Anticancer Drugs, 1993 Feb, 4(1), 65 - 75 Targeting of GM2-bearing tumor cells with the cytolytic Clostridium perfringens delta toxin; Jolivet-Reynaud C et al.; The cytolytic Clostridium perfringens delta toxin lyses selectively cells which express ganglioside GM2 . In this study, we investigated whether delta toxin can be used to characterize GM2 on tumor cell membranes and as an antitumor agent . The sensitivity to lysis by delta toxin of various murine and human malignant cell lines and also normal tissues was quantified using a 51Cr-release assay . The cytotoxicity titers were correlated with the 125I-labeled toxin binding capacity of sensitive and insensitive cells . Seven of eight human melanomas tested were lysed by the toxin and, of these, four were very sensitive (cytotoxicity titers below 12 ng of toxin) . All neuroblastomas, gliomas and the retinoblastoma tested were lysed with 3-18 ng of toxin . Three of six carcinomas and one of two sarcomas were also very sensitive (cytotoxicity titers 0.6-15 ng) whereas leukemias and lymphoma cells were insensitive . Normal human tissues were insensitive (erythrocytes, skin fibroblasts) or poorly sensitive (brain, lung, spleen) . The in vivo antitumor activity of delta toxin was tested in tumor-bearing mice . Daily intra-tumor injections of 0.5-1 mg of toxin for 4-5 days in carcinoma Me180- and melanoma A375-bearing nude mice, and neuroblastoma C1300-bearing A/J mice significantly inhibited tumor growth for 12-36 days . Intravenous administration of 100 ng of toxin per day for 5 days in Me180-bearing nude mice and C1300-bearing A/J mice gave significant inhibition of tumor growth only during the treatment period, and 10 injections of the same dose of toxin had no significant effect on SK-MEL28, a tumor lacking GM2. Toxicon, 1993 Feb, 31(2), 181 - 6 Production of monoclonal antibodies against Clostridium difficile cytotoxin using immunosorbent binding bioassay procedure; Depitre C et al.; In the following study, a novel screening approach was used to develop monoclonal antibodies specific for toxin B of Clostridium difficile . The approach, which consisted of an immunosorbent binding bioassay (ISBBA), is based on antigen immunocapture by monoclonal antibodies and detection of biological activity . Our results showed ISBBA, which uses unpurified antigen, to be more sensitive than the neutralization assay and ELISA for the detection of toxin B antibody. FEMS Microbiol Lett, 1993 Feb 1, 106(3), 327 - 33 Development of a selective and sensitive polymerase chain reaction assay for the detection of Mycoplasma pirum; Grau O et al.; A new assay using the polymerase chain reaction to amplify a 173-nucleotide DNA fragment within the 16S ribosomal RNA gene of Mycoplasma pirum has been developed . The assay selectively amplified DNA from all strains of M . pirum tested with a high level of sensitivity, even in a context of human DNA . DNA from other mollicute species, including those closely related to M . pirum, from bacteria phylogenetically close to mollicutes (Clostridium innocuum, C . ramosum and Bacillus subtilis), from Escherichia coli and from human peripheral blood mononuclear cells, did not produce the amplified DNA product specific for M . pirum. FEMS Microbiol Lett, 1993 Feb 1, 106(3), 265 - 70 Bioluminescence (lux) expression in the anaerobe Clostridium perfringens; Phillips-Jones MK; To determine whether bacterial luciferase is expressed in the anaerobe Clostridium perfringens to produce an oxygen-requiring bioluminescence reaction, a suitable plasmid vector possessing the luxA and luxB genes of Vibrio fischeri was constructed and introduced into C . perfringens cells . luxAB were placed under the transcriptional control of the C . perfringens alpha-toxin gene promoter region . Suitable ribosome binding sites were introduced upstream of both genes . Bioluminescence was strongly expressed in C . perfringens transformants . Comparisons of in vivo and in vitro bioluminescence measurements demonstrated that in vivo data constituted a quantitative measure of gene expression . This is the first study to show that luxA and luxB genes can be expressed in an anaerobic bacterium and that bioluminescence can be used as a quantitative reporter system in future in vivo studies of gene expression in C . perfringens. Anaesth Intensive Care, 1993 Feb, 21(1), 117 - 9 Spontaneous Clostridium perfringens lung abscess unresponsive to penicillin; Baldwin L et al.; Clostridial necrotising pneumonia is a rare complication of aspiration, bronchial tumour or foreign body, pulmonary infarction, trauma and debilitating medical conditions . Although spontaneous clostridial pneumonia has been reported previously, close scrutiny of those case reports suggests that most of the patients had a recognised predisposing cause . We report a case of true spontaneous Clostridium perfringens pneumonia complicated by septic shock, pneumothorax and pulmonary necrosis . The patient responded poorly to conventional treatment with benzylpenicillin, and although the addition of metronidazole produced dramatic resolution of the sepsis, lobectomy was required to effect cure. J Appl Bacteriol, 1993 Feb, 74(2), 143 - 8 Freeze-dried mixed cultures as reference samples in quantitative and qualitative microbiological examinations of food; Peterz M et al.; Two simulated food samples in the form of mixtures of bacteria, freeze-dried in small glass vials, were tested for their suitability as reference materials in food microbiology . Stability and homogeneity of the samples were assessed during a 1 year period when stored at 2-6 degrees C . Samples were examined for aerobic plate counts, coliform counts, Staphylococcus aureus, Clostridium perfringens, Bacillus cereus, enterococci, yeasts and salmonellas . The samples were homogeneous but the number of colony forming units of some organisms declined during the time period studied . The maximum rate of decline (0.8 log units per year) was observed for the coliforms . Precision estimates (reproducibility and repeatability) for the different testing methods used are given . When compared with these data, the rate of decline was considered to be of less importance. Proteins, 1993 Feb, 15(2), 183 - 90 A novel parameterization scheme for energy equations and its use to calculate the structure of protein molecules; Snow ME; A novel scheme for the parameterization of a type of "potential energy" function for protein molecules is introduced . The function is parameterized based on the known conformations of previously determined protein structures and their sequence similarity to a molecule whose conformation is to be calculated . Once parameterized, minima of the potential energy function can be located using a version of simulated annealing which has been previously shown to locate global and near-global minima with the given functional form . As a test problem, the potential was parameterized based on the known structures of the rubredoxins from Desulfovibrio vulgaris, Desulfovibrio desulfuricans, and Clostridium pasteurianum, which vary from 45 to 54 amino acids in length, and the sequence alignments of these molecules with the rubredoxin sequence from Desulfovibrio gigas . Since the Desulfovibrio gigas rubredeoxin conformation has also been determined, it is possible to check the accuracy of the results . Ten simulated-annealing runs from random starting conformations were performed . Seven of the 10 resultant conformations have an all-C alpha rms deviation from the crystallographically determined conformation of less than 1.7 A . For five of the structures, the rms deviation is less than 0.8 A . Four of the structures have conformations which are virtually identical to each other except for the position of the carboxy-terminal residue . This is also the conformation which is achieved if the determined crystal structure is minimized with the same potential . The all-C alpha rms difference between the crystal and minimized crystal structures is 0.6 A.(ABSTRACT TRUNCATED AT 250 WORDS) J Gen Microbiol, 1993 Feb, 139 ( Pt 2), 307 - 16 Gene sequence and properties of CelI, a family E endoglucanase from Clostridium thermocellum; Hazlewood GP et al.; The Clostridium thermocellum celI gene, coding for endoglucanase I (CelI), consists of an open reading frame (ORF) of 2640 nucleotides and codes for a protein of M(r) 98531 . The ORF was confirmed as celI by comparing the N-terminal sequence of purified recombinant CelI with that deduced from the nucleotide sequence . CelI hydrolysed lichenan and carboxymethylcellulose, but was principally active against barley beta-glucan . It exhibited significant sequence identity with subfamily E2 endoglucanases, and by analogy with others in this group contains a catalytic domain of around 500 residues located in the N-terminal half of the protein . The C-terminal region of CelI was highly homologous with the cellulose-binding domain of the non-catalytic cellulosome subunit, S1 . A repeated segment, previously shown to be highly conserved in xylanase Z and in other endoglucanases from C . thermocellum, was absent from CelI . Antiserum raised against purified recombinant CelI cross-reacted with proteins contained in the cellulosomes of two strains of C . thermocellu, suggesting that CelI is either a component of the cellulosome or is homologous to other cellulosome proteins . A second gene, located upstream of celI, consisted of an ORF of 1671 nucleotides, coding for a protein of M(r) 61042 . Based on its homology with the Escherichia coli tar gene product, the polypeptide encoded by the second gene is tentatively identified as a sensory transducer. Appl Environ Microbiol, 1993 Feb, 59(2), 623 - 7 Cloning of the beta-amylase gene from Bacillus cereus and characteristics of the primary structure of the enzyme; Nanmori T et al.; The gene encoding the beta-amylase of Bacillus cereus BQ10-S1 (SpoII) was cloned into Escherichia coli JM 109 . A sequenced DNA fragment of 2,001 bp contains the beta-amylase gene . The N-terminal sequences (AVNGKG MNPDYKAYLMAPLKKI), the C-terminal sequences (SHTSSW), and the amino acid sequences of the five regions in the beta-amylase molecules were determined . The mature beta-amylase contains 514 amino acid residues with a molecular mass of 57,885 Da . The amino acid sequence homology with those of known beta-amylases was 52.7% for Bacillus polymyxa, 52.0% for Bacillus circulans, 43.4% for Clostridium thermosulfurogenes, 31.8% for Arabidopsis thaliana, 31.5% for barley, 29.9% for sweet potato, and 28.9% for soybean . Ten well-conserved regions were found between the N terminus and the area around residue 430, but the C-terminal region of 90 residues has no similarity with those of the plant beta-amylases . The homology search revealed that this C-terminal region has homology with C-terminal regions of the beta-amylase from C . thermosulfurogenes, some bacterial alpha-amylases, cyclodextrin glucanotransferase, and glucoamylase . Some of these sequences are known as the raw-starch-binding domain . These results suggest that B . cereus beta-amylase has an extra domain which has raw-starch-binding ability and that the domain has considerable sequence homology with those of other amylases or related enzymes from a wide variety of microorganisms. Appl Environ Microbiol, 1993 Feb, 59(2), 496 - 501 Esterase electrophoretic polymorphism of human and animal strains of Clostridium perfringens; Pons JL et al.; Esterase electrophoretic polymorphism in human and animal strains of Clostridium perfringens was studied by using polyacrylamide-agarose gel electrophoresis . Five types of esterases, designated E-I to E-V and defined by their hydrolytic specificities toward five synthetic substrates, were found in protein extracts of bacteria grown without glucose (glucose-containing media allowed only the expression of esterase E-I) . Mobility variants of esterase E-I, which hydrolyzes alpha- and beta-naphthyl acetates and butyrates, were used as a basis for the distribution of strains into 11 zymogroups . When all five types of esterases and their electrophoretic variants were considered, 77 electrophoretic types (ETs) could be described for the 89 strains tested . Animal strains did not constitute a distinctive subpopulation, as revealed by their distribution in the zymogroups and by clustering analysis . Statistical analysis also emphasized the importance of esterase E-IV (which hydrolyzes only naphthyl acetates) and esterase E-V (which hydrolyzes only alpha-naphthyl acetate) in clustering by the relatedness of the ETs . ETs allowed the epidemiological characterization of stool isolates recovered from elderly inpatient residents and from adolescent chronic-care psychiatric patients . These results indicate that esterase electrophoretic typing may be a marker for epidemiological and ecological analyses. J Clin Microbiol, 1993 Feb, 31(2), 413 - 5 Typing of Clostridium difficile by western immunoblotting with 10 different antisera; Kato H et al.; Western blotting (immunoblotting) with antisera against each of 10 reference serogroups was evaluated as a means of typing Clostridium difficile . A total of 164 clinical isolates of C . difficile were tested . Variations in band profiles in each serogroup were used to type isolates into subserogroups . This technique was useful for an epidemiological investigation. J Clin Microbiol, 1993 Feb, 31(2), 315 - 7 Mammalian epithelial cell line kit for detection of Clostridium difficile toxin; Walpita P et al.; The performance characteristics of a mammalian epithelial (MEP) cell line kit (Cytotoxi Test; Advanced Clinical Diagnostics, Toledo, Ohio) for the detection of Clostridium difficile toxin was compared with that of conventional tissue culture assays with human embryonic lung (HEL) cells in shell vials and human foreskin fibroblasts (HFFs) in test tubes . One hundred forty-nine stool samples were tested . The MEP cells were at least as sensitive as the HEL cells for use in C . difficile toxin detection . Results for the MEP cells were also obtained considerably more rapidly than those for HEL cells when the cells were examined at 4 and 24 h and then every 24 h for up to 5 days . Approximately one-third of all positive MEP cells were detected at 4 h and 95% were detected by 48 h . In comparison, in the HEL shell vial monolayers, only 6% of the positive cells were detectable at 4 h and 76% were detectable at 48 h . The times for C . difficile toxin-induced cytotoxicity in HFF cells were similar to those in HEL cells . Shell vials carrying HEL cell monolayers (ViroMed Laboratories Inc., Minnetonka, Minn.) are a sensitive and reliable commercial source for the detection of C . difficile toxin, although they cannot detect C . difficile as rapidly as the Cytotoxi test with the MEP cell monolayers. J Clin Microbiol, 1993 Feb, 31(2), 279 - 82 Enzymatic reactions of Clostridium difficile in aerobic and anaerobic environments with the RapID-ANA II identification system; Peiffer S et al.; The RapID-ANA II anaerobic identification system (Innovative Diagnostic Systems, Inc., Atlanta, Ga.) was used to determine whether the incubation environment affects enzyme detection . Twenty strains of Clostridium difficile were tested in aerobic, anaerobic, and low-CO2 anaerobic incubation environments . The percentages of enzymes detected in reactions with the following substrates were noted in the three incubation environments: phenylalanine-beta-naphthylamide, aerobic, 0%; anaerobic, 35%; low-CO2 anaerobic, 35%; arginine-beta-naphthylamide, aerobic, 5%; anaerobic, 55%; low-CO2 anaerobic, 75%; pyrrolidonyl-beta-naphthylamide, aerobic, 5%; anaerobic, 65%; low-CO2 anaerobic, 65% . When the aerobic incubation environment was compared with either the anaerobic or the low-CO2 anaerobic incubation environments, the results were statistically different with respect to enzyme detection in reactions with the substrates listed above . The results for the anaerobic and low-CO2 anaerobic environments were not statistically different . The study was repeated twice . Statistical comparisons between the three environments were consistent with the results presented above, with the following exceptions . The aerobic and the anaerobic environments were not different in a reaction with phenylalanine-beta-naphthylamide in one of the runs, and there was no significant difference between the three environments in a reaction with arginine-beta-naphthylamide in another run . These results suggest that some of the enzymes used in the identification of clinical anaerobes appear to be inactive in an environment containing oxygen. Ann Thorac Surg, 1993 Feb, 55(2), 476 - 81 Surgical management and radiological characteristics of bronchogenic cysts; Suen HC et al.; Forty-two patients with bronchogenic cysts were treated over a 30-year period (1962 to 1991) . The location was mediastinal in 37 and intrapulmonary in 5 . Cysts were symptomatic in 21 patients (50%) and complications occurred in 11 (26%) . The complications included infection in 5 patients, hemorrhage into the cyst in 2 patients, dysphagia due to esophageal compression in 2, adenocarcinoma arising from a bronchogenic cyst in an 8 1/2-year-old girl, and an esophagobronchopleurocutaneous fistula as a result of previous incomplete resection in 1 patient . Magnetic resonance imaging has been found to provide specific diagnostic information about bronchogenic cysts . All but 2 patients were treated with complete excision . One patient was managed by observation and another had drainage of the cyst by mediastinoscopy . Complications of treatment occurred in only 2 patients . One had a minor wound infection and the other had Clostridium difficile enterocolitis . Only 4 patients were lost to follow-up . No late complication or recurrence developed in those patients having complete excision . We recommend complete excision in most instances to confirm the diagnosis, relieve symptoms, and prevent complications. Proc Natl Acad Sci U S A, 1993 Feb 1, 90(3), 1078 - 82 The unusual metal clusters of nitrogenase: structural features revealed by x-ray anomalous diffraction studies of the MoFe protein from Clostridium pasteurianum; Bolin JT et al.; Nitrogenase (EC 1.18.6.1) catalyzes the conversion of dinitrogen to ammonia, the central reaction of biological nitrogen fixation . X-ray anomalous diffraction data were analyzed to probe the structures of the metal clusters bound by nitrogenase MoFe protein . In addition to one FeMo cofactor, each half-molecule of MoFe protein binds one large FeS cluster of a type not previously observed in a protein . The FeS cluster contains roughly eight Fe atoms, comprises two subclusters, and is separated from the FeMo cofactor by an edge-to-edge distance of 14 A . The inorganic framework of the FeMo cofactor is not resolved into subclusters, but the Mo atom is located at its periphery . FeMo cofactors in different half-molecules are 70 A apart and cannot promote binuclear activation of dinitrogen by two Mo atoms. J Med Microbiol, 1993 Feb, 38(2), 109 - 13 Characterisation of Clostridium difficile strains by polymerase chain reaction with toxin A- and B-specific primers; Wren BW et al.; A total of 218 Clostridium difficile strains was examined for production of toxin A by ELISA, production of toxin B by a cytotoxin assay and the presence of toxin A and B gene-associated sequences by the polymerase chain reaction (PCR) . After saturation amplification with toxin B-specific primers, the characteristic amplification product (591 bp) was detected in all 184 toxigenic strains examined . PCR with toxin A-specific primers gave positive results with all but one of the toxigenic strains . By contrast, PCR with toxin A- and toxin B-specific primers yielded negative results with all 34 non-toxigenic strains tested . This suggests that PCR detection of either the toxin A or B gene is a good indication of toxin production . PCR did not require DNA extraction or hybridisation and was convenient, sensitive and rapid . Toxigenic C . difficile could be detected in mixed cultures, suggesting a role for PCR in the identification of toxigenic C . difficile in primary culture. J Med Microbiol, 1993 Feb, 38(2), 103 - 8 In-vitro and in-vivo characterisation of resistance to colonisation with Clostridium difficile; Larson HE et al.; In hamsters, resistance to colonisation by Clostridium difficile appears to be mediated by micro-organisms that are present in the gut in relatively low concentrations . Small amounts of normal caecal contents inhibited the growth of C . difficile when added to cultures in vitro or given to animals which had been treated with clindamycin . Filtrates of caecal contents, frozen and thawed contents and contents diluted to 0.1% wet weight lost their inhibitory properties . However, caecal contents retained their protective capacity after culture for 7 days in vitro . Antibiotic treatment altered resistance to colonisation by only a few species of clostridia . Faeces of animals treated with ampicillin but not clindamycin recovered colonisation resistance after incubation at 37 degrees C in vitro . Since human faeces could also restore colonisation resistance to hamsters, the hamster model may be useful for the study of resistance to colonisation by C . difficile in man. Infect Immun, 1993 Feb, 61(2), 457 - 63 Comparison of the alpha-toxin genes of Clostridium perfringens type A and C strains: evidence for extragenic regulation of transcription; Katayama S et al.; The Clostridium perfringens plc gene encoding phospholipase C (alpha-toxin) was cloned from type C NCIB 10662, a strain which produces low levels of phospholipase C activity . The nucleotide sequence of a cloned 3.1-kb HindIII fragment was determined . The same fragment was also cloned from type A NCTC 8237, a phospholipase C-overproducing strain . In this case, an open reading frame (ORF2) truncated in the previously cloned 2-kb fragment was also sequenced . Comparison of the nucleotide sequence between the 3.1-kb fragments of the two type strains shows some differences both in the plc gene and in ORF2 . However, when the 3.1-kb fragment was cloned into plasmid pUC19 and expressed in Escherichia coli, the plc genes from both type strains were similarly expressed and the toxins produced showed similar levels of activity . Northern blot analysis revealed that the type A strain produced 16 to 23 times more plc mRNA than the type C strain . These results indicate that in C . perfringens the two plc genes are transcribed at different rates, probably because of a difference in a locus lying outside of the cloned fragments . Gel retardation analysis showed that the type A strain possessed two different proteins that bound different regions of the plc gene . However, one of these proteins, which binds within the plc coding region, was not found in the type C strain, suggesting that it plays a role in the regulation of the plc gene expression. J Infect Dis, 1993 Feb, 167(2), 455 - 8 Detection of toxigenic Clostridium difficile in stool specimens by the polymerase chain reaction; Kato N et al.; Polymerase chain reaction (PCR) amplification of a segment of the toxin A gene was used to detect toxigenic Clostridium difficile directly from stool specimens of patients with antibiotic-associated diarrhea . Although PCR-inhibitory substances were recognized in DNA prepared from stool specimens, the inhibitory substances were eliminated by using an ion-exchange column after phenol-chloroform extraction . Eventually, 39 stool specimens were evaluated by PCR . PCR results for detection of toxigenic C . difficile were in complete agreement with cell culture assay results; all 12 PCR-positive stool specimens were positive by cytotoxin assay, and all 27 PCR-negative specimens were negative by cytotoxin assay . Toxigenic C . difficile was cultured from all PCR-positive specimens . These results suggest that PCR amplification may be an effective method for laboratory diagnosis of C . difficile-associated diarrhea and colitis. J Infect Dis, 1993 Feb, 167(2), 451 - 4 A massive outbreak of type E botulism associated with traditional salted fish in Cairo; Weber JT et al.; In April 1991, 91 hospitalized patients in Cairo were reported to the Egyptian Ministry of Health with botulism intoxication . To define the spectrum of illness and identify a food vehicle, 45 patients were interviewed and a case-control investigation was conducted among families of 5 hospitalized patients . Clinical specimens and specimens of implicated food were tested for toxin and cultured for Clostridium botulinum . Hospitalized patients had symptoms consistent with botulism; 18 (20%) of 91 reported patients died . Illness was associated with eating faseikh (uneviscerated, salted mullet fish; lower 95% confidence limit of odds ratio = 6.6, P < .001) . All 5 case-families purchased faseikh from one shop . Very high levels of type E botulinal toxin were detected in faseikh reported to be purchased from the implicated shop; C . botulinum type E was isolated from cultures of clinical specimens and from the faseikh . This is the first documented outbreak of botulism in Egypt and the largest type E outbreak ever reported. Mol Microbiol, 1993 Feb, 7(3), 447 - 59 Characterization of genes involved in molybdenum transport in Azotobacter vinelandii; Luque F et al.; Expression of alternative nitrogenases in Azotobacter vinelandii is repressed by molybdenum . Two strains with Tn5 insertion mutations showed alternative nitrogenase-dependent diazotrophic growth in the presence of Mo . The mutations were in a region which contained four open reading frames (ORFs 1-4) . The genetic structure and predicted products of ORFs 2, 3 and 4 are typical of the membrane-associated elements of the ATP-binding cassette (ABC) superfamily of transport systems . The products of ORF3 and ORF4 are homologous with the products of the Escherichia coli genes chlD and the partially sequenced chlJ, respectively, both of which are implicated in molybdenum transport . ORF1, which is in the relative position of bacterial permease genes commonly specifying periplasmic binding proteins, encodes a 29 kDa protein with a novel primary structure . It lacks a potential signal sequence, and its C-terminal half consists of a tandem repeat of a segment which is homologous with the M(r) 7 kDa molybdenum-pterin binding protein Mop from Clostridium pasteurianum . This suggests that a substituted pterin may be involved in the initial capture or early metabolism of molybdenum. Glycoconj J, 1993 Feb, 10(1), 50 - 6 Isolation and properties of the natural and the recombinant sialidase from Clostridium septicum NC 0054714; Zenz KI et al.; The natural sialidase of Clostridium septicum was purified and characterized in parallel with the recombinant enzyme expressed by Escherichia coli . The two enzymes exhibit almost identical properties . The maximum hydrolytic activity was measured at 37 degrees C in 60 mM sodium acetate buffer, pH 5.3 . Glycoproteins like fetuin and saponified bovine submandibular gland mucin, most of them having alpha(2-6) linked sialic acids, are preferred substrates, while sialic acids from gangliosides, sialyllactoses, or the alpha(2-8) linked sialic acid polymer (colominic acid) are hydrolysed at lower rates . alpha(2-3) Linkages are more rapidly hydrolysed than alpha(2-6) bonds of sialyllactoses . The cleavage rate is markedly reduced by O-acetylation of the sialic acid moiety . These properties are similar to those of other secreted clostridial sialidases . The enzyme exists in mono-, di- and trimeric forms, the monomer exhibiting a molecular mass of 125 kDa, which is close to the protein mass of 111 kDa deduced from the nucleotide sequence of the cloned gene. Glycoconj J, 1993 Feb, 10(1), 40 - 4 Inhibition of sialidases from viral, bacterial and mammalian sources by analogues of 2-deoxy-2,3-didehydro-N-acetylneuraminic acid modified at the C-4 position; Holzer CT et al.; The inhibition of sialidase activity from influenza viruses A and B, parainfluenza 2 virus, Vibrio cholerae, Arthrobacter ureafaciens, Clostridium perfringens, and sheep liver by a range of 2-deoxy-2,3-didehydro-N-acetylneuraminic acid analogues modified at the C-4 position has been studied . All substitutions tested resulted in a decrease in the degree of inhibition of the bacterial and mammalian sialidases . For sialidases from influenza viruses A and B, on the other hand, most of the substitutions tested either had no significant effect on binding or, in the case of the basic amino and guanidino substituents, resulted in significantly stronger inhibition . The results for parainfluenza 2 virus sialidase were mostly intermediate, in that inhibition was neither significantly increased nor decreased by most of the modifications . We conclude that only the influenza A and B sialidase active sites possess acid groups correctly positioned to participate in charge-charge interactions in the region of C-4 of bound substrate, and that the C-4 binding pockets of the bacterial and mammalian sialidases examined are considerably smaller than is observed for either the influenza virus or parainfluenza virus sialidases. Jpn J Med Sci Biol, 1993 Feb, 46(1), 51 - 6 Immunological comparison of intracellular toxin A and extracellular toxin A from Clostridium difficile; Meng XQ et al.; Intracellular toxin A and extracellular toxin A of Clostridium difficile were compared immunologically . Immunodiffusion tests with anti-intracellular toxin A and anti-extracellular toxin A sera showed that these toxins were identical . In neutralization tests, both antisera neutralized the homologous and heterologous toxins with regard to cytotoxicity, mouse lethality and loop response at nearly the same titers . Although intracellular toxin A lacks the hemagglutination (HA) activity, anti-intracellular toxin A serum neutralized HA activity of extracellular toxin A at the same titer as did anti-extracellular toxin A serum . These results suggest that these toxins are immunologically indistinguishable and that the intracellular toxin A molecule has an antigenic site(s) for the HA activity. Biosci Biotechnol Biochem, 1993 Feb, 57(2), 273 - 7 Nucleotide sequence of the Clostridium stercorarium xynA gene encoding xylanase A: identification of catalytic and cellulose binding domains; Sakka K et al.; The nucleotides of the xynA gene of Clostridium stercorarium were sequenced . The structural gene consists of an open reading frame of 1533 bp encoding 511 amino acids with an M(r) of 56,519 . The signal peptide cleavage site was identified by comparison with the N-terminal amino acid sequence of the enzyme produced by a recombinant Escherichia coli . Xylanase A consists of a catalytic domain belonging to family G at the N-terminus and two direct repeats of about 90 amino acids with a short spacing at the C-terminus . Deletion analysis showed that the repeated sequences were responsible for binding the enzyme to Avicel and were not essential for catalytic activity . The catalytic domain of this enzyme is highly homologous to xylanase A of Clostridium acetobutylicum (identity: 69%) and xylanase B of Bacillus pumilus (identity: 64%). Biosci Biotechnol Biochem, 1993 Feb, 57(2), 268 - 72 Nucleotide sequence of the Clostridium stercorarium xylA gene encoding a bifunctional protein with beta-D-xylosidase and alpha-L-arabinofuranosidase activities, and properties of the translated product; Sakka K et al.; The nucleotides of the beta-xylosidase (xylA) gene from Clostridium stercorarium were sequenced . A single open reading frame of 473 codons specifying the subunit (MW 53,340) of xylosidase was identified . The N-terminal amino acid sequence and molecular weight estimated by SDS-polyacrylamide gel electrophoresis of the purified enzyme were quite in agreement with those deduced from the nucleotide sequence . Analysis of the enzyme by gel filtration on an HPLC column gave a molecular weight of 220,000, suggesting that the native enzyme is a tetramer composed of 4 identical subunits . The pH optimum was 7.0 and quite stable over the pH range of 5 to 10 at 4 degrees C . The optimum temperature was 65 degrees C . Vm was estimated to be 5.9 nmol/min/micrograms for p-nitrophenyl-beta-D-xylopyranoside and 16.7 nmol/min/micrograms for p-nitrophenyl-alpha-L-arabinofuranoside, while Km was estimated to be 2.5 mM for p-nitrophenyl-beta-D-xylopyranoside and 17.6 mM for p-nitrophenyl-alpha-L-arabinofuranoside. Biochem Biophys Res Commun, 1993 Jan 29, 190(2), 609 - 15 Stimulation of rat liver microsomal cGMP-inhibited cAMP phosphodiesterase (PDE III) by phospholipase C and D; Carpenedo F et al.; The specific activity of cGMP-inhibited cAMP phosphodiesterase (PDE III) of rat liver microsomal membranes is increased in a concentration-dependent way by adding phospholipase C from Clostridium perfringens or phospholipase D from Streptomyces chromofuscus . The effect depends on an increase in Vmax of the enzyme . Treatment of microsomal membranes with phospholipase C causes a marked increase in the relative amounts of phosphatidylserine and phosphatidylinositol, and mild stimulation of PDE III activity . Treatment with phospholipase D increases phosphatidic acid and strongly increases PDE III activity . These data suggest that phosphatidic acid is the most important regulator of membrane-bound PDE III activity in liver. Biochim Biophys Acta, 1993 Jan 17, 1175(2), 140 - 6 Identification of heterotrimeric and low molecular weight GTP-binding proteins in rabbit skeletal muscle longitudinal sarcoplasmic reticulum; Kutchai H et al.; Direct photoaffinity labeling of proteins of longitudinal sarcoplasmic reticulum (LSR) of rabbit skeletal muscle with {32P}GTP revealed GTP-binding proteins of about 52, 45 and 30 kDa . ADP-ribosylation with {32P}NAD in the presence of cholera toxin (CTX) or pertussis toxin (PTX) indicates the existence of a CTX substrate (about 45 kDa); no PTX substrates were observed . Western blots of LSR probed with RM/1, an antiserum against a decapeptide from the C-terminus of Gs alpha, showed an immunoreactive band at about 45 kDa . {32P}GTP overlays of Western blots of LSR showed a heavily-labeled protein of about 29 kDa and one or more additional slightly smaller proteins that were more weakly labeled . When LSR was subjected to mild trypsin hydrolysis, the Western blot overlay revealed three bands at about 23, 25 and 29 kDa . Western blots of LSR proteins showed no significant immunoreactivity with the anti-(pan)-ras monoclonal antibodies 142-24E05 and Ras 11 . ADP-ribosylation of LSR with {32P}NAD in the presence of C3 exoenzyme of Clostridium botulinum yielded a labeled band at about 23 kDa . Our results indicate the presence in rabbit LSR of a Gs alpha, the absence of Gi and G(o), and the presence of several low molecular weight GTP-binding proteins, distinct from p21 ras, one of which belongs to the rho or rac family. Eur J Biochem, 1993 Jan 15, 211(1-2), 37 - 45 Flavin dynamics in reduced flavodoxins . A time-resolved polarized fluorescence study; Leenders R et al.; The time-resolved fluorescence and fluorescence anisotropy characteristics of reduced flavin mononucleotide in solution as well as bound in flavodoxins isolated from the bacteria Desulfovibrio gigas, Desulfovibrio vulgaris, Clostridium beijerinckii MP and Megasphaera elsdenii have been examined . All fluorescence and fluorescence anisotropy decays were analyzed by two different methods: (a) least-squares fitting with a sum of exponentials and (b) the maximum entropy method to yield distributed lifetimes and correlation times . The results of both approaches are in excellent agreement . The fluorescence decay of the free as well as protein-bound reduced flavin chromophore is made up of three components . The shortest component proves to be relatively sensitive to the environment and can therefore be used as a diagnostic tool to probe the microenvironment of the reduced isoalloxazine ring system . The other two longer fluorescence lifetime components are insensitive to the chromophore environment and seem therefore to be related to intrinsic, photophysical properties of the reduced chromophore . Fluorescence anisotropy decays show that the flavin mononucleotide in all four reduced flavodoxins is immobilized within the protein matrix, as indicated by the recovery of a single rotational correlation time, reflecting the rotation of the whole protein . No indications are found that rapid structural fluctuations occur in reduced flavodoxins, and the mechanism of electron transfer from flavodoxin to other redox proteins seems to involve immobilized reduced flavin. Gene, 1993 Jan 15, 123(1), 93 - 7 Sequence and arrangement of genes encoding enzymes of the acetone-production pathway of Clostridium acetobutylicum ATCC824; Petersen DJ et al.; The nucleotide sequence of three open reading frames in the acetone-production locus of Clostridium acetobutylicum ATCC824 has been established . The three gene products, corresponding to acetoacetate decarboxylase (EC 4.1.1.4) and both subunits of the acetoacetyl-CoA:acetate/butyrate:CoA transferase (EC 2.8.3.9) are transcribed in two convergently arranged operons . The intervening DNA region separating the two transcripts is characterized by an inverted repeat which appears capable of forming a stem-loop structure functioning as a Rho-independent transcription terminator in both directions. Ugeskr Laeger, 1993 Jan 11, 155(2), 108 - 9 {Botulism in Ammassalik}; Sorensen HC et al.; Botulism is a rare and serious form of food poisoning and was diagnosed for the first time in the East Coast of Greenland . Historical reports suggest that outbreaks of this condition have occurred previously in this region . In 1990, however, the presence of Clostridium botulinum type E could be confirmed with certainty . Eight individuals partook of a meal which consisted of raw seal meat and raw seal intestines . Four of these developed symptoms of botulism and two of these required assisted ventilation . On the basis of this experience, the medical officers of health in Greenland recommend that all hospitals in Greenland should maintain a supply of antitoxin. JAMA, 1993 Jan 6, 269(1), 71 - 5 Diagnosis and treatment of Clostridium difficile colitis; Fekety R et al.; Pseudomembranous colitis associated with antibiotic therapy is almost always due to an overgrowth of Clostridium difficile . If untreated, pseudomembranous colitis can lead to severe diarrhea, hypovolemic shock, toxic dilatation of the colon, cecal perforation, hemorrhage, and death . However, C difficile-associated colitis can mimic the more common "benign" antibiotic-associated diarrhea that is not caused by C difficile . An algorithm for diagnosis management of hospitalized patients with antibiotic diarrhea and C difficile colitis is presented in this review . Diagnosis depends on sigmoidoscopy and/or stool tests for C difficile toxins in all patients with antibiotic-associated diarrhea . If the results of these tests are positive, either metronidazole or vancomycin is recommended for treatment of mild illness, and vancomycin is recommended for treatment of severe illness . Oral therapy is always preferred because it is more reliable . In patients with recurrent or relapsing colitis, treatment with either metronidazole or vancomycin is effective for that episode, but novel approaches, such as the oral or rectal introduction of competing nonpathogenic organisms, may prove to be more successful in prevention of relapses. J Immunol Methods, 1993 Jan 4, 157(1-2), 125 - 33 An immunoassay for the rapid and specific detection of three sialidase-producing clostridia causing gas gangrene; Roggentin T et al.; A rapid and methodologically unusual diagnostic test was developed for the specific detection of Clostridium perfringens, C . septicum and C . sordellii, which cause clostridial myonecrosis . Sialidases (EC 3.2.1.18) secreted by these bacterial species were bound to polyclonal antibodies raised against the respective enzyme and immobilized onto microtiter plates . The activity of bound sialidase was determined with the fluorogenic substrate 4-methylumbelliferyl-alpha-D-N-acetylneuraminic acid . The assay permits the detection of a minimum sialidase activity of about 0.1-1 mU/ml of sample solution within 2 h . The sensitivity of the test was reduced by about three-fold for sialidase activities in samples containing 50% serum . Only a few, low cross-reactivities, which never exceeded 10% of the homologous reaction, were observed with 12 sialidases from other bacterial sources . Clinical isolates of the three clostridial species were analysed by this assay and gave positive signals in the homologous test . The assay for the detection of C . perfringens was applied to nine samples from patients suspected to be suffering from clostridial myonecrosis . There was a high correlation between the results of the immunoassay and the bacteriological analysis of infection. Folia Microbiol (Praha), 1993, 38(1), 15 - 21 Identification of the product of tetP gene: a possible mechanistic basis for tetracycline resistance in Clostridium perfringens; Saksena NK et al.; Using cloning and in vitro protein synthesis we identified the polypeptide product of the tetP gene of Clostridium perfringens which is responsible for conferring resistance to tetracycline . Two EcoRI fragments invariably share the resistance determinant in all of the Clostridium perfringens isolates that we studied . Likewise, two proteins of 10 and 20 kDa were found to be conserved in all of the recombinant clones . The 10 kDa protein appears to be responsible for the constitution of the expression of tetP gene in C . perfringens. Arch Microbiol, 1993, 159(4), 345 - 53 Anaerobic transformation of 2,4,6-trinitrotoluene (TNT); Preuss A et al.; A sulfate-reducing bacterium using trinitrotoluene (TNT) as the sole nitrogen source was isolated with pyruvate and sulfate as the energy sources . The organism was able to reduce TNT to triaminotoluene (TAT) in growing cultures and cell suspensions and to further transform TAT to still unknown products . Pyruvate, H2, or carbon monoxide served as the electron donors for the reduction of TNT . The limiting step in TNT conversion to TAT was the reduction of 2,4-diamino-6-nitrotoluene (2,4-DANT) to triaminotoluene . The reduction proceeded via 2,4-diamino-6-hydroxylaminotoluene (DAHAT) as an intermediate . The intermediary formation of DAHAT was only observed in the presence of carbon monoxide or hydroxylamine, respectively . The reduction of DAHAT to triaminotoluene was inhibited by both CO and NH2OH . The inhibitors as well as DANT and DAHAT significantly inhibited sulfide formation from sulfite . The data were taken as evidence for the involvement of dissimilatory sulfite reductase in the reduction of DANT and/or DAHAT to triaminotoluene . Hydrogenase purified from Clostridium pasteurianum and carbon monoxide dehydrogenase partially purified from Clostridium thermoaceticum also catalyzed the reduction of DANT in the presence of methyl viologen or ferredoxin, however, as the main reduction product DAHAT rather than triaminotoluene was formed . The findings could explain the function of CO as an electron donor for the DANT reduction (to DAHAT) and the concomitant inhibitory effect of CO on triaminotoluene formation (from DAHAT) by the inhibition of sulfite reductase.(ABSTRACT TRUNCATED AT 250 WORDS) Anticancer Res, 1993 Jan-Feb, 13(1), 107 - 11 Antimetastatic activity induced by Clostridium butyricum and characterization of effector cells; Chen HY et al.; The effects of a bacterial vaccine, heat-killed Clostridium butyricum MII 588 cells, on the metastasis of B16-F10 melanoma in BDF1 mice was investigated . The vaccine stimulated natural killer (NK) cell cytotoxic activity against YAC-1 target cells, which peaked at 72 hr after the pretreatment, whereas maximum macrophage cytotoxic activity was obtained on days 9 to 11 . These stimulated cytotoxic activities were also observed in B16-F10 tumor-bearing BDF1 mice . The important role of stimulated NK cells and/or macrophage in the antimetastatic effect was confirmed using Anti-asialo GM1 antibody, whole body x-ray irradiation and carrageenan treatment . In addition, the vaccine could induce a high titer of interferon-gamma (IFN-gamma), an important lymphokine which may account for a significant portion of its antimetastatic activity. Parasitol Res, 1993, 79(2), 114 - 8 Effect of various digestive enzymes on the interaction of Toxoplasma gondii with macrophages; de Carvalho L et al.; The participation of resident, elicited, and activated macrophage surface components during internalization of tachyzoites of Toxoplasma gondii was analyzed using neuraminidase, phospholipase C, trypsin, protease, and hyaluronidase . Treatment of these macrophages with neuraminidase from Vibrio cholerae, phospholipase C from Bacillus cereus and Clostridium perfringens, protease, and hyaluronidase prior to their interaction with parasites increased the penetration of host cells by T . gondii . Incubation of macrophages with trypsin significantly inhibited the uptake of parasites . Our findings confirm previous observations that treatment of the macrophages with cytochalasin D under conditions that completely block the typical phagocytic process partially inhibits infection of the cells by T . gondii . The results of simultaneous treatment of the macrophages with enzymes and cytochalasin D suggested that the observed enhancement of cell infection by treatment with neuraminidase and hyaluronidase was attributable to a classic phagocytic process, whereas that obtained using phospholipase resulted from active penetration. Neuroradiology, 1993, 35(3), 218 - 20 Diffuse pneumocephalus due to Clostridium septicum cerebritis in haemolytic uraemic syndrome: CT demonstration; Randall JM et al.; The computed tomography finding of diffuse pneumocephalus due to infection by gas-forming organisms is very unusual . We report such a case due to secondary infection by Clostridium septicum in a child with diarrhoea-associated haemolytic uraemic syndrome. J Gen Microbiol, 1993 Jan, 139 ( Pt 1), 79 - 86 The complete nucleotide sequence of the gene encoding the nontoxic component of Clostridium botulinum type E progenitor toxin; Fujii N et al.; We have analysed the genes borne on a 6.0 kb HindIII fragment cloned from the chromosome of Clostridium botulinum type E strain Mashike . This fragment, cloned within plasmid pU9EMH, contains part of the structural gene for botulinum toxin type E neurotoxin as well as the entire structural gene for a nontoxic component of botulinum type E progenitor neurotoxin gene, ent-120 . ent-120 is transcribed in the same direction as the neurotoxin gene and consists of one open reading frame encoding 1162 amino acid residues . Western blotting with anti-nontoxic component sera demonstrates that ent-120 encodes a protein of 120 kDa which forms part of the nontoxic component . ent-120 is homologous to an analogous gene found in botulinum type C strains (69.3% identity at the nucleotide level and 56.1% at the amino acid level) . Two stretches of amino acids at the N-terminus of the ent-120 protein are highly homologous to amino acid sequences within the type E neurotoxin . The stop codon of the ent-120 gene is situated 27 nucleotides upstream from the start codon of the neurotoxin gene. Infection, 1993 Jan-Feb, 21(1), 54 - 6 Clostridium perfringens septicemia with massive hemolysis; Rogstad B et al.; Massive hemolysis and renal failure are rare complications of infection with Clostridium perfringens, resulting in a very high mortality rate (70-100%) . The severity of the infection depends on the presence of underlying conditions such as malignancies and diabetes mellitus . In patients without underlying disorders, massive hemolysis and anuria have been observed in only eight cases, according to recent reports . This case report describes a 61-year-old man who died of C . perfringens septicemia with massive hemolysis and anuria less than 4 h after admittance to the hospital, despite rapid and adequate antibiotic treatment . No focal infection was found. Postgrad Med J, 1993 Jan, 69(807), 45 - 7 The prevalence and nosocomial acquisition of Clostridium difficile in elderly hospitalized patients; Rudensky B et al.; Rectal swabs obtained from 10 of 49 chronic-care geriatric patients were positive for Clostridium difficile, for a prevalence rate of 20.4% . Simultaneous sampling of 29 patients in an acute geriatric ward revealed four colonized patients, for a prevalence rate of 13.7% . A prospective study of C . difficile colonization in 100 consecutive patients admitted to an acute geriatric ward was carried out . All patients were sampled upon admission and biweekly during hospitalization . Two patients (2%) were positive on admission, and 12 of the 98 initially negative patients became colonized, representing a nosocomial acquisition rate of 12.2% . A major determinant for C . difficile colonization in asymptomatic patients appears to be length of hospitalization . Previous antibiotic administration was not found to be a significant factor. Monatsschr Kinderheilkd, 1993 Jan, 141(1), 33 - 5 {Botulism in infancy . A rare cause of floppy infant syndrome}; Greve M et al.; In Europe only a few cases of infantile botulism have been identified . A two month old infant fell ill with feeding difficulties and profound general weakness . Clostridium botulinum and botulinal toxin, the causative agents, were isolated from the patients stool . We describe the typical clinical symptoms and the investigations confirming the diagnosis. Ann Gastroenterol Hepatol (Paris), 1993 Jan-Feb, 29(1), 15 - 8 {Can antibiotic-associated diarrhea be prevented?}; Loizeau E; Post-antibiotic diarrhea is common but rarely serious . The chief cause is changes in the normal intestinal flora . Decrease in the number of bacteria leads to maldigestion of carbohydrates, resulting in osmotic diarrhea . Disappearance of the flora encourages the emergence of resistant strains, e.g . Clostridium difficile . General measures concern the prescription of antibiotics and the use of probiotics . The latter restore and replace the normal flora and prevent more than half of all cases of diarrhea, in particular serious forms and pseudo-membranous colitis . Antibiotics are not accompanied by any complication in 80% of instances . Probiotics should be used in high risk patients: elderly, seriously ill or hospitalised for long periods. Appl Environ Microbiol, 1993 Jan, 59(1), 7 - 14 Precipitation of cadmium by Clostridium thermoaceticum; Cunningham DP et al.; Cadmium at an initial concentration of 1 mM was completely precipitated by cultures of Clostridium thermoaceticum in complex medium . The precipitation was energy dependent and required cysteine, although cysteine alone did not act as a growth substrate . Electron microscopic analysis revealed localized areas of precipitation at the surfaces of nonstarved cells as well as precipitate in the surrounding medium . The addition of cadmium had no apparent effect on growth or acetogenesis . However, nickel and cadmium were synergistically toxic at a concentration (1 mM) at which neither alone was toxic . The amount of protein extracted from cadmium-treated cultures was twofold higher than that in control extracts, and the amount of total sulfide was fourfold higher in cultures containing cadmium than in control cultures . Comparable levels of cysteine desulfhydrase activity were observed in extracts of both cadmium-treated and control cultures, but the enzyme activity was expressed maximally about 24 h earlier in the cadmium-treated cultures than in the untreated controls. Infect Control Hosp Epidemiol, 1993 Jan, 14(1), 36 - 9 Inactivation of Clostridium difficile spores by disinfectants; Rutala WA et al.; OBJECTIVE: The current study was designed to evaluate the activity of glutaraldehyde-based disinfectants against Clostridium difficile using the Association of Official Analytical Chemists' (AOAC) sporicidal test . This study was undertaken because gastrointestinal endoscopes that may be contaminated with C difficile spores are most commonly disinfected between patients using glutaraldehyde-based disinfectants . DESIGN: Using the AOAC test, the following disinfectants were tested: 2% alkaline glutaraldehyde, 2% acid glutaraldehyde, a 1:16 dilution of a 2% glutaraldehyde-7.05% phenol-1.20% sodium phenate, and a 1:20 dilution of a 10% glutaraldehyde-0.5% phenylphenol-0.1% amylphenol . RESULTS: Test results of the four disinfectants against C difficile spores at exposure times of 10, 20, and 60 minutes were as follows (number of positive penicylinders per 30 replicates): 0, 0, and 0 for 2% alkaline glutaraldehyde; 6, 3, and 0 for 2% acid glutaraldehyde; 30, 29, and 30 for a 1:16 dilution of glutaraldehyde-7.05% phenol-1.20% sodium phenate; and 30, 30, and 30 for a 1:20 dilution of glutaraldehyde-0.5% phenylphenol-0.1% amylphenol . CONCLUSIONS: C difficile spores are more susceptible to inactivation by glutaraldehyde-based disinfectants than the spore-forming organisms recommended in the AOAC sporicidal test (i.e., Bacillus subtilis and Clostridium sporogenes) . Diluting glutaraldehyde-based disinfectants below 2% led to the inability to inactivate spores of C difficile using exposure times commonly employed to disinfect semicritical items such as gastrointestinal endoscopes. Int J Syst Bacteriol, 1993 Jan, 43(1), 107 - 10 Phylogeny of the ammonia-producing ruminal bacteria Peptostreptococcus anaerobius, Clostridium sticklandii, and Clostridium aminophilum sp . nov; Paster BJ et al.; In previous studies, gram-positive bacteria which grew rapidly with peptides or an amino acid as the sole energy source were isolated from bovine rumina . Three isolates, strains C, FT (T = type strain), and SR, were considered to be ecologically important since they produced up to 20-fold more ammonia than other ammonia-producing ruminal bacteria . On the basis of phenotypic criteria, the taxonomic position of these new isolates was uncertain . In this study, the 16S rRNA sequences of these isolates and related bacteria were determined to establish the phylogenetic positions of the organisms . The sequences of strains C, FT, and SR and reference strains of Peptostreptococcus anaerobius, Clostridium sticklandii, Clostridium coccoides, Clostridium aminovalericum, Acetomaculum ruminis, Clostridium leptum, Clostridium lituseburense, Clostridium acidiurici, and Clostridium barkeri were determined by using a modified Sanger dideoxy chain termination method . Strain C, a large coccus purported to belong to the genus Peptostreptococcus, was closely related to P . anaerobius, with a level of sequence similarity of 99.6% . Strain SR, a heat-resistant, short, rod-shaped organism, was closely related to C . sticklandii, with a level of sequence similarity of 99.9% . However, strain FT, a heat-resistant, pleomorphic, rod-shaped organism, was only distantly related to some clostridial species and P . anaerobius . On the basis of the sequence data, it was clear that strain FT warranted designation as a separate species . The closest known relative of strain FT was C . coccoides (level of similarity, only 90.6%) . Additional strains that are phenotypically similar to strain FT were isolated in this study.(ABSTRACT TRUNCATED AT 250 WORDS) Med Pediatr Oncol, 1993, 21(1), 70 - 2 Clostridium cadaveris bacteremia in the immunocompromised host; Gucalp R et al.; Clostridium cadaveris, usually considered a non-pathogen, was isolated from blood cultures of two febrile patients with cancer . The bacteremias appeared to have originated from the abdomen . This organism has not been previously reported as the etiological agent in this setting. Arch Biochem Biophys, 1993 Jan, 300(1), 483 - 8 Purification and characterization of Clostridium difficile glutamate dehydrogenase; Anderson BM et al.; Recombinant Clostridium difficile glutamate dehydrogenase (L-glutamate:NAD oxidoreductase, EC 1.4.1.2) was purified 177-fold to electrophoretic homogeneity with a 62% recovery through a four-step procedure involving gel filtration and ion-exchange and dye affinity chromatography . The approximate molecular weights of the native enzyme by gel filtration and subunits by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were consistent with a hexameric structure for the purified enzyme . The enzyme-catalyzed glutamate oxidation was an NAD-dependent sequential process in which NADP could not be substituted as coenzyme . Several dinucleotide analogs of NAD structurally altered in either the pyridine or the purine moiety were observed to function as coenzymes when substituted for NAD . Nicotinamide mononucleotide did not serve as a coenzyme for glutamate oxidation . Product inhibition by NADH was competitive with respect to NAD . In deadend inhibition studies, adenosine diphosphoribose was shown to be an effective coenzyme-competitive inhibitor. J Clin Gastroenterol, 1993 Jan, 16(1), 26 - 30 Role of infectious agents in exacerbations of ulcerative colitis in India . A study of Clostridium difficile; Kochhar R et al.; Fifty patients with idiopathic ulcerative colitis--25 with acute exacerbation of the disease (Group I) and 25 in quiescent phase (Group II)--were studied . None of the patients had a history of recent exposure to antimicrobial drugs or hospitalization . Evidence of infection with protozoal and bacterial agents and/or presence of Clostridium difficile toxin was demonstrated in eight (32%) patients in group I and one (4%) patient in group II (group I vs . group II, p < 0.05; chi 2 test with Yate's correction) . In five of the six patients with demonstrable Clostridium difficile toxin in the stool, one patient with Entamoeba trophozoites, and another with Salmonella infection, the exacerbation responded clinically and endoscopically to specific antimicrobial therapy . Another two patients who had Entamoeba histolytica cysts in the stool had no change in their clinical status with metronidazole . We conclude that infectious agents are responsible for some of the exacerbations in patients of ulcerative colitis in a tropical country like India, where careful microbiologic examination is in order in every acute exacerbation of this disease . This contrasts with the findings in developed countries. J Appl Bacteriol, 1993 Jan, 74(1), 67 - 77 Purification, partial characterization and plasmid-linkage of pediocin SJ-1, a bacteriocin produced by Pediococcus acidilactici; Schved F et al.; Pediococcus acidilactici SJ-1, isolated from a naturally-fermented meat product, produced an antibacterial agent active against selected strains of Lactobacillus spp., Clostridium perfringens and Listeria monocytogenes . The agent was bactericidal against sensitive indicators, and sensitive to proteolytic enzymes; it was identified as a bacteriocin, and was designated as pediocin SJ-1 . It was stable over a wide pH range (3-9), and apparently most stable in the lower part of that range . At pH 3.6, pediocin SJ-1 was stable at heat-processing temperatures within the range 65-121 degrees C; its activity decreased significantly, however, when it was heated at pH 7.0 . The activity of pediocin SJ-1 on sensitive indicator cells was lost in the presence of alpha-amylase, suggesting that it contains a glyco moiety, necessary for its antibacterial action . Native pediocin SJ-1 exists in the form of monomers and aggregates (with molecular weights in the range 80-150 kDa) . Pediocin SJ-1 was purified 262-fold by direct application of cell-free supernatant fluids to a cation-exchange chromatography column, and was resolved by SDS-PAGE as a single peptide band with a MW of ca 4 kDa . The original pediocin SJ-1-producing strain (bac+) harbours three plasmids of 4.6, 23.5, and 45.7 MDa . Production of pediocin SJ-1, but not immunity to SJ-1, is associated with the 4.6 MDa plasmid. J Appl Bacteriol, 1993 Jan, 74(1), 61 - 6 Detection by in vitro amplification of the alpha-toxin (phospholipase C) gene from Clostridium perfringens; Fach P et al.; A polymerase chain reaction (PCR) with thermostable DNA polymerase from Thermus aquaticus is described for the specific amplification of the phospholipase C (alpha-toxin) gene of Clostridium perfringens . A set of primers selected for their high specificity could detect Cl . perfringens in stools with a detection limit of approximately 5 x 10(2) bacteria, after bi-amplification . A modified PCR without thermal steps was performed to rapidly amplify, with a yield of 60%, the DNA template . With this PCR method Cl . perfringens alpha-toxin gene could be detected within 2 h . The PCR method detected alpha-toxin positive Cl . perfringens but did not react with phospholipase C-producing Bacillus cereus, Pseudomonas aeruginosa, Cl . sordellii and Cl . bifermentans . The amplified PCR products were screened through ethidium bromide agarose gel electrophoresis or, in only 1 h, with the PhastSystem (Pharmacia) . This PCR satisfies the criteria of specificity, sensitivity and rapidity required for a useful tool in epidemiology and for the diagnosis of the pathogen Cl . perfringens as it may be used directly on stool samples. Am J Med, 1993 Jan, 94(1), 21 - 8 Anaerobic osteomyelitis and arthritis in a military hospital: a 10-year experience; Brook I et al.; PURPOSE: The methods of collecting, transporting, cultivating, and identifying aerobic bacteria in bone and joint infections have improved markedly since the early 1980s . In addition, many of the anaerobes have been reclassified and renamed . The purpose of this study was to provide more current information regarding the incidence of recovery of anaerobic bacteria from clinical specimens of infected bone and joint . MATERIALS AND METHODS: Specimens from 73 infected bone specimens and 65 infected joints inoculated on media supportive for aerobic and anaerobic bacteria showed bacterial growth . RESULTS: One hundred fifty-seven organisms (2.2 isolates/specimen), consisting of 122 anaerobic bacteria (1.7 isolates/specimen) and 35 facultative or aerobic bacteria (0.5 isolate/specimen), were recovered from the 73 bone specimens . Anaerobic bacteria were recovered with aerobe or facultative bacteria in 24 (33%) instances . The predominant anaerobes were Bacteroides species (49 isolates), anaerobic cocci (45), Fusobacterium species (11), Propionibacterium acnes (7), and Clostridium species (6) . Conditions predisposing to bone infections were vascular disease, bites, contiguous infection, peripheral neuropathy, hematogenous spread, and trauma . Pigmented Prevotella and Porphyromonas species were mostly isolated in skull and bite infections (7 of 19), members of the Bacteroides fragilis group in hand and feet infection (12 of 16), and Fusobacterium species in skull, bite, and hematogenous long bone infections . Seventy-four organisms (1.1 isolates/specimen), consisting of 67 anaerobic bacteria (1.0 isolate/specimen) and 7 facultative or aerobic bacteria (0.1 isolate/specimen), were isolated from 65 joint specimens . The predominant anaerobes were P . acnes (24 isolates), anaerobic cocci (17), Bacteroides species (10), and Clostridium species (5) . Predisposing conditions to joint infection were trauma, prior surgery, presence of a prosthetic joint, and contiguous infection . P . acnes isolates were associated with prosthetic joints, members of the B . fragilis group with hematogenous spread, and Clostridium species with trauma . The clinical presentation of these cases is discussed . CONCLUSION: These data highlight the importance of anaerobic bacteria in bone and joint infection. J Med Microbiol, 1993 Jan, 38(1), 69 - 73 Purification and characterisation of intracellular toxin A of Clostridium difficile; Meng XQ et al.; After sonic disintegration of Clostridium difficile cells, intracellular toxin A was purified to homogeneity by thyroglobulin affinity chromatography (TGAC) followed by anion-exchange (Mono Q) by fast protein liquid chromatography (FPLC) . High haemagglutinating (HA) activity was detected in TGAC-unbound fractions (2(9)/50 microliters), but not in TGAC thermal eluates (2(0)/50 microliters) . The low HA titre of the thermal eluates was markedly increased to 2(5)/50 microliters after dialysis against 0.02 M Tris-HCl (pH 7.5) . A disparity in the position of the peaks containing cytotoxic and HA activity was observed in the first Mono Q-FPLC step . Intracellular toxin A without HA activity was obtained by a second Mono Q-FPLC step . The M(r) of the intracellular toxin A was estimated by polyacrylamide gel electrophoresis (PAGE) to be 580 kDa under non-denaturing conditions . The minimum doses of the toxin causing cytotoxicity, mouse lethality and enterotoxicity were 0.83 ng, 8.7 ng and 5 micrograms, respectively. Cornell Vet, 1993 Jan, 83(1), 77 - 81 Abdominal pain associated with an umbilical abscess in a llama; St-Jean G et al.; A 3-month-old llama with a presenting complaint of lethargy, anorexia, and a painful, distended abdomen was evaluated . The llama had intermittently strained to defecate during the 3 days prior to admission . Physical examination results, hematologic data and lateral abdominal radiographs were used to diagnose a large umbilical abscess, which was causing a partial obstruction of the gastrointestinal tract . Under general anesthesia, 3 liters of purulent exudate were drained from the abscess . The abscess cavity was then lavaged with saline solution and its capsule was marsupialized to the skin . Cultures of the abscess content yielded Proteus sp, Streptococcus equisimilis, and Clostridium septicum . Two days after surgery, the llama was drinking, eating, and passing feces . The abscess was lavaged daily for a total of 11 days . Six months after surgery, the llama was the same size as other llamas of the same age, and the owners were pleased with the cosmetic appearance of the ventral abdomen . Umbilical abscesses can vary in size and clinical presentation; they should be recognized as a possible cause of abdominal pain with a potential for causing intestinal obstruction in llamas. Mol Cell Biol, 1993 Jan, 13(1), 72 - 9 Involvement of rho p21 and its inhibitory GDP/GTP exchange protein (rho GDI) in cell motility; Takaishi K et al.; Evidence is accumulating that rho p21, a ras p21-related small GTP-binding protein (G protein), regulates the actomyosin system . The actomyosin system is known to be essential for cell motility . In the present study, we examined the action of rho p21, its inhibitory GDP/GTP exchange protein (named rho GDI), its stimulatory GDP/GTP exchange protein (named smg GDS), and Clostridium botulinum ADP-ribosyltransferase C3, known to selectively ADP-ribosylate rho p21 and to impair its function, in cell motility (chemokinesis) of Swiss 3T3 cells . We quantitated the capacity of cell motility by measuring cell tracks by phagokinesis . Microinjection of the GTP gamma S-bound active form of rhoA p21 or smg GDS into Swiss 3T3 cells did not affect cell motility, but microinjection of rho GDI into the cells did inhibit cell motility . This rho GDI action was prevented by comicroinjection of rho GDI with the GTP gamma S-bound form of rhoA p21 but not with the same form of rhoA p21 lacking the C-terminal three amino acids which was not posttranslationally modified with lipids . The rho GDI action was not prevented by Ki-rasVal-12 p21 or any of the GTP gamma S-bound form of other small GTP-binding proteins including rac1 p21, G25K, and smg p21B . Among these small G proteins, rhoA p21, rac1 p21, and G25K are known to be substrates for rho GDI . The rho GDI action was not prevented by comicroinjection of rho GDI with smg GDS . Microinjection of C3 into Swiss 3T3 cells also inhibited cell motility . These results indicate that the rho GDI-rho p21 system regulates cell motility, presumably through the actomyosin system. J Urol, 1993 Jan, 149(1), 142 - 4 Postoperative clostridium difficile gastroenteritis; Godet AS et al.; Clostridium difficile gastroenteritis can be the cause of an enigmatic postoperative syndrome of high temperature and marked leukocytosis, out of proportion to the initially mild constitutional symptoms . Patients may suffer delayed onset of diarrhea, which will test positive for the C . difficile enterotoxin by latex agglutination . We report 5 cases of C . difficile gastroenteritis that occurred within a 2-year period . We believe that the combination of preoperative bowel preparation, and intraoperative and postoperative systemic antibiotics is the primary operant factor . All patients responded rapidly when oral antibiotics specific for C . difficile were instituted . The sequelae of C . difficile colitis can include toxic megacolon with perforation and peritonitis, increasing the importance of early recognition and appropriate treatment. J Clin Microbiol, 1993 Jan, 31(1), 157 - 9 Use of electrophoretic polymorphisms of esterases for differentiation of Clostridium argentinense strains; Bories PN et al.; Esterase electrophoresis was used to study 10 strains of Clostridium argentinense, including 7 toxigenic and 3 nontoxigenic strains . On the basis of the electrophoretic mobilities and hydrolytic specificities toward five synthetic substrates, different esterase profiles could be defined for almost all strains, revealing the heterogeneity of bacterial clones . Therefore, electrophoretic polymorphism of esterases can be used for differentiation of C . argentinense in population genetic or epidemiological studies. Can J Infect Control, 1993 Summer, 8(2), 37 - 40 Clostridium difficile enteritis in a Canadian tertiary care hospital; MacDonald KS et al.; Nosocomial Clostridium difficile enteritis causes epidemic and endemic infection . The authors report the incidence of C difficile enteritis in a tertiary care hospital . The overall incidence rate over two years in the authors' institution was 22.2 cases per 100,000 in-patient-days; this rate appeared to decline over the study period (from 30.0 cases per 100,000 in 1990 to 11.2 at study end in 1992) . Continued surveillance will determine whether this is a durable trend . An average of 5.25 cases per month occurs in the authors' hospital . A 50% increase overall was selected as the threshold for outbreak investigation . Surveillance of ward and specialty specific rates will allow clustering detection. Biol Chem Hoppe Seyler, 1993 Jan, 374(1), 85 - 90 On the role of two different cobalt(II) species in coenzyme B12-dependent 2-methyleneglutarate mutase from Clostridium barkeri; Zelder O et al.; Purified 2-methyleneglutarate mutase from Clostridium barkeri contains adenosylcobalamin (coenzyme B12) and varying amounts of oxygen-stable cob(II)alamin . The content of the latter was estimated by EPR spectroscopy at 6-11% of the total cobalamin (2-4 mol/mol enzyme) . Tryptic digestion of the enzyme liberated the prosthetic groups, cob(II)alamin being oxidized by air to aquocobalamin . HPLC analysis of the released cobamides from several preparations revealed > 90% adenosylcobalamin and < 10% aquocobalamin . Treatment of active 2-methyleneglutarate mutase with 8M urea followed by gelfiltration yielded an inactive enzyme from which 50% of the adenosylcobalamin and up to 70% of the cob(II)alamin was removed . Addition of adenosylcobalamin to the urea-treated enzyme resulted in complete reactivation, but the content of cob(II)alamin was not increased . These data suggest that the oxygen-stable cob(II)alamin is not involved in catalysis . In the presence of the competitive inhibitor itaconate (methylenesuccinate, Ki = 0.7mM), an alteration of the UV/visible spectrum at 470 nm as well as a new line in the EPR spectrum of the enzyme (around g = 2.1) was observed . The results indicate the formation of an unusual, oxygen sensitive Co(II) species during catalysis . The EPR signal of the oxygen-stable cob(II)alamin (gx,y = 2.24) remained unchanged under those conditions. Langenbecks Arch Chir, 1993, 378(4), 249 - 54 {Pneumatosis cystoides intestinii}; Bertram P et al.; Pneumatosis cystoides intestinalis (PCI), a condition involving submucosal or subserosal gas-containing cysts of the wall of the gastrointestinal tract, is a rare entity . It is mostly diagnosed between the third and fifth decades of life without a clear sexual predominance . Different aetiopathogenetic factors are under discussion, the most probable being a bacteriologic cause (Clostridium perfringens) in combination with minimal leaks in the mucosal barrier . There are no pathognomonic symptoms; the clinical picture ranges from incidental findings to haematochezia . Diagnosis is based on plain abdominal film and X-ray following barium enema . Methods of treatment in symptomatic cases are oxygen and antibiotic (metronidazole) therapies and, in severe cases, resection of the diseased part of the intestine. Folia Microbiol (Praha), 1993, 38(3), 188 - 92 Effect of alkyldimethylamine oxides on anaerobic sporulating bacteria of genus Clostridium; Ciganekova V et al.; Antibacterial activity of alkyldimethylamine oxides and Iodaminox was determined in vitro in 11 strains of clostridia . The most efficient was (1-methyldodecyl)dimethylamine oxide (MIC = 7.8-78 mumol/L) . Iodaminox was about twice as efficient (MIC = 3.9-31 mumol/L) . The bactericidal activity of both compounds was tested at different periods of exposure, ambient pH and cultivation temperature on three species of clostridia . The activity of both compounds was the highest at pH 6 and 40 degrees C. J Gynecol Obstet Biol Reprod (Paris), 1993, 22(4), 425 - 8 {Infectious endocarditis of gyneco-obstetric origin . Apropos of 15 cases}; Nkoua JL et al.; With the object of analyzing current characteristics of post-partum and post-abortum infective endocarditis (IE), authors carried out a retrospective study of 15 cases between september 1985 and may 1992 . Mean age was 22.2 +/- 4.0 years . Origin of sepsis was delivery (1 case), abortion (14 cases) . There was no underlying cardiac lesion in 9 cases, rheumatic heart disease in 6 cases . Infecting organisms were Staphylococcus aureus (n = 6), streptococcus D (n = 3), Clostridium perfringens (n = 2) . There were 10 acute and 5 subacute IE, 7 right-sided, 7 left-sided, and 1 right and left-sided IE . Vegetations were determined by transthoracic echocardiography in 12 cases (80%) . The main complications were heart failure (15 cases), and pulmonary or arterial embolism (7 cases) . Lethality was 53.3% and was not different in tricuspid acute IE and left-sided IE . Treatment was the more difficult as cardiac surgery is expensive or even inaccessible . Accordingly, prevention is primordial . It consist of antibiotic prophylaxis and fight against illicit abortion. Microbiol Immunol, 1993, 37(5), 395 - 8 Similarity in nucleotide sequence of the gene encoding nontoxic component of botulinum toxin produced by toxigenic Clostridium butyricum strain BL6340 and Clostridium botulinum type E strain Mashike; Fujii N et al.; The complete nucleotide and deduced amino acid sequence of the nontoxic component of botulinum type E progenitor toxin is determined in recombinant plasmid pU9BUH containing about 6.0 kb HindIII fragment obtained from chromosomal DNA of Clostridium butyricum strain BL6340 . The open reading frame (ORF) of this nontoxic component gene is composed of 3,486 nucleotide bases (1,162 amino acid residues) . The molecular weight calculated from deduced amino acid residues is estimated 13,6810.1 . The present study revealed that 33 nucleotide bases of 3,486 are different in the nontoxic component gene between C . butyricum strain BL6340 and C . botulinum type E strain Mashike . This corresponds to the difference of 17 amino acid residues in these nontoxic component. Gastroenterol Clin Biol, 1993, 17(4), 283 - 6 Detection of Clostridium difficile toxins in stools . Comparison between a new enzyme immunoassay for toxin A and other routine tests; Depitre C et al.; An enzyme-linked immunosorbent assay (ELISA) for detection of Clostridium difficile toxin A (enterotoxin) using a monoclonal antibody is described . No cross-reaction was observed with any of the Clostridium species tested except for toxigenic Clostridium difficile . One hundred and eight stool specimens from hospitalized patients harbouring C . difficile in their intestine and 43 samples negative for C . difficile isolation were studied to compare this test with a cytotoxicity assay, the isolation of toxigenic C . difficile and the commercial EIA Premier test (Meridian) . As compared with the cytotoxicity assay and EIA, specificity was 91 and 100%, while sensitivity 83 and 74% respectively . This ELISA technique could be used as a routine test for toxin A detection in stools. Appl Biochem Biotechnol, 1993 Spring, 39-40, 149 - 58 Structural features of the Clostridium thermocellum cellulase SS gene; Wang WK et al.; The Clostridium thermocellum cellulase SS is a subunit of the extracellular cellulase complex (cellulosome) . It has previously been shown that SS, hydrolyzes crystalline cellulose synergistically with another subunit, SL . To study this synergism further, the authors cloned the gene coding for SS (celS) and compared its sequence to other known cel genes . The celS, although unique in its DNA sequence, has many structural features similar to those found in other cel genes . These features include a ribosome biding site, signal peptide sequence, the existence of a conserved reiterated amino acid sequence, and a palindromic structure downstream from its open reading frame. Schweiz Arch Tierheilkd, 1993, 135(11-12), 328 - 32 {Hepatic necrosis caused by Clostridium perfringens in a dog}; Bornand-Jaunin V et al.; A case of acute hepatic necrosis in a dog is clinically and pathologically described . The occurrence of Clostridium perfringens in the liver lesions was revealed by histological examination and bacteriological isolation . Based on these findings we conclude that Clostridium perfringens is the cause of the infarcts . The case is discussed on the base of the literature. Compr Ther, 1993, 19(4), 151 - 6 Epidemiology of nosocomial Clostridium difficile infection; Samore MH; C . difficile is a frequent cause of nosocomial diarrhea and is associated with substantial morbidity . C . difficile carriage may arise either from overgrowth of endogenous organisms or from exogenous acquisition . Within hospitals, asymptomatically colonized patients typically outnumber symptomatic patients by a ratio of approximately 3:1 . Patient-to-patient transmission of C . difficile has been well documented in studies of nosocomial outbreaks, utilizing typing methods to supplement epidemiologic investigations . The mechanisms and frequency of transmission probably vary between institutions, but contamination of environmental surfaces and personnel hand carriage both appear to be important . Nosocomial C . difficile infection can be remarkably difficult to control, particularly after it becomes endemic in an institution . Policies targeting both environmental contamination and personnel hand-washing/glove use practices may be most successful . Prophylaxis of C . difficile diarrhea in high-risk patients on antibiotics may also have merit if an effective and safe biologic agent can be identified . Asymptomatically colonized patients should not be treated because they are at low risk for the development of diarrhea and because further antibiotic treatment may prolong the carrier state. Vet Med (Praha), 1993, 38(10), 589 - 96 {The role anaerobes in dermatitis digitalis et interdigitalis in cattle}; Koniarova I et al.; Altogether 52 tissue samples and swabs from the corium of the claw of diseased cattle were investigated . Microbiological examinations revealed the presence of 11 strictly anaerobic bacterial strains that belonged to the species Peptococcus asacharolyticus, Pc . sacharolyticus, Peptostreptococcus anaerobius, Bacteroides asacharolyticus and B . melaninogenicus . The remaining isolates were classified as Bacteroides, Fusobacterium, Streptococcus and Clostridium . In the above-mentioned strains observations of their biochemical, morphological and enzymatic properties (Tab . I) were carried out . Their resistance to antibiotics was evaluated from the qualitative viewpoint (Tab . II) . Capillary isotachophoresis was used to determine the amounts of volatile fatty acids that developed during glucose fermentation (Tab . III) . In the single clinical samples the above-mentioned strains occurred as association of microorganism . The anaerobic genera Bacteroides and Fusobacterium were found in 52% of the samples (Bacteroides sp . and Fusobacterium sp . were recorded in 25% and 12% of the samples, respectively, whereas in 15% of the samples both species occurred) . According to our results the etiology of dermatitis digitalis et interdigitalis is closely connected with the incidence of anaerobes . Of the latter, those belonging to the genera Bacteroides and Fusobacterium are of major importance. Surg Gynecol Obstet, 1993, 177 Suppl, 55 - 64; discussion 65-70 Emerging problems in gram-positive infections in the postoperative patient; Barie PS; Gram-positive bacteria are increasingly prevalent in the postoperative patient population and are important as pathogens . As increasingly ill and elderly patients undergo surgical treatment and as increased use is made of invasive or immunosuppressive treatment modalities, this trend is likely to accelerate . The increasing use of broad-spectrum antibiotics results in the emergence of resistant pathogens or superinfections that are increasingly difficult to treat . Examples of such pathogens include methicillin-resistant Staphylococcus epidermidis, vancomycin-resistant enterococci and Clostridium difficile . No acute care setting, from large urban teaching hospital to small rural community hospital, is immune to the trend . The clinical challenge posed by these organisms is compounded by the fact that serious gram-positive infections can be impossible to distinguish on clinical grounds from their gram-negative counterparts . The host inflammatory response to gram-positive infection is quite similar to that caused by gram-negative infection, including elaboration of cytokine mediators and progression to visceral organ dysfunction . The clinician must be cognizant that serious infections, especially of nosocomial origin, may be caused by gram-positive bacteria. Nord Med, 1993, 108(11), 283 - 5 {Antibiotic-induced diarrhea and pseudomembranous colitis}; Schreiner A; Although not all antibiotic-induced diarrhoea is pseudomembranous, and not all pseudomembranous colitis is antibiotic-induced, by far the greater proportion of intestinal lesions and diarrhoea occurring in conjunction with Clostridium difficile infection are attributed to previous antibiotic treatment . The article consists in a review of pathogenic, diagnostic, treatment and nosocomial aspects. Microbiol Immunol, 1993, 37(6), 447 - 50 Effect of Clostridium perfringens epsilon toxin on rat isolated aorta; Nagahama M et al.; The effect of Clostridium perfringens epsilon toxin on rat isolated aorta was investigated . The toxin caused contraction of the isolated aorta in a dose-dependent manner . The toxin induced no contraction of the isolated aorta in low-Na medium and of the tissue stored at 4 C for 7 days . However, tetrodotoxin (TTX) had no effect on the toxin-induced contraction . The toxin-induced contraction was significantly inhibited by phentolamine and prazosin, but did not by atropine, mecamylamine, chlorpheniramine and methysergide . These data suggest that the toxin-caused contraction is mediated through nervous system in rat isolated aorta. Med Dosw Mikrobiol, 1993, 45(4), 483 - 6 {Susceptibility of Clostridium difficile strains to teicoplanin and ramoplanin}; Dworczynski A et al.; The in vitro activity of teicoplanin and ramoplanin against 113 Clostridium difficile strains isolated from different origins was determined . The minimum inhibitory concentrations (MICs) of antibiotics were determined by the agar dilution method . MIC of teicoplanin and ramoplanin were twofold lower for Clostridium difficile strains isolated from patient with pseudomembranous colitis . No correlation between the origin of the strains toxigenicity and susceptibility to teicoplanin and ramoplanin was observed . Our result indicate that teicoplanin and ramoplanin are potentially effective alternative antibiotics for treatment of infections caused by Clostridium difficile. DNA Seq, 1993, 4(2), 105 - 11 The nucleotide sequence of genes involved in the leucine biosynthetic pathway of Clostridium pasteurianum; Oultram JD et al.; A 2.2 kb SphI/ClaI fragment of the Clostridium pasteurianum chromosome has previously been cloned and shown to complement leuB401 and leuC171 mutations in Escherichia coli . The nucleotide sequence of this fragment has been determined (2327 bp) and carries three open reading frames . The products of translation of these reading frames display significant homologies with the alpha-isopropylmalate isomerase subunit (leuD) gene of Salmonella typhimurium, the beta-isopropylmalate dehydrogenase (leuB) genes of several organisms, and the dihydroxyacid dehydrase (ilvD) gene of E . coli. Nat Toxins, 1993, 1(6), 361 - 8 Effect of okadaic acid on the cytotoxic activity of Clostridium difficile toxin B and Clostridium sordellii toxin L; Baldacini O et al.; Clostridium difficile toxin B and Clostridium sordellii toxin L, which are immunologically related toxins, possess a cytotoxic activity inducing depolymerization of microfilaments and cellular retraction of cell bodies that are different for toxin B- and toxin-L-treated cells . The biological mechanisms responsible for these effects are unknown, but a previous study revealed that both toxins induce modification of phosphorylation of cellular proteins extracted from toxin B- and toxin L-treated cells without changes in protein kinase C activity or cAMP concentration . In the present study, we have investigated the effect of okadaic acid, an inhibitor of protein phosphatases, on the cytotoxic activity of toxins B and L in MacCoy cells . Firstly, we reveal by cytotoxic assay and staining of F-actin that okadaic acid (1 microM or higher) induces depolymerization of microfilaments and cellular morphological modifications which are similar to that of cells treated with toxin L . Secondly, we show that 1 microM okadaic acid potentials the cytotoxic activity of toxin L but not of toxin B . These observations suggest that the cytotoxic mechanisms induced by okadaic acid and toxin treatment are partly common, indicating that an increase in phosphorylation favors the cytotoxicity of toxin L . Since okadaic acid had no influence on the cytotoxicity of toxin B, we suggest that toxin B and L, alter the cells by different cellular biological mechanisms. Acta Orthop Belg, 1993, 59(4), 416 - 9 Clostridium septicum gangrene complicating a closed femoral fracture; Mulier T et al.; One of the basic principles of orthopedic surgery is that gas gangrene does not develop in closed fractures . That there are exceptions to this rule is demonstrated by the following report of an extremely rare case . Clostridium septicum myonecrosis developed after surgical treatment of a closed femoral fracture in a fit 45-year-old man, treated by antibiotics . The patient survived after having a right hip disarticulation and administration of high doses of intravenous penicillin . Prophylactically administered cephalosporins failed to prevent the Clostridial infection. Antonie Van Leeuwenhoek, 1993-94, 64(3-4), 273 - 83 Genetic interrelationships of proteolytic Clostridium botulinum types A, B, and F and other members of the Clostridium botulinum complex as revealed by small-subunit rRNA gene sequences; Hutson RA et al.; The phylogenetic interrelationships of members of the Clostridium botulinum complex of species was investigated by direct sequencing of their 16S rRNA genes . Comparative analysis of the 16S rRNA sequences demonstrated the presence of four phylogenetically distinct lineages corresponding to: i) proteolytic C . botulinum types A, B, and F, and C . sporogenes, ii) saccharolytic types B, E and F, iii) types C and D and C . novyi type A, and iv) type G and C . subterminale . The phylogenetic groupings obtained from the 16S rRNA were in complete agreement with the four divisions recognised within the the 'species complex' on the basis of phenotypic criteria. Nat Toxins, 1993, 1(6), 369 - 75 Effects of purified Clostridium difficile toxin A in the small intestine of the rat in vivo; Beubler E et al.; The action of highly purified Clostridium difficile toxin A was studied in the jejunum of rats in vivo . C . difficile toxin A reversed dose-dependently net fluid absorption into net fluid secretion, accompanied by an increase in prostaglandin E2 but not 5-hydroxytryptamine output into the gut lumen . Accordingly, indomethacin but not the 5-hydroxytryptamine receptor antagonists ketanserin plus tropisetron were able to inhibit toxin A-induced fluid secretion . Atropine and hexamethonium were without effect on the action of toxin A, such excluding a nervous mechanism . The cyclic nucleotides cyclic AMP and cyclic GMP appear not to be involved in the mediation of the secretory response . The reduced cyclic GMP levels are most likely the result of a complete destruction of the villus membranes, where the guanylate cyclase is located . Histological studies revealed massive damage to intestinal villi, whereas the majority of the crypts seem to be unaffected . In conclusion, toxin A-induced intestinal fluid secretion appears to be caused mainly by severe mucosal damage . PGE2-release may be the consequence of the inflammation accompanying this damage . The mechanism seems to be completely different to those of cholera toxin or Escherichia coli heat stable enterotoxin. J Gen Microbiol, 1993 Jan, 139 ( Pt 1), 59 - 65 A reporter gene vector to investigate the regulation of glutamine synthetase in Bacteroides fragilis Bf1; Abratt VR et al.; The Clostridium acetobutylicum eglA gene, encoding a beta-1,4-endoglucanase (EG), was shown to be a useful reporter gene for the study of gene expression in Bacteroides fragilis . The eglA reporter gene has the advantages that it can be easily identified in both Escherichia coli and B . fragilis on agar media containing carboxymethylcellulose, and EG production can be rapidly quantified in liquid medium . Since the B . fragilis glutamine synthetase (GS) is inactivated in permeabilized cells and cell extracts, the eglA reporter gene was used to study the regulation of GS production in B . fragilis . Gene fusions containing the GS glnA promoter region fused to the promoterless eglA gene showed that glnA expression was regulated by nitrogen in B . fragilis at the transcriptional level . A glnA upstream region containing a near-perfect direct repeat sequence was essential for efficient GS expression and for regulation by nitrogen. Acta Microbiol Pol, 1993, 42(3-4), 251 - 7 Comparison of Delmée and Polish serogroup-specific Clostridium difficile strains; Martirosian G et al.; In this study we compared Delmee and Polish serogroup-specific Clostridium difficile strains by slide agglutination, observation of flagella and SDS-PAGE protein patterns . Among Delmee serogroup specific strains we observed flagella in groups A, D and K . Among Polish serogroup-specific Clostridium difficile strains we did not observe flagella . The Polish serogroup-specific strains gave different protein patterns compared to Delmee strains . SDS-PAGE protein pattern analyses of Polish serogroup-specific strains divide these strains into 3 groups: 71, 88 and third embracing 18, 27, 70, 72, and 89. Biochemistry, 1992 Dec 29, 31(51), 12870 - 5 Function and CO binding properties of the NiFe complex in carbon monoxide dehydrogenase from Clostridium thermoaceticum; Shin W et al.; Adding 1,10-phenanthroline to carbon monoxide dehydrogenase from Clostridium thermoaceticum results in the complete loss of the NiFeC EPR signal and the CO/acetyl-CoA exchange activity . Other EPR signals characteristic of the enzyme (the gav = 1.94 and gav = 1.86 signals) and the CO oxidation activity are completely unaffected by the 1,10-phenanthroline treatment . This indicates that there are two catalytic sites on the enzyme; the NiFe complex is required for catalyzing the exchange and acetyl-CoA synthase reactions, while some other site is responsible for CO oxidation . The strength of CO binding to the NiFe complex was examined by titrating dithionite-reduced enzyme with CO . During the titration, the NiFeC EPR signal developed to a final spin intensity of 0.23 spin/alpha beta . The resulting CO titration curve (NiFeC spins/alpha beta vs CO pha beta) was fitted using two reactions: binding of CO to the oxidized NiFe complex, and reduction of the CO-bound species to a form that exhibits the NiFeC signal . Best fits yielded apparent binding constants between 6000 and 14,000 M-1 (Kd = 70-165 microM) . This sizable range is due to uncertainty whether CO binds to all or only a small fraction (approximately 23%) of the NiFe complexes . Reduction of the CO-bound NiFe complex is apparently required to activate it for catalysis . The electron used for this reduction originates from the CO oxidation site, suggesting that delivery of a low-potential electron to the CO-bound NiFe complex is the physiological function of the CO oxidation reaction catalyzed by this enzyme. Biochemistry, 1992 Dec 29, 31(51), 12863 - 9 Copurification of rho protein and the rho-GDP dissociation inhibitor from bovine neutrophil cytosol . Effect of phosphoinositides on rho ADP-ribosylation by the C3 exoenzyme of Clostridium botulinum; Bourmeyster N et al.; The substrate of the C3 exoenzyme from botulinum toxin is a protein which is particularly abundant in the cytosol of neutrophils {Stasia, M . J., Jouan, A., Bourmeyster, N., Boquet, P., & Vignais, P . V . (1991) Biochem . Biophys . Res . Commun . 180, 615-622} . Optimal conditions for the ADP-ribosylation of the C3 substrate have been established in order to follow the course of its purification from bovine neutrophil cytosol . In particular, phosphoinositides at micromolar concentrations were found to enhance the ADP-ribosylation capacity of the C3 substrate in crude neutrophil cytosol and partially purified fractions . A {32P}ADP-ribosylatable protein, migrating on SDS-PAGE with a mass of 24 kDa, was copurified with a 29-kDa protein by a series of chromatographic steps on DEAE-Sephacel, Biogel P60, and Mono Q . In the case of the C3 substrate, isoelectric focusing revealed two major labeled bands with pI values of 6.2 and 5.6; the pI of the 29-kDa protein was 4.8-5.0 . On the basis of the amino acid sequence of peptides resolved after proteolytic digestion, the 24-kDa protein and the 29-kDa protein were identified respectively as rho and the GDP dissociation inhibitor (GDI), suggesting that rho and GDI copurify from bovine neutrophil cytosol in the form of a complex . The presence of a number of amino acid residues specific of rho A in the enzymatic digest originating from rho indicates that, among the rho proteins, at least rho A belongs to the GDI-rho complex. Biochem Biophys Res Commun, 1992 Dec 15, 189(2), 807 - 12 Site-directed mutagenesis of conserved residues of Clostridium thermocellum endoglucanase CelC; Navas J et al.; Four conserved residues of Clostridium thermocellum endoglucanase CelC were replaced by site-directed mutagenesis . Proteins mutated in His-90, Asn-139 and Glu-140 showed strongly reduced activity, in agreement with predictions of sequence alignments . Mutations in Glu-140 did not result in any detectable change in Km, or apparent size, suggesting that Glu-140 is directly involved in catalysis . The pH optimum of the proteins carrying the Glu-140/Ala and Glu140/Gln mutations was lower than that of the wild type, whereas the activity vs . pH profile of Glu-140/Asp CelC was similar to that of the wild type, suggesting that Glu-140 may act as a proton donor. FEMS Microbiol Lett, 1992 Dec 15, 79(1-3), 147 - 53 Possible function of tRNA(Thr)ACG in regulation of solvent formation in Clostridium acetobutylicum; Sauer U et al.; The mutation of Clostridium acetobutylicum mutant AA2, defective in the formation of acetone and butanol, was shown to be caused by a single insertion of Tn916 close to the structural gene thrA, encoding the tRNA(Thr)ACG . The DNA region containing the thrA gene was cloned and sequenced . Start and end points of the transcript were determined by primer extension and S1-mapping analysis . The results obtained were identical to predictions derived from the DNA sequence by various RNA-analysing computer programs . The rarely used ACG codon seems to be confined to genes expressed at the end of the exponential growth phase or involved in uptake or turnover of minor C or N substrates . Evolutionary aspects of this codon selection and a possible translational regulation mechanism are discussed. Clin Infect Dis, 1992 Dec, 15(6), 950 - 4 Clostridium sordellii bacteremia: case report and review; Spera RV Jr et al.; Clostridium sordellii is a gram-positive, anaerobic bacillus that has rarely been implicated as a human pathogen . It produces several exotoxins, which contribute to the progressive edema and refractory shock frequently seen with human infection . There have been eight prior reports of bacteremic C . sordellii infection and seven prior reports of nonbacteremic infections not due to myonecrosis of skeletal muscle . Mortality was 50% in the bacteremic group and 71% in the nonbacteremic group . Mortality correlated with both shock and leukemoid reaction at presentation . We present a case of C . sordellii sepsis in an asplenic patient with sickle beta thalassemia and inflammatory bowel disease, and we review the literature. Med J Aust, 1992 Dec 7-21, 157(11-12), 751 - 4 Crocodile attacks in the Northern Territory of Australia; Mekisic AP et al.; OBJECTIVE: To examine crocodile attacks in the Northern Territory with particular reference to risk factors, range of injuries, microorganisms isolated from wounds, and surgical management; and to make recommendations for optimal treatment . DESIGN AND SETTING: The case notes of patients treated at the Royal Darwin Hospital within the last decade were reviewed retrospectively . Autopsy and newspaper reports for the same period were also reviewed . RESULTS: There were 16 reported crocodile attacks in Northern Territory waters from June 1981 to June 1991 . Four of these were fatal . Most attacks resulted from swimming or wading in shallow water (13/16) . Half the victims were known to be affected by alcohol . The majority of attacks occurred in failing light or at night (10/16) . Injuries in survivors ranged from minor lacerations and puncture wounds to major abdominal, chest and limb trauma . Death in fatal attacks was caused by transection of the torso or decapitation . Microorganisms isolated from wound swabs included Pseudomonas, Enterococcus, Aeromonas and Clostridium species . CONCLUSIONS: Most attacks in this series could have been prevented by taking adequate precautions . The treatment of crocodile injuries must include (i) adequate wound cultures, (ii) antitetanus prophylaxis, (iii) meticulous wound debridement, (iv) appropriate broad spectrum prophylactic antibiotics and (v) allowing healing by secondary intention or delayed primary closure where appropriate. FEMS Microbiol Lett, 1992 Dec 1, 78(2-3), 251 - 5 Plasmid localization of a type E botulinal neurotoxin gene homologue in toxigenic Clostridium butyricum strains, and absence of this gene in non-toxigenic C . butyricum strains; Hauser D et al.; It has been shown recently that two Clostridium butyricum strains (ATCC 43181 and ATCC 43755) contain a botulinal neurotoxin type E (BoNT/E) gene closely related to that of C . botulinum type E . In this study, we show that this gene is located on a large plasmid in the two toxigenic C . butyricum strains and is absent in 18 non-toxigenic C . butyricum and C . beijerinckii strains . Interestingly, the 230 bp upstream and the 1260 bp downstream of the neurotoxin coding sequence are not present in either the non-toxigenic C . butyricum or C . beijerinckii strains . Our data suggest a BoNT/E gene transfer from C . botulinum E to originally non-toxigenic C . butyricum strains. FEMS Microbiol Lett, 1992 Dec 1, 78(2-3), 181 - 6 Identification of the cellulose-binding domain of the cellulosome subunit S1 from Clostridium thermocellum YS; Poole DM et al.; The 3' region of a gene designated cipB, which shows strong homology with cipA that encodes the cellulosome SL subunit of Clostridium thermocellum ATCC 27405, was isolated from a gene library of C . thermocellum strain YS . The truncated S1 protein encoded by the cipB derivative bound tightly to cellulose . The cellulose-binding domain in this polypeptide consisted of a C-terminal proximal 167 residue sequence which showed complete identity with residues 337-503 of mature SL from C . thermocellum strain ATCC 27405 . The cellulose-binding domain interacted with both crystalline and amorphous cellulose, but not with xylan. Toxicon, 1992 Dec, 30(12), 1583 - 9 Effect of oral Saccharomyces boulardii treatment on the activity of Clostridium difficile toxins in mouse digestive tract; Corthier G et al.; Human antibiotic-associated diarrhoea and pseudomembranous colitis are partly due to toxin production by Clostridium difficile . It is now well documented that Saccharomyces boulardii protects against C . difficile induced diseases . In an attempt to understand better the mechanism of this protective effect, the action of S . boulardii on a crude toxin preparation was studied in vitro and in vivo . The results showed that the yeast had no effect on the toxins in vitro but was able to protect mice inoculated with these toxins . Furthermore, the observation by scanning electron microscopy that the mucosa of S . boulardii protected mice was not damaged suggest that the yeast mainly acts on the intestinal mucosa. Antimicrob Agents Chemother, 1992 Dec, 36(12), 2850 - 1 Influence of ceftriaxone on emergence of Clostridium difficile; Lejko-Zupanc T et al.; The influence of long-term ceftriaxone administration on the emergence of Clostridium difficile was studied with 80 patients receiving ceftriaxone for 14 days . In five patients (6.3%) C . difficile was cultured . The appearance of gastrointestinal disturbances during treatment with ceftriaxone was common, but the rate of emergence of C . difficile in immunocompetent patients was not high. Toxicol Lett, 1992 Dec, 64-65 Spec No, 695 - 9 Cell injury and death caused by bacterial protein toxins; Donelli G et al.; Bacterial protein toxins, such as Clostridium difficile toxin A and the Escherichia coli cytotoxic necrotizing factor 1 are known to exert their cytotoxic action via a modification of some cytoskeletal components . The changes in actin organization caused by these toxins appear to be the primary events in the mechanism leading to cell death. Surg Gynecol Obstet, 1992 Dec, 175(6), 523 - 7 Surgical laparoscopic experience during the first year on a teaching service; Miller RE et al.; Recently, general surgeons have become actively involved in laparoscopic operations . The best method for teaching these techniques to surgical residents is unclear . Since June 1990, at St . Luke's-Roosevelt Hospital Center in New York City, we have instituted a formal course of instruction for surgical residents . This includes a reference syllabus, didactic instruction, use of an inanimate training device and a hands-on practice in swine . Clinically, the residents progress from observer to camera operator and, finally, operator . During the first year of this program, the authors performed 90 laparoscopic cholecystectomies, of which 71 were elective and 19 were for acute cholecystitis . There were seven morbidly obese patients, while 25 had undergone prior abdominal operations . The first 25 operations performed by the authors averaged 93.2 minutes, while the last 40 operations performed primarily by the surgical residents with assistance of the authors averaged 70 minutes . There were nine complications, including postoperative pancreatitis in two patients, Clostridium difficile enterocolitis in two and one each of prolonged paralytic ileus, postoperative transfusion and umbilical incision dehiscence . Two patients had postoperative common duct stones . There were no wound infections, bile duct injuries or deaths . Complications were evenly distributed throughout the series and did not correlate with whether the surgeon was a resident or an attending surgeon . The results of this plan have been quite successful and thus far, 12 residents have completed this program. J Pediatr, 1992 Dec, 121(6), 915 - 7 Pseudomembranous colitis in sickle cell disease responding to exchange transfusion; Baruchel S et al.; We report a case of ischemic colitis with pseudomembrane formation in a 6 1/2-year-old boy with sickle cell disease that responded to medical management including exchange transfusion . This case was not associated with Clostridium difficile infection but with an elevation of serum levels of tumor necrosis factor alpha . This rare complication of sickle cell disease has been reported only in adults. Cancer, 1992 Dec 1, 70(11), 2658 - 63 Emphysematous cystitis due to Clostridium perfringens and Candida albicans in two patients with hematologic malignant conditions; Greene MH; BACKGROUND . Fever, abdominal pain, and hematuria developed in two patients with hematologic malignant conditions (multiple myeloma and agnogenic myeloid metaplasia) . Each patient was found to have emphysematous cystitis (EC), secondary to Clostridium perfringens and Candida albicans, respectively . Both patients had debilitated general medical conditions, compromised immune function, prior treatment with broad-spectrum antibiotics and corticosteroids, bladder outlet obstruction, and indwelling Foley catheters as predisposing factors to EC . Neither was diabetic . METHODS . These cases provide an opportunity to review the related medical literature on the pathophysiology and management of this uncommon entity . RESULTS . Treatment consists of control of underlying diabetes (if present), administration of appropriate antibiotics, establishment of urinary drainage, provision of supportive general medical care, exclusion of the presence of a bladder fistula, and surgical debridement only when unavoidable . CONCLUSIONS . EC should be part of the differential diagnosis in patients with cancer who have fever, abdominal pain, and hematuria. J Bacteriol, 1992 Dec, 174(23), 7642 - 7 Differential expression of a Clostridium acetobutylicum antisense RNA: implications for regulation of glutamine synthetase; Fierro-Monti IP et al.; The Clostridium acetobutylicum glutamine synthetase (GS) DNA region is characterized by a downstream promoter, P3, oriented toward the glnA gene, which controls the transcription of an RNA complementary to the start of the glnA mRNA . Expression of the predicted 43-base antisense RNA was demonstrated in C . acetobutylicum and Escherichia coli cells containing the cloned glnA DNA . Antisense RNA transcription from P3 was not regulated by nitrogen in E . coli cells, but the expression of antisense RNA was associated with decreased levels of GS activity . In C . acetobutylicum, GS activity and the transcription of glnA mRNA and antisense RNA were regulated by nitrogen . GS activity and glnA mRNA were repressed in cells grown in nitrogen-rich medium . Repression ratios for GS activity varied from 1.6 to 9.0, depending on the sampling time . The relative number of glnA transcripts was approximately 25% lower in cells grown for 72 h in nitrogen-rich medium than in cells grown in nitrogen-limiting medium . This finding contrasted with the expression of antisense RNA, which was repressed in nitrogen-limiting medium but induced in nitrogen-rich medium . The relative number of antisense RNA transcripts was increased approximately sixfold in cells grown in nitrogen-rich medium . There was a 1.6-fold excess of antisense RNA over glnA mRNA under conditions that repressed GS activity . Under conditions that induced GS activity, glnA mRNA transcripts exceeded antisense RNA transcripts by fivefold. J Bacteriol, 1992 Dec, 174(24), 8158 - 62 Analysis of operons encoding 23S rRNA of Clostridium botulinum type A; East AK et al.; Southern hybridization analysis of Clostridium botulinum type A chromosomal DNA indicated the presence of six copies of the 23S rRNA gene . Fragments of DNA encoding 23S rRNA were amplified by polymerase chain reaction and cloned in Escherichia coli . Three clones examined by restriction enzyme and sequence analysis were found to be derived from different operons . Sequence determination of the entire insert of two clones revealed nine nucleotide changes in the genes coding for 23S rRNA (99.7% sequence identity) between operons encoded on the same chromosome, showing microheterogeneity in the rRNA operons of this organism. Appl Environ Microbiol, 1992 Dec, 58(12), 4032 - 7 Purification and properties of NADP-dependent glutamate dehydrogenase from Ruminococcus flavefaciens FD-1; Duncan PA et al.; Glutamate dehydrogenase (GDH) (L-glutamate:NADP+ oxidoreductase, deaminating, EC 1.4.1.4) from the cellulolytic ruminal bacterium Ruminococcus flavefaciens has been purified and characterized . The native enzyme and subunit are 280 and 48 kDa, respectively, suggesting that the native enzyme is a hexamer . The enzyme requires 0.5 M KCl for optimal activity and has a pH optimum of 6.9 to 7.0 . The Kms for ammonia,