was also found in FCS-elicited macrophages which appeared not to be responsive to the challenge with interferon gamma plus LPS, as measured by the generation of NO and tumor necrosis factor alpha . Resident as well as FCS-elicited macrophages also generated appreciable quantities of LXs (A(4) and B(4)) . Thioglycollate-elicited macrophages, which expressed a state of 'responsive' macrophages, showed a block of the LT and LX synthesis . This block was also present in C . parvum-elicited macrophages which expressed a fully 'activated' phenotype, reflected by their capacity of releasing NO and tumor necrosis factor alpha even though they were not challenged . These results provide the first evidence that the level of 'responsive' as well as 'activated' macrophages was associated with of a simultaneous block of LTB(4) and LXs. Prev Vet Med, 2000 Apr 28, 44(3-4), 205 - 20 Udder health and risk factors for subclinical mastitis in organic dairy farms in Switzerland; Busato A et al.; 1+ were submitted for somatic cell counting (SCC), bacteriological examination and to test for antibiotic susceptibility . The SCC and germ-cell counts of monthly bulk-tank milk samples were available through Dairy Inspection and Advisory Services and milk production data of 567 herd-book cows were available from breeding associations . Possible individual and environmental predictors of subclinical mastitis were identified using logistic models adjusted for clustering of the data at herd and cow levels . Data were analyzed separately for cows from 7 to 100 and from 101 to 305 days post partum . Prevalences of subclinical mastitis at the quarter level were 21.2% for lactation period 7-100 days and 34.5% for 101-305 days post partum . The geometric mean SCC in bulk-tank milk samples was 85.6x10(3)cells/ml . Samples at 7-100 and 101-305 days post partum were positive for Staphylococcus aureus in 16.0 and 7.4%, for coagulase-negative Staphylococci in 51.5 and 50.6%, for Streptococcus agalactiae in 0.0 and 0.8%, for other Streptococci in 19.4 and 15.6%, for E . coli in 1.0 and 0.4%, and for Corynebacterium bovis in 25.7 and 45.1%, respectively . Risks of subclinical mastitis increased significantly with increasing days post partum and advancing age of cow . Cows that were sampled when staying in alpine dairies had considerably higher risks of subclinical mastitis than cows staying in home barns . Significantly lower risks of subclinical mastitis were observed in farms where CMT was performed regularly as a control measure . Bacteria in milk from cows with mastitis exhibited antibiotic resistance at a comparable frequency as found previously in conventional farms. Mol Microbiol, 2000 Apr, 36(1), 68 - 84 Corynebacterium diphtheriae genes required for acquisition of iron from haemin and haemoglobin are homologous to ABC haemin transporters; Drazek ES et al.; Corynebacterium diphtheriae and Corynebacterium ulcerans use haemin and haemoglobin as essential sources of iron during growth in iron-depleted medium . C . diphtheriae and C . ulcerans mutants defective in haemin iron utilization were isolated and characterized . Four clones from a C . diphtheriae genomic library complemented several of the Corynebacteria haemin utilization mutants . The complementing plasmids shared an approximately 3 kb region, and the nucleotide sequence of one of the plasmids revealed five open reading frames that appeared to be organized in a single operon . The first three genes, which we have termed hmuT, hmuU and hmuV, shared striking homology with genes that are known to be required for haemin transport in Gram-negative bacteria and are proposed to be part of an ABC (ATP-binding cassette) transport system . The hmuT gene encodes a 37 kDa lipoprotein that is associated with the cytoplasmic membrane when expressed in Escherichi coli and C . diphtheriae . HmuT binds in vitro to haemin- and haemoglobin-agarose, suggesting that it is capable of binding both haemin and haemoglobin and may function as the haemin receptor in C . diphtheriae . This study reports the first genetic characterization of a transport system that is involved in the utilization of haemin and haemoglobin as iron sources by a Gram-positive bacterium. Mol Microbiol, 2000 Mar, 35(6), 1454 - 68 Manganese homeostasis in Bacillus subtilis is regulated by MntR, a bifunctional regulator related to the diphtheria toxin repressor family of proteins; Que Q et al.; The Bacillus subtilis yqhN gene encodes a metalloregulatory protein distantly related to the Corynebacterium diphtheriae diphtheria toxin repressor (DtxR) . While DtxR mediates the iron-dependent repression of iron uptake, we demonstrate that yqhN (herein renamed mntR) encodes a manganese modulated regulator of manganese transport . An mntR mutant strain is sensitive to both manganese and cadmium, suggesting that the transport of these metals is derepressed . We selected Tn10 insertions that suppress the Mn(II) sensitivity of the mntR mutant or that increase the Cd(II) tolerance of wild-type cells, and in both cases we recovered insertions in mntH (formerly ydaR) . MntH is a member of the NRAMP family of proton-coupled, metal ion transporters . MntR also regulates expression of a Mn(II) ABC transporter (MntABCD) . The MntH and MntABCD transporters are both selectively repressed by Mn(II) and this regulation requires MntR . In high Mn(II) conditions, MntR functions as a Mn(II)-dependent repressor of mntH transcription . In contrast, MntR acts as a positive regulator of the mntABCD operon under low Mn(II) growth conditions . Biochemical studies demonstrate that MntR binding to the mntH control region requires Mn(II), while interaction with the mntABCD control region does not depend on Mn(II). Prikl Biokhim Mikrobiol, 2000 Jan-Feb, 36(1), 59 - 67 {Storage of industrial microorganisms entrapped into polymer matrices}; Fedorov AIu et al.; Our study of the techniques of long-term storage of the biomass of various strains of microorganisms, which cause breakdown or transformation of synthetic organic compounds, demonstrates that desiccated agar beads with immobilized microbial cells can be used for this purpose . In addition, the cells can be stored in desiccated matrices of agar or polyvinyl alcohol, coating synthetic cords . Such dry biocatalysts may be used for quick starting of bioreactors and in other biotechnological processes . The technique is applicable to storage of various strains of Pseudomonas, Corynebacterium, Rhodococcus, and, to a lesser extent, Enterobacteriaceae. J Biol Chem, 2000 Jun 9, 275(23), 17494 - 500 Histidine 20, the crucial proximal axial heme ligand of bacterial heme oxygenase Hmu O from Corynebacterium diphtheriae; Chu GC et al.; The hemin complex of Hmu O, a 24-kDa soluble heme degradation enzyme in Corynebacterium diphtheriae, is coordinated axially to a neutral imidazole of a proximal histidine residue in Hmu O . To identify which of the eight histidines in Hmu O is the proximal heme ligand, we have constructed and expressed the plasmids for eight His --> Ala Hmu O mutants . Reconstituted with hemin, the active site structures and enzymatic activity of these mutants have been examined by EPR, resonance Raman, and optical absorption spectroscopy . EPR of the NO-bound ferrous heme-Hmu O mutant complexes reveals His(20) as the proximal heme ligand in Hmu O, and this is confirmed by resonance Raman results from the ligand-free ferrous heme-H20A . All eight His --> Ala mutants bind hemin stoichiometrically, proving that none of the histidines is essential for hemin-Hmu O formation . However, His(20) is crucial to Hmu O catalysis . Its absence by point mutation has inhibited the conversion of hemin to biliverdin . The ferric heme-H20A complex is pentacoordinate . Resonance Raman of the CO-bound ferrous heme-H20A corroborates this and reveals an Fe-C-O bending mode, delta(Fe-C-O), the first reported for a pentacoordinate CO-bound hemeprotein . The appearance of delta(Fe-C-O) in C . diphtheriae Hmu O H20A but not mammalian HO-1 mutant H25A indicates that the heme environment between the two heme oxygenases is different. J Clin Microbiol, 2000 Apr, 38(4), 1385 - 9 Rapid enzyme immunoassay for determination of toxigenicity among clinical isolates of corynebacteria; Engler KH et al.; A rapid enzyme immunoassay (EIA) was developed for the phenotypic detection of diphtheria toxin among clinical isolates of corynebacteria . The assay uses equine polyclonal antitoxin as the capture antibody and an alkaline phosphatase-labeled monoclonal antibody, specific for fragment A of the toxin molecule, as the detecting antibody . The assay is rapid, sensitive, and specific: a final result is available within 3 h of colony selection, and the limits of detection are 0.1 ng of pure diphtheria toxin/ml . Toxigenicity could be detected with isolates grown on a diverse range of culture media, including selective agars . Toxin detection using the EIA was compared to that with the Elek test and PCR detection of fragment A of the diphtheria toxin (tox) gene, using 245 isolates of corynebacteria . The results for the EIA were in complete concordance with those of the Elek test: 87 toxigenic and 158 nontoxigenic isolates . Ten of the phenotypically nontoxigenic strains were found to contain fragment A of the tox gene but did not express the toxin protein . These isolates were found to be nontoxigenic in the Vero cell tissue culture cytotoxicity assay and were therefore nontoxigenic for diagnostic purposes . The EIA is a simple rapid phenotypic test which provides a definitive result on toxigenicity within one working day. J Microbiol Methods, 2000 Mar, 40(1), 63 - 6 Usefulness of the BACTEC MYCO/F lytic system for detection of mycobacteremia in a clinical microbiology laboratory; Esteban J et al.; 278 BACTEC MYCO/F lytic system blood cultures for mycobacteria were evaluated between 1997 and 1999 . Sixty of them were read as positive by the system, being considered 15 of them as false positives . Twenty-seven yielded mycobacterial growth (13 Mycobacterium avium from 3 patients and 14 Mycobacterium tuberculosis from 8 patients) . Other bacteria isolated were coagulase-negative Staphylococcus (13 samples), Corynebacterium sp . (5 samples), Salmonella enteritidis (2 samples) and Klebsiella pneumoniae (1 sample) . Five of these isolates were considered as true episodes of bacteremia . The average time for detection of mycobacteria was 12.6 days for M . avium and 26.4 days for M . tuberculosis . BACTEC MYCO/F lytic system is useful for detection of mycobacteremia in clinical microbiology laboratories. Proteins, 2000 Apr 1, 39(1), 68 - 75 Molecular modeling of substrate binding in wild-type and mutant Corynebacteria 2,5-diketo-D-gluconate reductases; Khurana S et al.; 2,5-diketo-D-gluconic acid reductase (2,5-DKGR; E.C . 1.1.1.-) catalyzes the Nicotinamide adenine dinucleotide phosphate (NADPH)-dependent stereo-specific reduction of 2, 5-diketo-D-gluconate (2,5-DKG) to 2-keto-L-gulonate (2-KLG), a precursor in the industrial production of vitamin C (L-ascorbate) . Microorganisms that naturally ferment D-glucose to 2,5-DKG can be genetically modified to express the gene for 2,5-DKGR, and thus directly produce vitamin C from D-glucose . Two naturally occurring variants of DKGR (DKGR A and DKGR B) have been reported . DKGR B exhibits higher specific activity toward 2,5-DKG than DKGR A; however, DKGR A exhibits a greater selectivity for this substrate and significantly higher thermal stability . Thus, a modified form of DKGR, combining desirable properties from both enzymes, would be of substantial commercial interest . In the present study we use a molecular dynamics-based approach to understand the conformational changes in DKGR A as the active site is mutated to include two active site residue changes that occur in the B form . The results indicate that the enhanced kinetic properties of the B form are due, in part, to residue substitutions in the binding pocket . These substitutions augment interactions with the substrate or alter the alignment with respect to the putative proton donor group . Proteins 2000;39:68-75 . Biosci Biotechnol Biochem, 2000 Feb, 64(2), 424 - 7 A new type of glycoglycerolipids from Corynebacterium aquaticum; Yanagi H et al.; A new type of glycoglycerolipids, S361A and S365A, were obtained from Corynebacterium aquaticum strains, S361 and S365, newly isolated from soils, and were identified as (2R)-1-{alpha-glucopyranosyl-(1alpha-3)-(6O-acyl-alpha-manno pyranosyl)}-3-O-acylglycerol and (2R)-1-{alpha-mannopyranosyl-(1alpha-3)-(6-O-acyl-alpha-mannopyran osyl)}-3-O-acylglycerol, respectively . S365A was identical to a novel glycoglycerolipid recently isolated from some bacteria, but S361A was a new analog having a glucosylmannosyl in place of the dimannosyl group . Our results indicate that this sn-2 lysotype of glyceroglycolipids may be widely distributed in bacteria. J Pharmacol Exp Ther, 2000 Apr, 293(1), 136 - 50 Effects of cannabinoid receptor agonist and antagonist ligands on production of inflammatory cytokines and anti-inflammatory interleukin-10 in endotoxemic mice; Smith SR et al.; Previous studies have shown that mice primed with Corynebacterium parvum produce higher levels of inflammatory cytokines than unprimed mice upon challenge with lipopolysaccharide (LPS) . Herein, we describe experiments in which two cannabinoid (CB) agonists, WIN 55212-2 {(R)-(+)-{2, 3-dihydro-5-methyl-3-{(4-morpholinyl)methyl}pyrrolo{1,2,3-de}1, 4-benzoxazin-6-yl}(1-naphthyl)methanone) and HU-210 {(-)-11-hydroxy-delta(8) tetrahydrocannabinol-dimethylheptyl}, were examined for their effects on LPS-induced cytokines in C . parvum-primed and unprimed mice . These agonists have been reported to bind selectively to the CB2 and CB1 receptor subtypes, respectively . WIN 55212-2 (3.1-50 mg/kg i.p.) and HU-210 (0.05-0.4 mg/kg i.p.) decreased serum tumor necrosis factor-alpha and interleukin-12 (IL-12) and increased IL-10 when administered to mice before LPS . The drugs also protected C . parvum mice (but not unprimed mice) against the lethal effects of LPS . The protection afforded to C . parvum mice could not be attributed to the higher levels of IL-10 present in these mice after agonist treatment . The WIN 55212-2- and HU-210-mediated changes in the responsiveness of mice to LPS were antagonized by SR141716A {N-(piperdin-1-yl)-5-(4-chloropheny)-1-(2, 4-dichloropheny)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride}, a selective CB1 receptor antagonist, but not by SR144528 {N-{(1S)-endo-1,3,3-trimethylbicyclo{2.2 . 1}heptan-2-yl}5-(4-choro-3-methylphenyl)-1-(4-methylbenzyl)p yrazole-3 -carboxamide}, a selective antagonist at the CB2 receptor . Therefore, both CB agonists modulated LPS responses through the CB1 receptor . Surprisingly, SR141716A itself modulated cytokine responses in a manner identical with that of WIN 55212-2 and HU-210 when administered alone to mice . The agonist-like effects of SR141716A, which were more striking in unprimed than in primed mice, suggested that the antagonist also could function as a partial agonist at the CB1 receptor . Our findings indicate a role for the CB1 receptor subtype in cytokine modulation by CB ligands. J Biochem (Tokyo), 2000 Feb, 127(2), 191 - 8 Influenza A virus-binding activity of glycoglycerolipids of aquatic bacteria; Nakata K et al.; As the aqueous sphere has been proposed to be an important source medium for the virus infection of land animals, the glycolipids of some aquatic organisms were examined for human influenza A virus-binding activity . Active compounds were not found among the eight echinoderm gangliosides, but two active non-sialylated glycoglycerolipids were isolated from an aquatic bacterium, Corynebacterium aquaticum . The structural formula of one of them, H632A, was elucidated to be 1-14-methyl-hexadecanoyl-3-alpha-D-galactopyranosyl-(1-->3)-6-(12-met hyl-tetradecanoyl)-1-alpha-D-mannopyranosyl}-sn-glycerol . The latter together with reported one elsewhere, S365A, 1-14-methyl-hexadecanoyl-3-{alpha-D-mannopyranosyl-(1-->3)-6-(12-meth yl-tetradecanoyl)-1-alpha-D-mannopyranosyl}-sn-glycerol, apparently bound to three human influenza viruses, A/PR/8/34 (H1N1), A/Aichi/2/68 (H3N2), and A/Memphis/1/71 (H3N2), exhibiting 7-12% (H632A) and 10-22% (S365A) of the activities of the control substances (Neu5Acalpha2-3-paragloboside and Neu5Acalpha2-6- paragloboside) . Additionally, these glycolipids were assumed to have virus-neutralizing activities for the following two reasons: (i) The hemagglutination and hemolysis activities of the viruses were inhibited by the glycolipid . (ii) The leakage of a cytosolic enzyme (lactate dehydrogenase) from Madin-Darby canine kidney cells on virus infection was prevented by the glycolipids to nearly the same extent as by fetuin . This is the first evidence of the binding- and neutralizing-abilities of native glycoglycerolipids as to influenza viruses. Mol Gen Genet, 2000 Feb, 263(1), 1 - 11 The 51,409-bp R-plasmid pTP10 from the multiresistant clinical isolate Corynebacterium striatum M82B is composed of DNA segments initially identified in soil bacteria and in plant, animal, and human pathogens; Tauch A et al.; The 51,409-bp DNA sequence of the multiresistance plasmid pTP10 from the gram-positive opportunistic human pathogen Corynebacterium striatum M82B has been determined . Fully automated genome interpretation led to the identification of 47 ORFs . Analysis of the genetic organization of pTP10 suggests that the plasmid is composed of eight DNA segments, the boundaries of which are represented by transposons and insertion sequences . The DNA segments of pTP10 are highly similar to (1) a plasmid-encoded erythromycin resistance region from the human pathogen Corynebacterium diphtheriae; (2) a chromosomal DNA region from Mycobacterium tuberculosis; (3) a plasmid-encoded chloramphenicol resistance region from the soil bacterium Corynebacterium glutamicum; (4) transposable elements from phytopathogenic gram-negative Pseudomonas, Xanthomonas and Erwinia species; and (5) a plasmid-encoded aminoglycoside resistance region from the gram-negative fish pathogen Pasteurella piscicida . The complete DNA sequence of pTP10 provides genetic information regarding the mechanisms of resistance to 16 antimicrobial agents that belong to six structural classes . In addition, the mosaic structure of pTP10 represents the evolutionary consolidation into a single plasmid molecule of antimicrobial resistances from microorganisms found in different habitats by means of mobile elements, resulting in the generation of a multiresistant bacterium that can infect humans. Electrophoresis, 2000 Feb, 21(3), 654 - 9 Two-dimensional electrophoretic analysis of Corynebacterium glutamicum membrane fraction and surface proteins; Hermann T et al.; An improved protocol for the two-dimensional analysis of proteins of the Corynebacterium glutamicum cytoplasmic membrane fraction is described . By use of increased 3-{(3-cholamidopropyl)dimethylammonio}-1-propanesulfonate (CHAPS) concentrations (2-4%) and an optimized electrophoresis protocol, horizontal streaking of proteins of the cytoplasmic membrane fraction was almost completely avoided . More important, in contrast to a previously published method, both a sample tray and IPG-phor isoelectric focusing unit can be used for the in-gel application of proteins . The described protocol was also found to be suitable for hydrophilic cytoplasmic proteins . Additionally, the preparation and analysis of C . glutamicum cell surface proteins is described . Proteins were extracted with lauroyl sarcosinate and 100-120 spots were separated on two-dimensional (2-D) gels in comparison to 18-20 spots observed previously by standard sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) . C . glutamicum proteins can now be separated into three distinct fractions resembling different functional units of the bacterial cell. Can Vet J, 2000 Feb, 41(2), 126 - 7 Caseous lymphadenitis caused by Corynebacterium ulcerans in the dromedary camel; Tejedor MT et al.; Caseous lymphadenitis that affected the dorsal and ventral superficial lymph nodes in the left cervicothoracic region of a young dromedary camel is described . The agent isolated was Corynebacterium ulcerans . To our knowledge, this is the first description of purulent lymphadenitis caused by C . ulcerans in a species belonging to the Camelidae. Int J Antimicrob Agents, 2000 Mar, 14(2), 99 - 105 Vancomycin resistance in Gram-positive bacteria other than Enterococcus spp; Krcmery V et al.; This is a review article on vancomycin resistance on gram positive bacteria other than enterococci . Epidemiology of varying resistance, its clinical relevance and therapeutic options in infections caused by vancomycin resistant Listeria spp., Corynebacteria, streptococci and staphylocci are discussed. Mol Microbiol, 2000 Mar, 35(5), 1026 - 41 Characterization of the in vivo acceptors of the mycoloyl residues transferred by the corynebacterial PS1 and the related mycobacterial antigens 85; Puech V et al.; Mycolic acids, long-chain (C70-C90) alpha-alkyl, beta-hydroxy fatty acids, are characteristic cell envelope components of mycobacteria; similar but shorter-chain substances occur in corynebacteria and related taxa . These compounds apparently play an important role in the physiology of these bacteria . The deduced N-terminal region of PS1, one of the two major secreted proteins of Corynebacterium glutamicum encoded by the csp1 gene, is similar to the antigens 85 complex of Mycobacterium tuberculosis which has been shown to be associated in vitro with a mycoloyltransferase activity onto trehalose . Overexpression of PS1 in the wild-type strain of C . glutamicum suggested the implication of the protein in the transfer of corynomycolates, evidenced by an increase esterification of the cell wall arabinogalactan with corynomycolic acid residues and an accumulation of trehalose dicorynomycolates . Overexpression of truncated forms of PS1 demonstrated that the crucial region for transfer activity of the protein involves all the region of homology with antigens 85 . To establish the putative mycoloyltransferase activity of PS1, a csp1-inactivated mutant of C . glutamicum was biochemically characterized . Inactivation of the gene resulted in: (i) a 50% decrease in the cell wall corynomycolate content; (ii) the alteration of the permeability of the C . glutamicum cell envelope; (iii) the decrease of the trehalose dicorynomycolate content; (iv) the accumulation of trehalose monocorynomycolate; and (v) the appearance of a glycolipid identified as 6-corynomycoloylglucose . Complementation of the mutant by the csp1 gene fully restored the wild-type phenotype . Finally, a mycoloyltransferase assay established that PS1 possesses a trehalose mycoloyltransferase activity . To define the in vivo function of antigens 85, the csp1-inactivated mutant was complemented with the fbpA, fbpB or fbpC genes . Complementation with the different fbp genes restored the normal cell wall corynomycolate content and permeability, but did not affect either the fate of trehalose corynomycolates or the occurrence of glucose corynomycolate . Thus, PS1 is one of the enzymes that transfer corynomycoloyl residues onto both the cell wall arabinogalactan and trehalose monocorynomycolate, whereas in the whole bacterium the mycobacterial antigens 85A, 85B and 85C can transfer mycolates only onto the cell wall acceptor in C . glutamicum. Biotechnol Bioeng, 2000 Mar 20, 67(6), 872 - 85 Metabolic flux distributions in Corynebacterium glutamicum during growth and lysine overproduction . Reprinted from Biotechnology and Bioengineering, Vol . 41, Pp 633-646 (1993) Vallino JJ, Stephanopoulos G. The two main contributions of this article are the solidification of Corynebacterium glutamicum biochemistry guided by bioreaction network analysis, and the determination of basal metabolic flux distributions during growth and lysine synthesis . Employed methodology makes use of stoichiometrically based mass balances to determine flux distributions in the C . glutamicum metabolic network . Presented are a brief description of the methodology, a thorough literature review of glutamic acid bacteria biochemistry, and specific results obtained through a combination of fermentation studies and analysis-directed intracellular assays . The latter include the findings of the lack of activity of glyoxylate shunt, and that phosphoenolpyruvate carboxylase (PPC) is the only anaplerotic reaction expressed in C . glutamicum cultivated on glucose minimal media . Network simplifications afforded by the above findings facilitated the determination of metabolic flux distributions under a variety of culture conditions and led to the following conclusions . Both the pentose phosphate pathway and PPC support significant fluxes during growth and lysine overproduction, and that flux partitioning at the glucosa-6-phosphate branch point does not appear to limit lysine synthesis . Appl Environ Microbiol, 2000 Mar, 66(3), 1202 - 4 Reductive cleavage of demeton-S-methyl by Corynebacterium glutamicum in cometabolism on more readily metabolizable substrates; Girbal L et al.; Corynebacterium glutamicum is able to biotransform demeton-S-methyl, an organophosphorus compound, during cometabolism with more readily metabolizable substrates . Among the cosubstrates used, fructose is the growth substrate that is most favorable for demeton-S-methyl biotransformation . The reaction mechanism of demeton-S-methyl biotransformation involves reductive cleavage of an S-C bond, which leads to accumulation of dimethyl thiophosphate in the culture medium. J Protein Chem, 1999 Oct, 18(7), 747 - 52 Determination of amino acid sequences of two subunits in sarcosine oxidase from Corynebacterium sp . U-96; Mukouyama EB et al.; The primary structures of the C and D subunits of sarcosine oxidase from Corynebacterium sp . U-96 were determined by sequencing the peptide fragments derived from their enzymatic digestions . The C and D subunits were shown to be composed of 199 and 92 residues, respectively . Each amino acid sequence showed a high homology with the sequence of the corresponding subunit from Corynebacterium sp . P-1 . However, there were some differences between these two species, that is, four N-terminal residues were truncated in the C subunit, but six C-terminal residues were truncated in the D subunit . The D subunit contained three cysteine residues, but no disulfide bonds are in the subunit . Overall sequences of both subunit showed no homology with any other protein in the data base. Infect Immun, 2000 Mar, 68(3), 1328 - 36 Subunit vaccination of mice against new world cutaneous leishmaniasis: comparison of three proteins expressed in amastigotes and six adjuvants; Aebischer T et al.; A mixture of well-defined recombinant antigens together with an adjuvant that preferentially stimulates specific gamma interferon (IFN-gamma)-secreting helper type 1 CD4(+) T cells (Th1 cells) presents a rational option for a vaccine against leishmaniasis . The potential of this approach was investigated in murine infections with Leishmania mexicana, which are characterized by the absence of a parasite-specific Th1 response and uncontrolled parasite proliferation . A mixture of three antigens (glycoprotein 63, cysteine proteinases, and a membrane-bound acid phosphatase), which are all expressed in amastigotes, the mammalian stage of the parasite, were used for the immunization of C57BL/6 mice in combination with six adjuvants (interleukin 12 {IL-12}, Detox, 4'-monophosphoryl lipid A, QS-21, Mycobacterium bovis BCG, and Corynebacterium parvum) . All six vaccine formulations containing the mixture of recombinant antigens were protective against challenge infections with promastigotes, the insect stage of the parasite, in that mice controlled and healed infections but developed transient and, in certain cases, accentuated disease . The most effective adjuvants were IL-12 followed by Detox . Further studies using these two adjuvants showed that a similar protective effect was observed with a mixture of the corresponding native proteins, and mice which had controlled the infection showed a preponderance of IFN-gamma-secreting CD4(+) T cells in the lymph nodes draining the lesion . Using the recombinant proteins individually, it is shown that the relatively abundant cysteine proteinases and glycoprotein 63, but not the acid phosphatase, are able to elicit a protective response . The results are discussed in comparison to previous studies with subunit vaccines and with respect to cell biological aspects of antigen presentation in Leishmania-infected macrophages. Anal Biochem, 2000 Mar 1, 279(1), 100 - 5 Pulsed high-field gradient in vivo NMR spectroscopy to measure diffusional water permeability in Corynebacterium glutamicum; Schoberth SM et al.; Pulsed high-field gradient in vivo NMR spectroscopy was used to measure diffusional water permeability in cell suspensions of the Gram-positive bacterium Corynebacterium glutamicum . Two different regions of H2O mobility were detected . One was characterized by the apparent coefficient of self-diffusion, D(1 app) = (4.6-12.7)x10(-8) cm(2) s(-1), depending on the observation time t . The other region was characterized by D(2) = 1.4x10(-5) cm(2) s(-1) . The value of D(2) was similar to the diffusion coefficient of H2O in free water and in extracellular biological fluids . Restricted diffusion could be demonstrated for the slower process (D(1)) . It was attributed to the cytoplasm of the cells . The membrane permeability, P(d H2O), for C . glutamicum was (4.8+/-0.4)x10(-3) cm s(-1) . It compared favorably with values reported for human erythrocytes and was higher by a factor of about 100 compared to the diffusional permeability for ethanol, P(d ethanol), in Zymomonas mobilis . Addition of HgCl2, a water channel inhibitor in eukaryotes, decreased P(d H2O) in C . glutamicum by a factor of approximately 8 . To our knowledge, these are the first functional studies of water transport in prokaryotes that yielded quantitative data, viz., transmembrane water permeability expressed through D(H2O) and P(d H2O) . Comp Immunol Microbiol Infect Dis, 2000 Mar, 23(2), 91 - 7 Natural killer activity in mice infected with rabies virus and submitted to P . acnes (Propionibacterium acnes) as immunomodulator; Megid J et al.; The natural killer (NK) activity and lethality were evaluated in swiss mice experimentally infected with street rabies virus and submitted to immunomodulation by P . acnes (formerly Corynebacterium parvum) . The infected animals were sacrificed at different times and spleen non-adherent cells were obtained through ficoll-hypaque gradient and depletion of glass-adherent cells . Immunosuppression was observed in rabies virus infected mice correlated with lower NK activity in clinically ill animals . Higher NK activity and percentual of survival were observed in the group submitted to P . acnes . The increased survival correlated with higher NK activity induced by P . acnes suggests a protective role of this natural barrier against rabies virus infection in mice. Actas Urol Esp, 1999 Nov-Dec, 23(10), 885 - 7 {Encrusted alkaline cystitis and malacoplakia}; Farina Perez LA et al.; Urinary infection due to urea splitting bacteria leads to a rise in urinary pH, favouring the precipitation of calcium salts and struvita crystals . If deposited on the surface of a bladder with chronic inflammation or some other previous lesion, may produce an alkaline encrusted cystitis, now a rare condition . In the case here presented, occurred in a 69-year-old male . Corynebacterium urealyticum grown in the urine, and some foci of malakoplakia were found in the area of encrustation endoscopically excised . This case seems to be the third example of alkaline encrusted cystitis associated with malakoplakia reported in the bibliography . These two conditions share similar clinical signs and may probably have a common aetiopathogenesis. Biosci Biotechnol Biochem, 1999 Dec, 63(12), 2216 - 8 Site-directed mutagenesis of two zinc-binding centers of the NADH-dependent phenylacetaldehyde reductase from styrene-assimilating Corynebacterium sp . strain ST-10; Wang JC et al.; Phenylacetaldehyde reductase (PAR) with a unique and wide substrate range from styrene-assimilating Corynebacterium sp . strain ST-10, which is a useful biocatalyst producing chiral alcohols, has been found to belong to a family of zinc-containing, long-chain alcohol dehydrogenases (ADHs) on the basis of the primary structure similarity . The enzyme contains 2 moles of zinc per mole of subunit . The amino acid residues assumed to be three catalytic and four structural zinc-binding ligands were characterized by site-directed mutagenesis, compared with other zinc-containing, long-chain ADHs . Sixteen PAR mutants gave measurable but rather low activities toward phenylacetaldehyde, n-hexyl aldehyde, and 2-heptanone, although they maintained the activities of 8 to 16% of that of wild-type PAR for an acetophenone substrate except that the D153N mutant showed quite low activity . The results suggested that the seven residues present in PAR were probably zinc-binding ligands, and mutation in these residues caused a change in activities for some substrates. J Hosp Infect, 2000 Feb, 44(2), 127 - 33 Bacterial interference among nasal inhabitants: eradication of Staphylococcus aureus from nasal cavities by artificial implantation of Corynebacterium sp; Uehara Y et al.; To evaluate the role of normal flora in the nares in preventing Staphylococcus aureus colonization, we conducted a replacement study in vivo . Staphylococcus epidermidis (rate of colonization: 100%), various species of corynebacteria (52.5%) and S . aureus (25.%) were the major bacterial inhabitants in the nares of 156 healthy volunteers . The low incidence of S . aureus colonization in the carriers with corynebacteria (8.5%), compared to non-carriers (44 . 5%) indicated the possibility of competition for survival between S . aureus and corynebacteria . To confirm this hypothesis, we artificially implanted a strain of Corynebacterium sp (API Coryne bioprofile; 5100304), denoted as Co304 into the nares of 17 S . aureus carriers . S . aureus was completely eradicated in 71% of carriers by up to 15 inoculations of Co304 . However, similar doses of 0.9% NaCl or S . epidermidis into the nares of 10 volunteers did not eradicate S . aureus . No bacteriocin-like activity against S . aureus was detectable, even after mitomycin C stimulation of Co304 . Thus Co304 interfered with S . aureus by a different mechanism to a bacteriocin-like activity . J Infect Dis, 2000 Feb, 181 Suppl 1, S237 - 43 Epidemic diphtheria in the Newly Independent States of the Former Soviet Union: implications for diphtheria control in the United States; Golaz A et al.; The re-emergence of diphtheria in the Newly Independent States of the former Soviet Union in the 1990s raised global awareness of the potential for resurgent disease in countries with long-standing immunization programs . In the United States, the large population of susceptible adults and the possibility of a reintroduction of toxigenic strains of diphtheria create a setting in which diphtheria could spread . In addition, at least one focus of continued circulation of endemic toxigenic Corynebacterium diphtheriae has been identified . Few physicians now have expertise in the diagnosis and treatment of persons with diphtheria, and laboratory capacity is lacking throughout the country . These concerns highlight the importance of maintaining high levels of age-appropriate diphtheria toxoid vaccination, surveillance, accessible and reliable laboratory testing, and training of health care providers . Although the risk of resurgence of diphtheria in the United States is low, public health authorities must ensure that the capacity to recognize, diagnose, and control diphtheria is maintained. J Infect Dis, 2000 Feb, 181 Suppl 1, S168 - 77 Molecular epidemiology of diphtheria; Popovic T et al.; Molecular subtyping of Corynebacterium diphtheriae identified significant genetic diversity within the species and led to the identification of a unique clonal group that emerged in Russia in 1990 at the beginning of the current epidemic . Strains of this group belong to a distinct electrophoretic type complex and are of ribotypes D1 and D4 . Identification of the group allowed for precise monitoring of the epidemic's progression and for rapid detection of cases imported to other countries . The evolution of this clonal group was monitored, and changes were identified . Molecular analysis revealed that no amino acid substitutions have occurred in the diphtheria toxin gene of the epidemic clone strains, reaffirming the use of the current vaccine as the single most effective preventive measure . Application of molecular subtyping methods and continuous monitoring of the spread of these clones has made it possible to distinguish rapidly between epidemic, endemic, and imported cases, allowing for implementation of timely and adequate preventive measures and providing reassurance that no secondary spread resulted from importations. J Infect Dis, 2000 Feb, 181 Suppl 1, S156 - 67 Biology and molecular epidemiology of diphtheria toxin and the tox gene; Holmes RK; Diphtheria toxin (DT) is an extracellular protein of Corynebacterium diphtheriae that inhibits protein synthesis and kills susceptible cells . The gene that encodes DT (tox) is present in some corynephages, and DT is only produced by C . diphtheriae isolates that harbor tox+ phages . The diphtheria toxin repressor (DtxR) is a global regulatory protein that uses Fe2+ as co-repressor . Holo-DtxR represses production of DT, corynebacterial siderophore, heme oxygenase, and several other proteins . Diagnostic tests for toxinogenicity of C . diphtheriae are based either on immunoassays or on bioassays for DT . Molecular analysis of tox and dtxR genes in recent clinical isolates of C . diphtheriae revealed several tox alleles that encode identical DT proteins and multiple dtxR alleles that encode five variants of DtxR protein . Therefore, recent clinical isolates of C . diphtheriae produce a single antigenic type of DT, and diphtheria toxoid continues to be an effective vaccine for immunization against diphtheria. J Infect Dis, 2000 Feb, 181 Suppl 1, S152 - 5 Direct polymerase chain reaction for detection of toxigenic Corynebacterium diphtheriae strains from the Republic of Georgia after prolonged storage; Kobaidze K et al.; A total of 226 paired nose and throat swab specimens from 113 clinical diphtheria cases from the republic of Georgia were analyzed by direct polymerase chain reaction targeting both A and B subunits of the diphtheria toxin gene, tox . Even after prolonged transport and extensive storage (7-14 months) of the clinical specimens in silica gel packages, direct polymerase chain reaction detected the diphtheria tox gene in 54% of the specimens . Specimens obtained by throat swab were three times more likely than those obtained by nose swab to be positive for Corynebacterium diphtheriae. J Infect Dis, 2000 Feb, 181 Suppl 1, S146 - 51 The European Laboratory Working Group on Diphtheria: A global microbiologic network; Efstratiou A et al.; Increasing international travel, migration to and from epidemic areas, and the emergence of epidemic Corynebacterium diphtheriae strains globally have highlighted the need for better microbiologic surveillance and greater clinical and laboratory awareness . As part of the strategy developed by the World Health Organization, one major goal is particularly applicable to the microbiology and epidemiology of diphtheria: to improve surveillance for early detection of emerging and reemerging diseases by establishing a network of national and international laboratories . The European Laboratory Working Group on Diphtheria (ELWGD) was therefore formed in 1993 as a result of the epidemic situation in Eastern Europe . Currently, there is participation from 20 different countries from Western and Eastern Europe, the United States, Australia, and Southeast Asia . ELWGD is a prime example of a collaborative and coordinated approach to the microbiology of diphtheria and should serve as a potential model for other infectious diseases networks. J Infect Dis, 2000 Feb, 181 Suppl 1, S138 - 45 Current approaches to the laboratory diagnosis of diphtheria; Efstratiou A et al.; Despite the success of mass immunization in many countries, diphtheria continues to play a major role as a potentially lethal resurgent infectious disease . Early, accurate diagnosis is imperative since delay in specific therapy may result in death . The microbiologic diagnosis of the disease, the identification of contacts and carriers, and the appropriate clinical management of these patients are therefore crucial . The epidemiology of diseases caused by Corynebacterium diphtheriae has changed dramatically over the decades, a situation that is highlighted by the resurgence of infections in the European region . These factors have strengthened the need for laboratories to screen for C . diphtheriae . Many modified and new methodologies are now used widely within laboratories for diphtheria diagnosis . Recent developments have focused upon methods for detection of the lethal and potent exotoxin produced by the causative organism, C . diphtheriae; this detection is the definitive test for the microbiologic diagnosis of diphtheria. J Infect Dis, 2000 Feb, 181 Suppl 1, S116 - 20 The pathology of diphtheria; Hadfield TL et al.; Diphtheria is an acute, communicable disease caused by Corynebacterium diphtheriae . The disease is generally characterized by local growth of the bacterium in the pharynx with pseudomembrane formation or, less commonly, in the stomach or lungs; systemic dissemination of toxin then invokes lesions in distant organs . Acute disease of the upper respiratory tract usually involves one or more of the following: tonsillar zones, larynx, soft palate, uvula, and nasal cavities . A recent epidemic in Russia emphasized the role of vaccination in reducing disease in children and adults. J Infect Dis, 2000 Feb, 181 Suppl 1, S104 - 9 Diphtheria epidemic in the Republic of Uzbekistan, 1993-1996; Niyazmatov BI et al.; The Republic of Uzbekistan, like the other Newly Independent States in the 1990s, experienced epidemic diphtheria during the 1990s . The outbreak in Uzbekistan began in 1993 in southern regions that bordered areas of Tajikistan that were experiencing a very intense diphtheria epidemic . However, the Uzbek epidemic rapidly spread and threatened to involve the entire country . From 1993-1996, 1169 cases of diphtheria were reported, compared with 58 in 1990-1992 . Unvaccinated or only partially vaccinated cases were more likely to have clinically severe forms of diphtheria than those who were fully vaccinated . Strong epidemiologic links with the Tajik diphtheria epidemic and the predominance of mitis biotype strains of Corynebacterium diphtheriae in Uzbekistan make it likely that the Uzbek outbreak arose independently of the predominantly biotype gravis epidemic that began in Russia . The epidemic appeared to be due to low population immunity and the large-scale reintroduction of toxigenic strains of C . diphtheriae . Several mass vaccination campaigns and general enhancement of routine immunization procedures led to control of the epidemic in 1996. J Infect Dis, 2000 Feb, 181 Suppl 1, S55 - 9 Diphtheria in Lithuania, 1986-1996; Usonis V et al.; Diphtheria reappeared in Lithuania in 1986 and rose to epidemic levels by 1992 . Between 1991 and 1996, 110 cases of diphtheria were registered, with an incidence of 0.03-1.15/100,000 population . Most cases (84%) and all 17 deaths occurred among persons >/=15 years, most of whom had never been vaccinated . Persons 40-49 years old had the highest average annual age-specific morbidity (1.70/100,000) and mortality (0.53/100,000) rates . Low levels of immunity among individuals 40-49 years old and migration to epidemic areas in Russia and Belarus contributed to the epidemic's occurrence . Between 1991 and 1995, toxigenic Corynebacterium diphtheriae strains were isolated from 84 of all registered patients (76%), and nontoxigenic strains were isolated from 13 (12%) . By 1996, two mass vaccination campaigns, which provided one dose of vaccine to individuals 25-30 years old and three doses of vaccine to persons 31-60 years old, helped reduce the number of cases . The first campaign achieved 69% coverage; the second achieved 48% coverage. J Infect Dis, 2000 Feb, 181 Suppl 1, S27 - 34 Diphtheria in the Russian Federation in the 1990s; Markina SS et al.; A resurgence of diphtheria spread throughout the Russian Federation in the early 1990s; diphtheria had been well controlled, but circulation of toxigenic strains of Corynebacterium diphtheriae had persisted since the implementation of universal childhood vaccination in the late 1950s . More than 115,000 cases and 3,000 deaths were reported from 1990 to 1997, and, in contrast to the situation in the prevaccine era, most of the cases and deaths occurred among adults . Contributing factors included the accumulation of susceptible individuals among both adults and children and probably the introduction of new strains of C . diphtheriae . Vaccine quality, vaccine supply, or access to vaccine providers did not significantly contribute to the epidemic . Mass vaccination of adults and improved childhood immunization controlled the epidemic . High levels of population immunity, especially among children, will be needed to prevent and control similar outbreaks in the future. Microbiol Immunol, 1999, 43(12), 1103 - 6 Complement receptor type 3 plays an important role in development of protective immunity to primary and secondary Corynebacterium pseudotuberculosis infection in mice; Lan DT et al.; The present study was performed to investigate the role of CR3, the type 3 complement receptor, in host defense against primary and secondary Corynebacterium (C.) pseudotuberculosis infection in mice . Treatment of mice with 5C6, an anti-CR3 monoclonal antibody (mAb), resulted in unrestricted multiplication of bacteria in the organs and dramatically increased mortalities of the infected mice . Histological examinations showed the inflammation, degeneration and necrosis of organs and revealed that the infection-enhancing effect of 5C6 mAb was associated with the failure of mice to focus mononuclear phagocytes at sites of bacterial multiplication . These results suggest that CR3 plays an important role in host defense against primary as well as secondary C . pseudotuberculosis infection in mice. Zentralbl Bakteriol, 1999 Dec, 289(5-7), 666 - 72 Investigation of skin samples from red foxes (Vulpes vulpes) in eastern Brandenburg (Germany) for the detection of Borrelia burgdorferi s . l; Heidrich J et al.; During earlier investigations a high prevalence of Borrelia (B.) burgdorferi s . l . in unfed Ixodes (I.) ricinus ticks in the Federal State of Brandenburg has been demonstrated . In the present study skin samples were obtained from 100 red foxes (Vulpes vulpes) from the districts where the highest B . burgdorferi prevalences had previously been found (i.e . Uckermark, Barnim, Markisch-Oderland, Oder-Spree) . BSK- and MKP-medium including inhibitory substances were used for cultivation of spirochaetes . Non-motile spirochaete-like organisms were observed in 26% of the samples . Additionally, by subcultures it was not possible to obtain motile helical forms characteristic for B . burgdorferi . On tryptose agar, the bacteria which produced nonmotile forms appeared as corynebacterium-like-colonies . Investigations by electron microscopy showed that the immobile spiral forms were giant whips (flagellae) which belonged to the contaminant flora . These forms proved to be negative for B . burgdorferi s . l . by the use of a nested-PCR . In a further study, the same skin samples were investigated for the presence of B . burgdorferi s . l.-DNA using a nested-PCR . Seven out of 100 samples were positive. Dermatology, 1999, 199(4), 341 - 5 Bacterial interdigital scaly erythema (Kitamura): a possible new clinical entity; Mochizuki T et al.; Four patients with scaly erythema on their finger webs and sides of their fingers during summer are described . These patients were working in either butcher's shops or a sushi bar, where they handled raw meat, chicken or fish for many hours . The eruptions first appeared as scaly erythema, sometimes accompanied by small pustules, on the second, third and fourth finger webs, and later the erythema extended to the sides of the fingers and palms . Maceration and/or shallow erosion sometimes appeared on the finger webs . Symptoms were usually mild; the patients complained of slight itching, irritation or pain . Direct examination of specimens stained with Parker blue-black ink containing KOH revealed scales containing bacterial granules or filaments . Several species of bacteria were cultured including Corynebacterium sp . Fungus was not detected in either KOH specimens or in cultures . The lesions responded rapidly to topical or oral antibiotics; however, they recurred frequently during hot and humid weather . Hitherto a similar condition has not been described and is possibly a new clinical entity . Copyright (R) 2000 S.Karger AG, Basel J Bacteriol, 2000 Jan, 182(2), 432 - 8 Characterization of specific nucleotide substitutions in DtxR-specific operators of Corynebacterium diphtheriae that dramatically affect DtxR binding, operator function, and promoter strength; Lee JH et al.; The diphtheria toxin repressor (DtxR) of Corynebacterium diphtheriae uses Fe(2+) as a corepressor . Holo-DtxR inhibits transcription from the iron-regulated promoters (IRPs) designated IRP1 through IRP5 as well as from the promoters for the tox and hmuO genes . DtxR binds to 19-bp operators with the consensus sequence 5'-TTAGGTTAGCCTAACCTAA-3', a perfect 9-bp palindrome interrupted by a single C . G base pair . Among the seven known DtxR-specific operators, IRP3 exhibits the weakest binding to DtxR . The message (sense) strand of the IRP3 operator (5'-TTAGGTGAGACGCACCCAT-3' {nonconsensus nucleotides underlined}) overlaps by 2 nucleotides at its 5' end with the putative -10 sequence of the IRP3 promoter . The underlined C at position +7 from the center of the IRP3 operator {C(+7)} is unique, because T is conserved at that position in other DtxR-specific operators . The present study examined the effects of nucleotide substitutions at position +7 or -7 in the IRP3 operator . In gel mobility shift assays, only the change of C(+7) to the consensus nucleotide T caused a dramatic increase in the binding of DtxR, whereas other nucleotide substitutions for C(+7) or replacements for A(-7) had only small positive or negative effects on DtxR binding . All substitutions for C(+7) or A(-7) except for A(-7)C dramatically decreased IRP3 promoter strength . In contrast, the A(-7)C variant caused increased promoter strength at the cost of nearly eliminating repressibility by DtxR . The message (sense) strand of the IRP1 operator (5'-TTAGGTTAGCCAAACCTTT-3') includes the -35 region of the IRP3 promoter . A T(+7)C variant of the IRP1 operator was also constructed, and it was shown to exhibit decreased binding to DtxR, decreased repressibility by DtxR, and increased promoter strength . The nucleotides at positions +7 and -7 in DtxR-specific operators are therefore important determinants of DtxR binding and repressibility of transcription by DtxR, and they also have significant effects on promoter activity for IRP3 and IRP1. Microbiology, 1999 Dec, 145 ( Pt 12), 3399 - 408 S-layer protein production by Corynebacterium strains is dependent on the carbon source; Soual-Hoebeke E et al.; Three strains of Corynebacterium producing various amounts of PS2 S-layer protein were studied . For all strains, more PS2 was produced if the bacteria were grown in minimal medium supplemented with lactate than if they were grown in minimal medium supplemented with glucose . The consumption of substrate and PS2 production was studied in cultures with mixed carbon sources . It was found that the inhibitory effect of glucose consumption was stronger than the stimulatory effect of lactate in one strain, but not in the other two strains . The regulation of gene expression involved in S-layer formation may involve metabolic pathways, which probably differ between strains . S-layer organization was also studied by freeze-fracture electron microscopy . It was found that low levels of PS2 production correlated with the partial covering of the cell surface by a crystalline array . Finally, it was found that PS2 production was mainly regulated by changes in gene expression and that secretion was probably not a limiting step in PS2 accumulation. J Biol Chem, 2000 Jan 14, 275(2), 735 - 41 Osmosensor and osmoregulator properties of the betaine carrier BetP from Corynebacterium glutamicum in proteoliposomes; Rubenhagen R et al.; The secondary glycine betaine uptake system BetP of Corynebacterium glutamicum was purified from Escherichia coli membranes in strep-tagged form after heterologous expression of the betP gene and was reconstituted in E . coli lipids . BetP retained its kinetic properties (V(max) and K(m) for betaine and Na(+)) as compared with intact cells . The influence of driving forces (Na(+) gradient and/or electrical potential) on betaine uptake was quantified in proteoliposomes . BetP was effectively regulated by the external osmolality and was stimulated by the local anesthetic tetracaine . A shift of the optimum of osmotic stimulation to higher osmolalities was linearly correlated with an increasing share of phosphatidyl glycerol, the major lipid of the C . glutamicum plasma membrane in the E . coli lipid proteoliposomes . This finding correlates with results demonstrating an identical shift when betP was expressed in E . coli instead of C . glutamicum . These data indicate that (i) BetP comprises all elements of osmosensing and osmoregulatory mechanisms of betaine uptake, (ii) osmoregulation of BetP is directly related to protein/membrane interactions, (iii) the turgor pressure presumably plays no major role in osmoregulation of BetP, and (iv) the regulatory properties of BetP may be related to the physical state of the surrounding membrane. Infection, 1999 Nov-Dec, 27(6), 361 - 4 Diphtheria immunity of Albanian and other eastern European immigrants in Greece compared with the local population--the risk of reemergence in Greece; Pournaras S et al.; A large number of immigrants have come to Greece from diphtheria-endemic countries during the past 8 years . Information on the immune status against diphtheria among immigrants from Albania and the New Independent States (NIS) of the former USSR, as well as the Greek population, is essential in planning public health control measures . The main objective of the study was to determine the risk of the spread of diphtheria if new cases arrived in the country . Diphtheria antitoxin (DAT) in sera from 185 healthy individuals from northern Greece and 227 immigrants from Albania and the NIS was titrated using a quantitative ELISA test . The participants were adults aged 22 to 46 years . Titers were considered as protective when DAT levels were higher or equal to 0.1 IU/ml . Albanian immigrants exhibited the highest rates of protective titers (99 of 167, 59.3%), while lower immunity rates were detected among immigrants from the NIS (20 of 60, 33.3%) (1.5 < OR = 2.9 < 5.4, p = 0.0007) as well as among Greek participants (47 of 185, 25.4%) (2.8 < OR = 4.4 < 7.0, p = 0.00001) . Immunity rates did not differ between Greek and NIS participants . DAT levels of persons with protective titers differed between the three population groups (analysis of variance, p = 0.0005), with the highest mean DAT values detected among NIS immigrants (0.613 IU/ml) . Lower values were found among protected Albanians (0.482 IU/ml) and Greeks (0.314 IU/ml) . There was no significant age- and sex-related difference in immunity rates and DAT levels among the groups . The low immunity rates among Greek adults might suggest that they have had no natural contact with toxigenic strains of corynebacteria . A booster dose of diphtheria toxoid for local adults is recommended, to reduce their risk of acquiring toxigenic strains from individuals who may carry the bacteria without exhibiting clinical disease. Heart . 2000 Jan;83(1):E3. Mycotic aneurysm of the ascending aorta following CABG; Prech M et al.; Mycotic aneurysm of the thoracic aorta is a rare and life threatening condition . Two patients are presented (both male, aged 66 and 59 years) in whom coronary artery bypass surgery was complicated by the development of a mycotic aneurysm . Fever preceded the radiological and echocardiographic signs of the aneurysm by at least several months in both cases . Blood cultures were negative for one patient and the source of Corynebacterium sp infection in the other was not determined for several months . Both patients died before surgery could correct the aneurysm. Eur J Clin Microbiol Infect Dis, 1999 Nov, 18(11), 761 - 70 Class 1 integrons, gene cassettes, mobility, and epidemiology; Fluit AC et al.; Integrons are genetic elements that, although unable to move themselves, contain gene cassettes that can be mobilized to other integrons or to secondary sites in the bacterial genome . The majority of approximately 60 known gene cassettes encode resistance to antibiotics . Recently, a number of gene cassettes encoding extended-spectrum beta-lactamases or carbapenemases have been described . Up to at least five cassettes may be present in an integron, which leads to multiresistance . Frequently, more than one integron is observed within the same bacterial cell . Integrons are widespread in their species distribution . Although integrons are normally reported from Enterobacteriaceae and other gram-negative bacteria, an integron has been described in Corynebacterium glutamicum, a gram-positive species . The gene cassette in this integron showed even higher expression when compared to the expression in Escherichia coli . Integrons have been reported from all continents and are found frequently . The widespread occurrence of integrons is thought to be due to their association with transposon plasmids, conjugative plasmids, or both . Integrons form an important source for the spread of antibiotic resistance, at least in gram-negative bacteria but also potentially in gram-positive bacteria . The aim of this review is to describe the versatility of integrons, especially their mobility and their ability to collect resistance genes. Commun Dis Public Health, 1999 Dec, 2(4), 250 - 7 Laboratory guidelines for the diagnosis of infections caused by Corynebacterium diphtheriae and C . ulcerans . World Health Organization; Efstratiou A et al.; These guidelines represent an application of the World Health Organization European Region's manual for the laboratory diagnosis of diphtheria for laboratories in the United Kingdom (UK), but they could be applied to laboratories overseas . The manual was rewritten in response to the re-emergence of diphtheria in eastern Europe and the emergence of other infections caused by Corynebacterium diphtheriae and C . ulcerans in the UK and overseas . The guidelines summarise our current recommendations and procedures for the microbiological diagnosis of infections caused by toxigenic and non-toxigenic isolates of corynebacteria, with particular reference to C . diphtheriae and C . ulcerans. J Vet Med Sci, 1999 Nov, 61(11), 1203 - 8 Tumor necrosis factor alpha and gamma interferon are required for the development of protective immunity to secondary Corynebacterium pseudotuberculosis infection in mice; Lan DT et al.; The production and role of endogenous cytokines during the course of secondary Corynebacterium (C.) pseudotuberculosis infection were investigated in mice . When immunized mice were challenged on day 28 after primary infection, tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) were found to appear at 3 hr and to reach the maximum at 24 hr after challenge . Spleen cells of mice primarily infected from 2 to 8 weeks before produced a significant amount of TNF-alpha and IFN-gamma when stimulated with formalin-killed bacteria . However, they could not produce detectable amounts of IL-4 . The administration of anti-TNF-alpha monoclonal antibody (MAb) and IFN-gamma MAb increased bacterial proliferation in the organs of immune mice and exacerbated the secondary infection . Injection of anti-CD4 MAb alone or anti-CD4 plus anti-CD8 MAbs resulted in significantly increased mortality and a marked suppression of bacterial elimination as well as cytokine production of secondarily infected mice, while the treatment with anti-CD8 MAb alone showed no effect on either the resistance or cytokine production of mice . These results suggest that CD4, probably Th1 T cells, play an important role for establishment of protective immunity against secondary C . pseudotuberculosis infection by secreting TNF-alpha and IFN-gamma. J Chromatogr A, 1999 Nov 26, 863(2), 243 - 8 Pyrolysis-gas-liquid chromatography with atomic emission detection for the identification of Corynebacterium species; Voisin S et al.; We report here the application of pyrolysis-gas chromatography followed by atomic emission detection (AED) for the characterisation of microorganisms . AED measured the quantity of carbon, sulfur and nitrogen in the molecules separated chromatographically . Twenty-three strains, representing eight Corynebacterium species, were tested in this preliminary study . Co-ordinate principal analysis grouped 11 strains in their respective species group . Most of the other strains appear randomly distributed, perhaps because these strains require additional nutrients . These preliminary results show that the method could be used as a tool for the taxonomic and perhaps the epidemiologic characterisation of bacteria. Am J Nephrol, 1999, 19(6), 682 - 5 Outcome of an opportunistic infection after polymicrobial peritonitis in an HIV-infected patient treated with peritoneal dialysis; Varela MP et al.; The prevalence of human immuodeficiency virus (HIV)-infected patients with end stage renal disease (ESRD) is likely to increase and many of them will be on peritoneal dialysis as renal replacement therapy . Infectious complications are a major problem associated with peritoneal dialysis (PD) . It has been speculated that the HIV-positive peritoneal dialysis population may develop peritonitis more frequently than other peritoneal dialysis patients . We present the complications and unexpected good response to medical management of PD-associated peritonitis in a young HIV-infected black male . He had two unusual and serious infections; the first was a polymicrobial peritonitis which predisposed the patient to an unusual infection caused by Corynebacteria JK for which he was successfully treated without catheter removal . Copyright Rev Sci Tech, 1999 Dec, 18(3), 681 - 90 The role of houseflies (Musca domestica) in harbouring Corynebacterium pseudotuberculosis in dairy herds in Israel; Braverman Y et al.; A study was conducted to assess the role of houseflies, Musca domestica L . in harbouring Corynebacterium pseudotuberculosis in dairy farms in Israel . The bacterium was isolated in June 1993 from 40 wild houseflies which had fed on a lesion on a cow, and from 28 laboratory flies fed on contaminated milk from a cow infected with mastitis . The bacterium was recovered from the body surface of 10 flies (of a total of 160) 10 min after being dipped entirely in a bacterial broth . The bacterium was recovered from the body surface of 10 flies (of a total of 40) 5 min after being fed on contaminated milk . When 110 flies were fed on contaminated sugar cubes, the bacterium was recovered externally from 70 flies 5 min later, and from an additional 20 flies 10 min after feeding . Of 110 flies, 80 excreted bacteria in saliva from 5 min to 3 h after feeding on contaminated milk . Bacteria were isolated from the intestine of 40 of 60 flies between 1 h and 4 h after feeding on contaminated milk . Bacteria were found in the faeces of 30 of 60 flies, between 1 h and 4 h after feeding on contaminated milk . In the light of these findings, and given the fact that this species of fly has a predilection to feed on milk residues of cow teats, the authors concluded that the housefly plays an important role in harbouring and disseminating C . pseudotuberculosis in dairy herds in Israel . In contrast, stable flies (Stomoxys calcitrans L.) are not important in the habouring and dissemination of the bacteria, since bacteria were not recovered 5, 10, 15, 30 min, 2 h or 24 h after membrane feeding on a mixture of bacterial broth and blood. Arch Pediatr, 1999 Nov, 6(11), 1196 - 8 {Acute Corynebacterium diphtheriae aortic endocarditis with negative blood cultures}; Durandy Y et al.; Acute endocarditis with negative blood culture is a challenge requiring close cooperation between several specialists . CASE REPORT: A 15-year-old boy presented with an acute aortic endocarditis due to Corynebacterium diphtheriae . Blood cultures were negative due to an empiric anti-infective therapy . Valvular replacement with a mechanical prosthesis failed to cure the sepsis, which resolved after a cryopreserved allograft implantation . CONCLUSION: This case underlines the efficiency of the highly specialized bacteriological centers and the advantages of using a cryopreserved allograft during the septic state. Mol Gen Genet, 1999 Oct, 262(3), 568 - 78 The Corynebacterium glutamicum insertion sequence ISCg2 prefers conserved target sequences located adjacent to genes involved in aspartate and glutamate metabolism; Quast K et al.; An IS element, termed ISCg2, was identified in the chromosome of Corynebacterium glutamicum ATCC 13032 . After screening a cosmid library of the C . glutamicum ATCC 13032 genome, six copies of ISCg2 including their flanking regions were sequenced and analyzed . ISCg2 is 1636 bp in length and has 26-bp imperfect inverted repeats flanked by 3-bp direct repeats . By comparisons with other IS elements, ISCg2 was classified as a member of the IS30 family of insertion sequences . The six copies of ISCg2 were identical at the nucleotide level and were located in intergenic, AT-rich regions of the chromosome . The regions in which the six copies of ISCg2 were inserted displayed significant similarities . This similarity extends over a region of 65 bp, which was assumed to be the target region for ISCg2 . Interestingly, five of the six copies of ISCg2 were located adjacent to genes that may be involved in aspartate and glutamate metabolism or its regulation . Investigation of the distribution of ISCg2 showed that the IS element is restricted to certain C . glutamicum strains . Analysis of various integration regions indicates active transposition of ISCg2 in C . glutamicum. Biosci Biotechnol Biochem, 1999 Oct, 63(10), 1806 - 10 Molecular analysis of the Corynebacterium glutamicum t