Clostridia

J Clin Microbiol, 1985 Dec, 22(6), 980 - 3
Evaluation of the RapID-ANA system for identification of anaerobic bacteria of veterinary origin; Adney WS et al.; This study evaluated the ability of the RapID-ANA system (Innovative Diagnostic Systems, Inc., Atlanta, Ga.) to accurately identify a spectrum of freshly isolated veterinary anaerobes . A total of 183 isolates were tested and included 7 Actinomyces spp., 53 Bacteroides spp., 32 Clostridium spp., 2 Eubacterium spp., 65 Fusobacterium spp., 1 Peptococcus spp., 22 Peptostreptococcus spp., and 1 Propionibacterium spp . All isolates were initially identified by conventional biochemical testing and gas-liquid chromatography of short-chain fatty acid metabolites . Additional tests were performed as required by the RapID-ANA system . Of these isolates, 81.4% were correctly identified to the genus level, including 59.6% to the species level, 14.2% were incorrectly identified at the genus level, and 4.4% were not identified . Initially, 20.2% of the strains were not identified because the microcodes were not in the code book . The majority of the incorrect identifications were caused by the misidentification of Fusobacterium spp . as Bacteroides spp . Errors also occurred when veterinary anaerobes not included in the data base were assigned an identification from the existing data base . The RapID-ANA system appears to be a promising new method for rapid identification of veterinary anaerobes; however, further evaluation with an extended data base is needed before the system can accurately identify all clinically significant anaerobes.

J Clin Microbiol, 1985 Dec, 22(6), 962 - 7
Rapid identification of Clostridium species by high-pressure liquid chromatography; Harpold DJ et al.; High-pressure liquid chromatography was evaluated as a rapid means of identifying various species of clostridia . Isolates were inoculated into a defined medium and incubated aerobically for 1 h at 35 degrees C . The organisms were removed, and the supernatants were derivatized for 1 min at room temperature by the addition of o-phthalaldehyde . The total time required to run each chromatogram was approximately 50 min . Standardized peak heights for each medium component and any new peaks formed were calculated for each isolate and compared with those for uninoculated control medium . Multiple isolates of various Clostridium species gave consistent patterns of medium utilization that could be used for identification . This rapid method can easily be adapted for laboratory use and has the potential for automation.

Antimicrob Agents Chemother, 1985 Dec, 28(6), 832 - 3
Pharmacokinetic and therapeutic trial of sultamicillin in acute sinusitis; Jones S et al.; Sultamicillin, an antibiotic combining ampicillin and the beta-lactamase inhibitor sulbactam, was administered to 13 patients diagnosed as having acute sinusitis . Specimens from sinus were obtained for all 13 patients by transantral puncture . Pharmacokinetics, bacteriology, and therapeutic efficacy were assessed . Eighty-five percent (11 of 13) were cured; two treatment failures were subsequently shown to have chronic (rather than acute) sinusitis during surgical exploration . Diarrhea was frequently encountered, and Clostridium difficile-associated enteritis was documented for one patient . Beta-lactamase-producing organisms were not encountered in this study; however, this study provides impetus for further controlled clinical trials.

Arch Biochem Biophys, 1985 Dec, 243(2), 447 - 53
Enzyme elements involved in the interconversion of L-carbamylaspartate and L-dihydroorotate by dihydroorotase from Clostridium oroticum; Pettigrew DW et al.; Enzyme elements that are involved in the reversible cyclization of L-carbamylaspartate to L-dihdroorotate catalyzed by dihydroorotase (EC 3.5.2.3) from Clostridium oroticum (ATCC 25750) have been studied . Removal of Zn(II) from the enzyme by chelators followed by incubation of apoenzyme with Co(II) results in replacement of two to three of the four Zn(II) ions per molecule by Co(II) . The catalytic properties of the Zn(II)Co(II) dihydroorotase are different from those of native enzyme . The Vmax is increased for both the synthesis and hydrolysis of L-dihydroorotate . The Km for L-dihydroorotate is unchanged, while the Km for L-carbamylaspartate is increased more than twofold . On the other hand, the kinetic properties of Zn(II)-reconstituted dihydroorotase are indistinguishable from those of native enzyme . The pH dependence of Vmax is also altered by the Co(II) substitution . For both Zn(II)- and Zn(II)Co(II)-dihydroorotase, this pH dependence is well described by a single ionization and the pK's for L-dihydroorotate synthesis and hydrolysis are different . Substitution with Co(II) increases the pK for both reaction directions to different extents . These results strongly support a role for the tightly bound metals in the catalytic mechanism . In addition, diethylpyrocarbonate rapidly inactivates the enzyme . The inactivation is prevented by L-dihydroorotate . This result is consistent with a role for at least one histidine in catalysis . The possibility that C . oroticum dihydroorotase may be useful model for the more complex mammalian enzyme is considered.

Infect Immun, 1985 Dec, 50(3), 844 - 51
Effect of thiol-activated toxins (streptolysin O, alveolysin, and theta toxin) on the generation of leukotrienes and leukotriene-inducing and -metabolizing enzymes from human polymorphonuclear granulocytes; Bremm KD et al.; The generation of leukotrienes (LTC4, LTD4, LTE4, and LTB4; 12-epi-LTB4 isomer) from human granulocytes by thiol-activated toxins (streptolysin O, alveolysin from Bacillus alvei, and theta toxin from Clostridium perfringens) is described . The release occurs under noncytolytic conditions . Although LTB4 is the major component after calcium ionophore stimulation, more LTC4 as compared with LTB4 is released with the toxins . The 5-lipoxygenase pathway of toxin-mediated activation can effectively be inhibited by caffeic acid, a lipoxygenase inhibitor . The toxins also induce the release of leukotriene-metabolizing enzymes such as gamma-glutamyltranspeptidase, which transfers LTC4 into LTD4, and dipeptidase, which metabolizes LTD4, into LTE4 . Dipeptidase activity is more pronounced than the gamma-glutamyltranspeptidase activity but still does not reach the levels obtained when cells were triggered with opsonized zymosan.

CMAJ, 1985 Dec 1, 133(11), 1141 - 6
Food-borne botulism in Canada, 1971-84; Hauschild AH et al.; Sixty-one outbreaks of food-borne botulism involving a total of 122 cases, of which 21 were fatal, were recorded from 1971 to 1984 in Canada . Most occurred in northern Quebec, the Northwest Territories or British Columbia . Of the 122 victims 113 were native people, mostly Inuit . Most of the outbreaks (59%) were caused by raw, parboiled or "fermented" meats from marine mammals; fermented salmon eggs or fish accounted for 23% of the outbreaks . Three outbreaks were attributed to home-preserved foods, and one outbreak was attributed to a commercial product . The causative Clostridium botulinum type was determined in 58 of the outbreaks: the predominant type was E (in 52 outbreaks), followed by B (in 4) and A (in 2) . Renewed educational efforts combined with a comprehensive immunization program would significantly improve the control of botulism in high-risk populations.

Eur J Epidemiol, 1985 Dec, 1(4), 264 - 73
Morphological alterations and changes in cellular cations induced by Clostridium perfringens type A enterotoxin in tissue culture cells; Sugimoto N et al.; The morphological alterations (bleb-balloon formation) induced by Clostridium perfringens type A enterotoxin in HeLa and Vero cells were studied under defined extracellular conditions . The action of enterotoxin was found to depend on the temperature but not on energy metabolism . The morphological alterations by the enterotoxin occurred in phosphate buffered saline containing Ca2+ and Mg2+ . Of the constituents of the buffered saline, Ca2+ was essential for the morphological alterations and other ions were interchangeable . The morphological alterations by the enterotoxin occurred also in 10 mM Hepes-Na buffer, pH 7.2 containing NaCl, KCl or choline chloride at a concentration of over ca . 50 mM and in 10 mM Hepes-Ca buffer, pH 7.2 containing CaCl2 at a concentration of over ca . 50 mM . Addition of sucrose to the medium prevented induction of the morphological alterations . The amount of sucrose necessary to protect the cells increased with increase in NaCl, KCl or CaCl2 concentration in the medium . A calcium ionophore A23187 mimicked the action of enterotoxin . Examination of the cation contents of the cells by atomic absorption spectrophotometry showed early and rapid increase of Ca2+ during intoxication with concomitant changes in Na+, K+ and Mg2+ that reduced the ion concentration gradients between inside and outside of the cell present before toxin treatment . The mechanism of action of C . perfringens type A enterotoxin is discussed on the basis of these findings.

J Bacteriol, 1985 Dec, 164(3), 1162 - 70
Regulation and genetic enhancement of beta-amylase production in Clostridium thermosulfurogenes; Hyun HH et al.; We studied the general mechanism for regulation of beta-amylase synthesis in Clostridium thermosulfurogenes . beta-Amylase was expressed at high levels only when the organism was grown on maltose or other carbohydrates containing maltose units . Three kinds of mutants altered in beta-amylase production were isolated by using nitrosoguanidine treatment, enrichment on 2-deoxyglucose, and selection of colonies with large clear zones on iodine-stained starch-glucose agar plates . beta-Amylase was produced only when maltose was added to cells growing on sucrose in wild-type and catabolite repression-resistant mutant strains, but the differential rate of enzyme synthesis in constitutive mutants was constant regardless of the presence of maltose . In carbon-limited chemostats of wild-type and catabolite repression-resistant mutant stains, beta-amylase was expressed on maltose but not on glucose or sucrose . beta-Amylase synthesis was immediately repressed by the addition of glucose . Therefore, we concluded that beta-amylase synthesis in C . thermosulfurogenes was inducible and subject to catabolite repression . The addition of cAMP did not eliminate the repressive effect of glucose . The mutants were generally characterized in terms of beta-amylase production, growth properties, fermentation product formation, and alterations in glucose isomerase and glucoamylase activities . A hyperproductive mutant produced eightfold more beta-amylase on starch medium than the wild type and more rapidly fermented starch to ethanol.

Experientia, 1985 Nov 15, 41(11), 1435 - 7
Increased formation of arginine deiminase by Clostridium perfringens FD-1 growing in the presence of caffeine; Sacks LE; Caffeine slowed growth and markedly increased the formation of arginine deiminase in growing C . perfringens FD-1 when dextrin, but not maltose or maltotriose, served as the energy source . It is postulated that the ability of caffeine to induce arginine deiminase is related to an inhibition of polysaccharide utilization, resulting in a shift-down condition known to induce arginine deiminase and other enzymes in bacteria.

S Afr Med J, 1985 Nov 9, 68(10), 760 - 2
{Gas gangrene: A discussion of 3 cases and review of the literature}; du Toit DF et al.; Three patients with gas gangrene of the lower limbs are presented . In 2 of the 3 patients gas gangrene developed after lower-limb amputation, indications for amputation being atherosclerotic and diabetic gangrene . In the third patient associated leukaemia was diagnosed . All 3 patients presented with the typical clinical manifestations of gas gangrene . Clostridium perfringens was isolated from the affected leg in each patient . The current application of surgery and hyperbaric oxygen therapy in the treatment of gas gangrene is discussed.

J Immunol Methods, 1985 Nov 7, 83(2), 241 - 8
An immunochemical method for fingerprinting Clostridium difficile; Sharp J et al.; The use of sodium dodecyl sulphate-polyacrylamide gel electrophoresis in association with electrophoretic transfer of proteins to nitrocellulose and subsequent probing with antisera appears useful as a method for fingerprinting Clostridium difficile . Thorough testing of the stability of the antigenic nature of isolates of the organism during subculture and antigen preparation has shown it to be remarkably stable both in vitro and in vivo . Minor differences in the method of antigen extraction do not markedly alter the immunoblot patterns produced . It has also been demonstrated that an individual may harbour more than one strain of the organism at any one time . Results show the possible usefulness of this technique in studying the epidemiology of diarrhoeal disease known to be associated with C . difficile . It is suggested that for any serious study several colonies should be subcultured from the primary isolation plate.

J Biol Chem, 1985 Nov 5, 260(25), 13509 - 12
13C NMR studies of butyric fermentation in Clostridium kluyveri; Smith GM et al.; The fermentation of 13C-labeled ethanol and acetate into butyrate and caproate by Clostridium kluyveri has been studied by using 13C NMR . The pathway involves the conversion of both ethanol and acetate into acetyl coenzymes A, two of which condense to form CoA-linked precursors of butyrate . If butyryl-CoA is involved in the condensation, caproate is the ultimate product . ATP is produced from acetyl-CoA via the reactions catalyzed by phosphotransacetylase and acetate kinase with acetate, a required carbon source, as a co-product . In spectra of whole cells incubated with the labeled carbon sources, label from ethanol appears rapidly in acetate, which then reaches a lower, steady-state concentration due to its re-entry into the pathway . The rapid initial production of acetate indicates equally rapid production of ATP . Label from acetate appears in ethanol only if ethanol is already present, indicating that this process is one of isotopic equilibration rather than net synthesis of ethanol from acetate . The ratio of butyrate to caproate produced depends strongly on the initial ratio of ethanol to acetate in the medium . The relative rates of utilization of ethanol and acetate vary as the fermentation proceeds . 13C-13C coupling in the butyrate and caproate produced from {1-13C}ethanol and {2-13C}acetate can be used to determine if the acetyl-CoA molecules arising from ethanol and acetate enter the same pool or if they remain separated . The data are consistent with random mixing of the acetyl-CoA produced from the two carbon sources.

Biochemistry, 1985 Nov 5, 24(23), 6527 - 33
Mode of hydrolysis of collagen-like peptides by class I and class II Clostridium histolyticum collagenases: evidence for both endopeptidase and tripeptidylcarboxypeptidase activities; Mookhtiar KA et al.; The action of three class I (beta, gamma, and eta) and three class II (delta, epsilon, and zeta) collagenases from Clostridium histolyticum on two series of peptides with collagen-like sequences has been examined . The peptides in the first series all contain 4-nitrophenylalanyl-Gly-Pro-Ala in subsites P1 through P3', but each is successively lengthened in the N-terminal direction by addition of an appropriate residue until subsite P5 is occupied . The second group of peptides all have cinnamoyl-Leu in subsites P2 and P1, respectively, but each is successively lengthened in the C-terminal direction by partial additions of the Gly-Pro-Leu triplet until subsite P6' is occupied . N-Terminal elongation causes the kcat/KM values to rise markedly and to level off after occupancy of subsite P6 for the class I enzymes and subsite P3 for the class II enzymes . C-Terminal elongation produces the best substrates for both classes of enzymes when subsites P3' or P4' are occupied by amino acids with free carboxyl groups . The kcat/KM values for the hydrolysis of both Leu-Gly bonds of cinnamoyl-Leu-Gly-Pro-Leu-Gly-Pro-Leu have been measured for both classes of enzymes . Both rates are large, but both classes preferentially hydrolyze the Leu-Gly bond of the C-terminal triplet . Thus, both classes of enzymes exhibit both endopeptidase and tripeptidylcarboxypeptidase activities.

Biochemistry, 1985 Nov 5, 24(23), 6520 - 6
Complementary substrate specificities of class I and class II collagenases from Clostridium histolyticum; Van Wart HE et al.; The substrate specificities of three class I (beta, gamma, and eta) and three class II (sigma, epsilon, and zeta) collagenases from Clostridium histolyticum have been investigated by quantitating the kcat/KM values for the hydrolysis of 53 synthetic peptides with collagen-like sequences covering the P3 through P3 subsites of the substrate . For both classes of collagenases, there is a strong preference for Gly in subsites P1' and P3 . All six enzymes also prefer substrates that contain Pro and Ala in subsites P2 and P2' and Hyp, Ala, or Arg in subsite P3' . This agrees well with the occupancies of these sites by these residues in type I collagen . However, peptides with Glu in subsites P2 or P2' are not good substrates, even though Glu occurs frequently in these positions in collagen . Conversely, all six enzymes prefer aromatic amino acids in subsite P1, even though such residues do not occur in this position in type I collagen . In general, the class II enzymes have a broader specificity than the class I enzymes . However, they are much less active toward sequences containing Hyp in subsites P1 and P3' . Thus, the two classes of collagenases have similar but complementary sequence specificities . This accounts for the ability of the two classes of enzymes to synergistically digest collagen.

Appl Environ Microbiol, 1985 Nov, 50(5), 1258 - 61
Inhibitory effect of a copper-dipeptide complex on the establishment of a Clostridium perenne strain in the intestinal tract of gnotobiotic mice; Dubos F et al.; A semisynthetic diet fed to axenic mice was found to prevent the establishment of a Clostridium perenne strain in their intestinal tract . This inhibitory effect did not occur when axenic mice were preinoculated with a strain of Clostridium difficile . The inhibitory effect was related to the presence in the intestinal contents of axenic mice of both dietary copper and a dipeptide, aspartic-epsilon-lysine . When C . difficile was inoculated into axenic mice, the dipeptide disappeared from the digesta, and C . perenne became established even in the presence of high concentrations of copper.

Appl Environ Microbiol, 1985 Nov, 50(5), 1238 - 43
Isolation and characterization of butanol-resistant mutants of Clostridium acetobutylicum; Hermann M et al.; In a wild-type strain of Clostridium acetobutylicum isolated from soil, solvent production appeared limited by butanol toxicity . Butanol-resistant mutants have been obtained which produced significantly higher solvent concentrations (about 30%) than the wild-type strain . Some other physiological differences were observed between a selected resistant mutant and the wild-type strain at the level of solvent resistance and sporulation.

Clin Nephrol, 1985 Nov, 24(5), 242 - 8
Clostridium difficile-associated colitis in uremic patients; Leung AC et al.; Five uremic patients managed in a renal unit developed Clostridium difficile-associated colitis . Four cases occurred in a cluster at about the same time . All patients had previously received or were on antibiotic therapy at the onset of diarrhea and one patient was also on oral steroid therapy . Cefotaxime, a third generation cephalosporin was involved in all five cases . All patients had severe diseases with explosive diarrhea and systemic toxicity . The diagnosis was confirmed in all cases by culture of C . difficile and demonstration of high titers of C . difficile cytotoxin in the stool . Histology from rectal biopsy in one patient showed classical pseudomembranous colitis . Response to treatment with vancomycin was generally good though one patient had two relapses . Uremic patients have impaired immune response and intestinal motility and are predisposed to C . difficile infection . Cross-infection can occur and the isolation of affected patients seems prudent.

Biol Chem Hoppe Seyler, 1985 Nov, 366(11), 1057 - 62
Observations on the elimination of water from 2-hydroxy acids in the metabolism of amino acids by Clostridium sporogenes; Machacek-Pitsch C et al.; Cell-free extracts of Clostridium sporogenes catalyse the water elimination from (2R)-phenyllactate in the presence of one of the energy-rich compounds acetyl-CoA, acetylphosphate or ATP and coenzyme A . Water is eliminated from (2R)-phenyllactoyl-CoA without any of the aforementioned additions . Cinnamoyl-CoA also acts catalytically . One molecule of cinnamoyl-CoA causes the elimination of water from more than 8 molecules phenyllactate . This is important from an energetic point of view since less than 2 mol ATP are formed per 2-3 mol metabolized amino acids . An activation of the hydroxy group of the alpha-hydroxy acid in form of a phosphate ester can also be excluded for energetic reasons.

Obstet Gynecol, 1985 Nov, 66(5), 737 - 8
Clostridium difficile colitis associated with single-dose cefazolin prophylaxis; McNeeley SG Jr et al.; Diarrhea and pseudomembranous colitis are associated with antimicrobial therapy and prophylaxis . Clostridium difficile colitis occurred in a patient who received a single dose of cefazolin for prophylaxis at cesarean section . Prompt remission occurred after treatment with oral vancomycin.

J Clin Microbiol, 1985 Nov, 22(5), 873 - 6
Survey of neuraminidase production by Clostridium butyricum, Clostridium beijerinckii, and Clostridium difficile strains from clinical and nonclinical sources; Popoff MR et al.; Neuraminidase production was investigated in 57 Clostridium butyricum strains, 16 Clostridium beijerinckii strains, and 25 Clostridium difficile strains . Neuraminidase activity was found only in C . butyricum strains originating from one human newborn with neonatal necrotizing enterocolitis, two newborns with hemorrhagic colitis, one infected placenta, and one adult with peritonitis, It was concluded that neuraminidase was not a major virulence factor in C . butyricum strains.

J Neurochem, 1985 Nov, 45(5), 1487 - 94
Solubilization of membrane-bound acetylcholinesterase by a phosphatidylinositol-specific phospholipase C; Futerman AH et al.; Phosphatidylinositol-specific phospholipase C (PIPLC) quantitatively solubilizes acetylcholinesterase (AChE) from purified synaptic plasma membranes and intact synaptosomes of Torpedo ocellata electric organ . The solubilized AChE migrates as a single peak of sedimentation coefficient 7.0S upon sucrose gradient centrifugation, corresponding to a subunit dimer . The catalytic subunit polypeptide of AChE is the only polypeptide detectably solubilized by PIPLC . This selective removal of AChE does not affect the amount of acetylcholine released from intact synaptosomes upon K+ depolarization . PIPLC also quantitatively solubilizes AChE from the surface of intact bovine and rat erythrocytes, but only partially solubilizes AChE from human and mouse erythrocytes . The AChE released from rat and human erythrocytes by PIPLC migrates as a approximately 7S species on sucrose gradients, corresponding to a catalytic subunit dimer . PIPLC does not solubilize particulate AChE from any of the brain regions examined of four mammalian species . Several other phospholipases tested, including a nonspecific phospholipase C from Clostridium welchii, fail to solubilize AChE from Torpedo synaptic plasma membranes, rat erythrocytes, or rat striatum.

Gastroenterology, 1985 Nov, 89(5), 1038 - 45
Antibiotic-associated colitis due to Clostridium difficile: double-blind comparison of vancomycin with bacitracin; Young GP et al.; A randomized double-blind study was carried out in patients with unresolving antibiotic-associated colitis due to Clostridium difficile, to compare the effect of bacitracin (80,000 U/day) with vancomycin (500 mg/day) on the resolution of symptoms, clearance of organism, and prevention of relapse . Forty-two patients with colitis, 9 of whom had a pseudomembrane, were randomized, 21 patients to each treatment group . The two groups were comparable in age, disease severity, and antibiotic exposure . For a 50% reduction in stool frequency the mean times (+/- SE) were 4.1 +/- 0.4 days for bacitracin and 4.2 +/- 0.4 days for vancomycin . Sixteen patients (76%) had symptom resolution after 7 days of treatment with bacitracin, compared with 18 patients (86%) given vancomycin . Patients who failed to respond were crossed over (blind) to the alternative antibiotic, but tended to be refractory to the alternative medication as well . Vancomycin-treated patients had negative toxin (83% vs . 53%, p = 0.04) and negative stool cultures (81% vs . 52%, p = 0.02) more frequently than did those patients given bacitracin . Similar numbers of patients in each group had symptomatic relapse during 1 mo of follow-up, but most of them relapsed yet again after blinded crossover therapy . Although bacitracin was significantly less effective than vancomycin in clearing C . difficile from the stools, both were of similar value in the control of symptoms in a group of patients with predominantly nonpseudomembranous colitis . In view of its low cost, bacitracin is a reasonable first-line alternative to vancomycin in the treatment of antibiotic-associated colitis.

J Appl Bacteriol, 1985 Nov, 59(5), 469 - 78
Multiple modes of inhibition of spore germination and outgrowth by reduced pH and sorbate; Blocher JC et al.; Germination and outgrowth of three strains of Clostridium botulinum in PYEG medium were measured by phase contrast microscopy . Reduction in pH from 7 to 5.5 completely inhibited germination of strain 12885A, reduced the extent of germination of strain 62A and had no effect on the extent of germination of strain 53B . At pH 5.5, 225 mg/l of undissociated sorbic acid had no effect on the germination of strain 53B, while at pH 6.5, 225 mg/l of undissociated sorbic acid completely inhibited germination of strains 62A and 12885A . Outgrowth of germinated spores of strains 62A and 53B was not inhibited at pH 5.5, but the addition of sorbate (225 mg/l undissociated sorbic acid) completely inhibited outgrowth . Sorbate inhibited germination of Cl . botulinum and Bacillus cereus spores triggered to germinate by amino acids . Inhibition occurred after germinant binding, as measured by commitment to germinate.

Cancer Res, 1985 Nov, 45(11 Pt 2), 5714 - 21
Similar effects of phospholipase C and phorbol ester tumor promoters on primary mouse epidermal cells; Jeng AY et al.; Interaction of tumor promoting phorbol esters with specific high affinity receptors is probably essential for many of the biological responses elicited by these agents . Since diacylglycerols which can be produced enzymatically from phospholipids by phospholipase C are postulated to be the physiological ligands for the phorbol ester receptor, we have examined primary cultures of mouse epidermal basal cells exposed to phospholipase C (Clostridium perfringens) for several biological and biochemical responses characteristic of treatment with 12-O-tetradecanoyl-phorbol-13-acetate, the most potent phorbol ester tumor promoter . Formation of diacylglycerols by treatment with phospholipase C was demonstrated by the dose-dependent release of radioactive diacylglycerols in cells prelabeled with {3H}arachidonic acid . Treatment with phospholipase C at 0.05 units/ml for 30 min led to the morphological changes and to the reduction in epidermal growth factor binding (90%) associated with 12-O-tetradecanoylphorbol-13-acetate treatment . Continuous treatment at the same dose led to the induction of the enzymes ornithine decarboxylase and transglutaminase with a time course and extent similar to the inductions by 12-O-tetradecanoylphorbol-13-acetate . Treatment with phospholipase C at 0.1 enzyme unit/ml yielded substantial suppression of the binding affinity of phorbol-12,13-dibutyrate for its receptors without reduction in total number of binding sites, consistent with the production by phospholipase C of a competitive inhibitor of phorbol ester binding . Several diacylglycerols at concentrations of 250 microM and above effectively competed for phorbol-12,13-dibutyrate binding, reduced epidermal growth factor binding, and to a lesser extent induced ornithine decarboxylase and transglutaminase . These results support the hypothesis that diacylglycerols can act through the phorbol ester receptors and thus produce biological and biochemical responses similar to those of the phorbol esters.

Arch Surg, 1985 Nov, 120(11), 1321 - 2
Clostridium difficile colitis mimicking acute peritonitis; Drapkin MS et al.; Five patients receiving penicillin V potassium or a cephalosporin antibiotic for 18 hours to 22 days developed fever, marked leukocytosis, and signs and symptoms that suggested right-lower-quadrant peritoneal irritation . All underwent emergency laparotomy, at which dilatation and inflammation of the ascending colon were found . Only one of the patients had profuse diarrhea, and two patients had no diarrhea prior to laparotomy . Postoperatively, Clostridium difficile colitis was diagnosed by stool toxin assay and was confirmed in one case by proctosigmoidoscopic biopsy results . Antibiotic-associated colitis must be considered in any patient who develops peritoneal signs while or after receiving antibiotics . Over a two-year period, the "acute abdomen" presentation accounted for 5.2% of all patients with C difficile colitis at our institutions . Early proctosigmoidoscopy or stool examination for C difficile or its toxin may avoid unnecessary laparotomy in such patients.

Zentralbl Bakteriol Mikrobiol Hyg {A}, 1985 Nov, 260(3), 319 - 28
Correlative properties for a differentiation of two Clostridium sordellii phenotypes and their distinction from Clostridium bifermentans; Roggentin P et al.; As the present classification (19) of Clostridium sordellii and C . bifermentans is based on properties which are not conclusive for most of our strains, we investigated 80 strains from various origin of this group regarding 30 selected properties . Four of these properties were correlative and therefore particularly important for a distinct differentiation of the strains investigated: urease activity (U), growth inhibition by 1% mannose (M), arginine deaminase activity (A), and sialidase (EC 3.2.1.18) activity (S) . Concerning these four characters three clusters were formed: cluster I was positive for U, M, A, and S and comprised 36 strains including C . sordellii type strain (ATCC 9714T); cluster II was positive for M and S and negative for U and A and comprised twelve strains including strain ATCC 35392; and cluster III was positive for A and negative for U, M, and S and comprised 32 strains including C . bifermentans type strain (ATCC 638T) . Only two of the correlative properties (U and S, U and A, A and M, or A and S) needed to be tested to determine the affiliation of any strain of the C . sordellii/bifermentans group to one of the three clusters . Clusters I and II, representing two phenotypes of C . sordellii, can now clearly be distinguished from C . bifermentans . Sialidase formed by cluster I and II strains was inhibited by antibodies produced against cluster I strain sialidase . No cross reaction was found with other clostridial sialidases . Pathogenicity, hitherto considered as one of the distinctive properties of C . sordellii and C . bifermentans, was found with various strains of all the three clusters . Therefore, in the case of an infection caused by these two species, care should be taken as to the pathogenicity especially of C . bifermentans and treatment should be accordingly.

Rev Infect Dis, 1985 Nov-Dec, 7 Suppl 4, S789 - 93
Clinical experience with aztreonam in the treatment of gram-negative bacteremia; Scully BE et al.; Aztreonam was used for the treatment of gram-negative bacteremia in 101 patients . In 34 instances a second antibiotic was prescribed for the treatment of suspected or documented gram-positive or anaerobic infection . The sources of bacteremia were the urinary tract (50 patients), an intraabdominal site (17), the respiratory tract (8), an intravascular site (9), and an unknown site (17) . The clinical response rate was 92% (91 of 99 patients) . The bacteriologic response rate was 97% (98 of 101 patients) . In six of seven patients, Pseudomonas aeruginosa bacteremia was cured . Twelve patients developed superinfection with gram-positive cocci or Candida, and one patient developed diarrhea associated with Clostridium difficile . No other serious toxic effects were noted.

Rev Infect Dis, 1985 Nov-Dec, 7 Suppl 4, S579 - 93
Discovery and development of the monobactams; Sykes RB et al.; A novel procedure designed to detect naturally occurring beta-lactam-containing molecules led to isolation of the monobactams - structurally unique, bacterially produced, monocyclic beta-lactam antibiotics . Although none of these monobactams exhibited impressive antimicrobial activity, side-chain variation - as with the penicillins and cephalosporins - resulted in potently active compounds . Aztreonam was chosen from hundreds of compounds for extended laboratory studies . In addition to a unique chemical structure, aztreonam has biologic properties that are unique in comparison with those of the classical penicillins and cephalosporins . Aztreonam is relatively inactive against gram-positive bacteria and anaerobes but is extremely effective against aerobic gram-negative bacteria, including Pseudomonas aeruginosa . The drug is highly resistant to enzymatic hydrolysis by beta-lactamases, particularly those known to be mediated by R plasmids, and is a poor inducer of chromosomal beta-lactamases . In the majority of drug combinations tested, aztreonam exhibits additive or synergistic activity . In a series of animal-model infections, the drug showed a high degree of efficacy that was consistent with findings in studies in vitro . In a hamster model for Clostridium difficile-induced pseudomembranous colitis, aztreonam did not induce any significant changes.

Pathol Biol (Paris), 1985 Nov, 33(9), 899 - 903
{Sensitivity of strict anaerobic bacteria to metronidazole, cefoxitin, and clindamycin}; Dubreuil L et al.; The minimal inhibitory concentrations of three antimicrobial agents were determined by two fold dilution in Wilkins Chalgren agar for over 569 anaerobes isolated and collected from 20 French hospitals . Metronidazole, cefoxitin and clindamycin inhibited respectively 95, 97 and 91% of tested strains . No metronidazole resistant strains could be found among Bacteroides fragilis group, Fusobacterium and Clostridium other than C . perfringens . Cefoxitin was the most active against C . perfringens, Gram positive cocci and non sporulated bacilli . Clindamycin resistance was observed mainly with B . fragilis and Clostridium other than C perfringens . This phenomenon occurred in most hospitals where strains were isolated.

Pathol Biol (Paris), 1985 Nov, 33(9), 891 - 5
{Comparative sensitivity of 207 strains of strict anaerobic bacteria to piperacillin and 4 other antibiotics}; Reynaud A et al.; The MIC of two hundred and seven anaerobic bacterial strains was determined by an agar dilution method for five antibiotics (piperacillin, ampicillin, carbenicillin, cefalotin, metronidazole) . 100% of the strains were susceptible to carbenicillin (MIC less than or equal to 128 mg/l), 97% to piperacillin (MIC less than or equal to 16), 86% to metronidazole (MIC less than or equal to 4), 70% to ampicillin (MIC less than or equal to 4) and 68% to cefalotin (MIC less than or equal to 8) . Amongst beta-lactam compounds, piperacillin and ampicillin determined the lowest MIC for Bacteroides fragilis, Fusobacterium, Clostridium and Gram-positive cocci . Amongst Bacteroides fragilis strains, the lowest MIC were obtained with metronidazole.

Vet Clin North Am Food Anim Pract, 1985 Nov, 1(3), 509 - 14
Enterotoxemia in neonatal calves; Fleming S; The incidence, bacterial characteristics, disease syndromes, diagnosis, treatment, and prevention of enterotoxemia of neonatal calves caused by Clostridium perfringens (Types A, B, C, D, and E) are reviewed.

Dis Colon Rectum, 1985 Nov, 28(11), 765 - 9
The significance of quantitative results of C . difficile cultures and toxin assays in patients with diarrhea; Church JM et al.; The clinical courses of 114 patients with positive Clostridium difficile cultures or toxin assays performed between 1981 and 1984 were reviewed to determine the relationship between outcome of treatment and quantitative bacteriologic test results . C . difficile culture was positive in 60 of 91 patients while toxin assay was positive in 99 of 114 . One third of the patients received supportive therapy only, and 30 percent of these failed to resolve their symptoms . Ninety-one percent of the patients treated with vancomycin resolved, although 11 percent of these suffered relapse . Patients with high toxin titers receiving supportive treatment alone showed a lower response rate than patients with lower toxin titers . This effect was not seen in patients treated with specific therapy nor with different culture quantities . C . difficile colitis has a range of clinical and microbiologic manifestations . Endoscopy is not always diagnostic, both culture and toxin assays are needed for diagnosis, and toxin titer may help in planning treatment . Patients with low toxin titers may be treated supportively, but high toxin titers are an indication for specific therapy . Quantitative culture results have little diagnostic or therapeutic value.

Appl Environ Microbiol, 1985 Nov, 50(5), 1165 - 70
Effects of butanol on Clostridium acetobutylicum; Bowles LK et al.; The internal pH of Clostridium acetobutylicum was determined at various stages during the growth of the organism . Even in the presence of significant quantities of acetic, butyric, and lactic acids, an internal pH of 6.2 was maintained . Experiments using N,N'-dicyclohexylcarbodiimide indicated that a functioning H+-ATPase is necessary for internal pH control . Butanol, one of the end products of the fermentation, had numerous harmful effects on C . acetobutylicum . At a concentration high enough to inhibit growth, butanol destroyed the ability of the cell to maintain internal pH, lowered the intracellular level of ATP, and inhibited glucose uptake . Experiments done at two different external pH values suggested that the butanol-mediated decrease in ATP concentration was independent of the drop in internal pH . Glucose uptake was not affected by arsenate, suggesting that uptake was not ATP dependent . The effects of butanol on C . acetobutylicum are complex, inhibiting several interrelated membrane processes.

Zh Mikrobiol Epidemiol Immunobiol, 1985 Nov, (11), 37 - 41
{Use of an experimental analytical method for equilibrating nutrient broths for Clostridium perfringens type A growth and toxin formation}; Artemenko VD et al.; A successful attempt to use analytico-experimental approach to the evaluation of experimental data for the scientifically based calculation of the composition of complex culture media, intended for growing pathogenic microorganisms, has been made . The method is based on the evaluation of the specific growth-stimulating and toxin-forming activity of the components of a given culture medium, which are determined by the number of cells grown in the variants of the medium with the limited amount of one of its components . The use of the analytico-experimental balancing method makes it possible to develop culture media with the optimal composition ensuring the definite yield of the target product rather quickly and economically by experimenting on the minimal number of variants equal to the number of the components of the medium . The investigation carried out by means of the analytico-experimental method has revealed that on the basis of peptic serum albumin hydrolysate, pancreatic casein hydrolysate and fodder yeast extract, alongside the culture medium described in an earlier work and containing these components in the proportion 4:2:1, two other media, containing the above components in the proportion 2:4:1 and 3:4:2, can be obtained, these media providing the optimal conditions for, respectively, the toxin formation and growth of C . perfringens, type A.

Hepatology, 1985 Nov-Dec, 5(6), 1126 - 31
Transformation of bile acids into iso-bile acids by Clostridium perfringens: possible transport of 3 beta-hydrogen via the coenzyme; Batta AK et al.; We have examined the mechanism for the bacterial transformation of chenodeoxycholic acid and lithocholic acid into the corresponding 3 beta-hydroxy epimers with the use of 3 alpha- and 3 beta-tritiated bile acids . The 3-oxo bile acids were transformed into the 3 alpha- (85%) and 3 beta- (15%) hydroxy bile acids after 20-hr incubation with Clostridium perfringens . Approximately 75% radioactivity was recovered in the aqueous medium when {3 beta-3H}chenodeoxycholic acid or {3 beta-3H}lithocholic acid was incubated with the bacteria, and approximately 15% of radioactivity in the bile acid fraction was associated with the 3 alpha-position of the iso-bile acids . When {3 beta-3H}chenodeoxycholic acid was incubated with unlabeled 3-oxo-5 beta-cholanoic acid, tritiated litho- and iso-lithocholic acids were recovered . These results can be explained only when a 3-oxo intermediate is postulated, and the 3 beta-hydrogen in the bile acids is transferred by the bacterial coenzyme (NAD+ or NADP+) to the 3 alpha-position in the iso-bile acids during the reduction of the 3-oxo compounds.

Infect Immun, 1985 Nov, 50(2), 442 - 8
Production and characterization of monoclonal antibodies against Clostridium perfringens type A enterotoxin; Wnek AP et al.; Hybridomas secreting monoclonal antibodies (MABs) specific for Clostridium perfringens type A enterotoxin were produced by fusion of P3X63Ag8.653 myeloma cells with spleen cells from BALB/c mice immunized with purified enterotoxin . Wells containing hybridomas secreting immunoglobulin G (IgG) antibodies against enterotoxin were specifically identified by an indirect enzyme-linked immunosorbent assay (ELISA), and 10 ELISA-positive hybridomas were selected and cloned twice by limiting dilution . All 10 hybridomas produced MABs containing immunoglobulin G1 heavy chains and kappa (kappa) light chains . These hybridomas were then grown as ascitic tumors in mice, and MABs were purified from the ascites fluids with DEAE Affi-gel blue . The specificity of the MABs for enterotoxin was demonstrated by immunoblotting and ELISA . Competitive radioimmunoassay with 125I-MABs suggests that these MABs recognized at least four epitopes on the enterotoxin molecule . The enterotoxin-neutralizing ability of MABs from both hybridoma culture supernatants and ascites fluids was assessed by using a 3H-nucleotide-release Vero (African green monkey kidney) cell assay . Only 2 of the 10 hybridomas produced MABs which completely (greater than 90%) neutralized the biologic activity of enterotoxin . Preincubation of 125I-enterotoxin with MABs demonstrated that MAB neutralizing ability correlated with MAB-specific inhibition of specific binding of enterotoxin to intestinal brush border membranes.

J Biol Chem, 1985 Oct 25, 260(24), 13181 - 9
Lactate reduction in Clostridium propionicum . Purification and properties of lactyl-CoA dehydratase; Kuchta RD et al.; Clostridium propionicum converts lactate to propionate (Cardon, B.P., and Barker, H.A . (1947) Arch . Biochem . Biophys . 12, 165-171) . We have obtained a soluble system that carries out this conversion as well as the hydration of acrylate to lactate and the reduction of acrylate to propionate . 3-Pentynyl-CoA inhibits reduction of acrylate and lactate to propionate, but not hydration of acrylate to lactate by cell extracts . The conversion probably involves CoA esters . When {beta-2H3} lactate is used as a substrate, the rate of propionate formation is reduced 1.8-fold, and the methyl group of the resulting propionate has lost 1.4 deuterium atoms . These results are consistent with the intermediate formation of acrylate (acrylyl-CoA) in the conversion of D-lactate to propionate . Two proteins, which we designate E I and E II, were purified to greater than 90% homogeneity . Together, they catalyze the hydration of acrylyl-CoA to lactyl-CoA . E I has an apparent molecular mass of 27,000 daltons and is rapidly and irreversibly inactivated by O2 . E II consists of two subunits of molecular mass 41,000 and 48,000 daltons and contains equal amounts of riboflavin and flavin mononucleotide . Hydration of acrylyl-CoA to lactyl-CoA requires Mg2+ and catalytic quantities of ATP . GTP can replace ATP, but ADP and adenylyl imidodiphosphate cannot . We were unable to detect any stable intermediate during acrylyl-CoA hydration . Finally, we proposed a mechanism for this reaction.

J Immunol Methods, 1985 Oct 24, 83(1), 141 - 50
A double antibody sandwich enzyme-immunoassay for Clostridium perfringens type A enterotoxin detection in stool specimens; Jackson SG et al.; A double antibody sandwich enzyme-immunoassay has been developed for detection of Clostridium perfringens enterotoxin . Anti-enterotoxin immunoglobulin G-alkaline phosphatase conjugates were prepared using a rapid minicolumn procedure . The assay can achieve a sensitivity of greater than or equal to 1 ng/ml with purified enterotoxin . Sensitivity for detection of cases of C . perfringens enteritis in a C . perfringens outbreak (86 individuals tested) was between 85.7 and 98.0 per cent depending upon stringency of criteria for defining positive cases . Specificity of the assay was demonstrated by the lack of positive results in 53 individuals involved in a gastroenteritis outbreak of unknown etiology.

Biochemistry, 1985 Oct 22, 24(22), 6145 - 52
Aldehyde and ketone substrate analogues inhibit the collagenase of Clostridium histolyticum; Grobelny D et al.; The collagenase from Clostridium histolyticum is a mixture of several collagenases, all of which are zinc metalloproteases . This enzyme catalyzes the cleavage of the X-Gly peptide bond in the repeating sequence of collagen: -Gly-Pro-X-Gly-Pro-X- . Thus the S3, S2, and S1 subsites on the enzyme appear to be occupied by the sequence -Gly-Pro-X- and the S1', S2', and S3' subsites also by -Gly-Pro-X- . Short peptides up to and including N alpha-acyltetrapeptides containing the repeat sequence do not detectably inhibit the enzyme (IC50 greater than 10 mM) . However, peptide aldehydes of the form aminoacyl-X-glycinal, presumably occupying the S1, S2, ..., Sn subsites, are inhibitors . The most potent of these was Pro6-Gly-Pro-glycinal, with an IC50 of 340 +/- 70 microM . The single peptide aldehyde investigated, which could occupy the S1' and S2' subsites, 4-oxobutanoyl-L-proline, did not inhibit collagenase (IC50 greater than 20 mM) . The peptide ketone 5-benzamido-4-oxo-6-phenylhexanoyl-Pro-Ala (XXV), which could occupy the S1-S3' subsites, inhibits collagenase with an IC50 of 120 +/- 50 microM, over 80-fold more potently than its parent peptide analogue benzoyl-Phe-Gly-Pro-Ala (XXIII) . The alcohol analogue of XXV, 5-benzamido-4-hydroxy-6-phenylhexanoyl-Pro-Ala (XXVI), is over 60-fold less potent with an IC50 of 8 +/- 2mM . Extending the peptide ketone XXV to occupy the S2-S3' subsites gave 5-(N alpha-carbobenzoxy-L-prolinamido)-4-oxo-6-phenylhexanoyl-Pro -Ala (XXVII) . Surprisingly, XXVII had an IC50 of only 5.2 +/- 2 mM.(ABSTRACT TRUNCATED AT 250 WORDS)

Vet Rec, 1985 Oct 19, 117(16), 408 - 13
Protection of the nursing pig against experimentally induced enteric colibacillosis by vaccination of dam with fimbrial antigens of E coli (K88, K99 and 987P); Nagy LK et al.; Pregnant gilts were vaccinated with two doses of alhydrogel adsorbed fimbrial antigens of Escherichia coli (K88ab, K88ac, K99 and 987P) supplemented with beta toxoid of Clostridium perfringens type C . Their piglets, and piglets of nonvaccinated gilts, were subsequently orogastrically challenged with one or other of the four fimbrial types of enteropathogenic E coli . Some of the vaccinated animals were reinjected with a single dose of the vaccine during second gestation and their piglets, and piglets of non-vaccinated sows, were challenged the same way as were litters of gilts . Blood serum and colostra were examined for antibodies to the four fimbrial antigens of E coli and for antitoxin to beta toxin of C perfringens type C . It was found that: (1) a highly significant reduction in mortality and morbidity was achieved in vaccinated litters against all four challenge strains of E coli; (2) excretion of K88ab and K88ac but not of K99 and 987P challenge strains was significantly reduced; (3) revaccination of sows by a single dose of the vaccine during second gestation conferred complete protection against mortality and highly significant protection against morbidity; (4) no correlation was noted between colostral or seroagglutinins to fimbrial antigens of E coli and mortality rates in litters challenged with homologous fimbrial types of E coli, but good correlation was found between colostral precipitins to K88 antigens and mortality rates in litters; (5) antitoxin value in 97 per cent of colostrum of vaccinated sows was 10 iu equivalent of C perfringens type C toxin or more per ml of colostrum.

Appl Environ Microbiol, 1985 Oct, 50(4), 1043 - 7
Transmembrane pH gradient and membrane potential in Clostridium acetobutylicum during growth under acetogenic and solventogenic conditions; Huang L et al.; The proton motive force and its electrical and chemical components were determined in Clostridium acetobutylicum, grown in a phosphate-limited chemostat, using {14C}dimethyloxazolidinedione and {14C}benzoic acid as transmembrane pH gradient (delta pH) probes and {14C}triphenylmethylphosphonium as a membrane potential (delta psi) indicator . The cells maintained an internal-alkaline pH gradient of approximately 0.2 at pH 6.5 and 1.5 at pH 4.5 . The delta pH was essentially constant between pH 6.5 and 5.5 but increased considerably at lower extracellular pH values down to 4.5 . Hence, the intracellular pH fell from 6.7 to 6.0 as the external pH was lowered from 6.5 to 5.5 but did not decrease further when the external pH was decreased to 4.5 . The transmembrane electrical potential decreased as the external pH decreased . At pH 6.5, delta psi was approximately -90 mV, whereas no negative delta psi was detectable at pH 4.5 . The proton motive force was calculated to be -106 mV at pH 6.5 and -102 mV at pH 4.5 . The ability to maintain a high internal pH at a low extracellular pH suggests that C . acetobutylicum has an efficient deacidification mechanism which expresses itself through the production of neutral solvents.

Eur J Cancer Clin Oncol, 1985 Oct, 21(10), 1159 - 63
Clostridium difficile colitis in leukemia patients; Panichi G et al.; Leukemia patients with diarrhea or other abdominal symptoms have been investigated for the presence of Clostridium difficile and its cytotoxin in stools . Of the patients studied 19% had C . difficile, in most cases together with cytotoxin . All patients but one had received antibiotics, while one had been treated with cytotoxic agents only . Symptoms of colitis were most often abdominal pain and distension rather than diarrhea . Owing to the not infrequent fatal evolution, it is recommended that routine search for C . difficile in leukemia patients with abdominal symptoms be performed and appropriate therapy started immediately.

J Clin Gastroenterol, 1985 Oct, 7(5), 387 - 90
Lack of relationship between Clostridium difficile toxin and inflammatory bowel disease in children; Hyams JS et al.; Conflicting reports have appeared concerning the role of Clostridium difficile toxin in chronic inflammatory bowel disease . Therefore, we prospectively evaluated the incidence of C . difficile toxin in 44 children with inflammatory bowel disease of variable clinical severity over a 1-year period . Only 3/128 stool specimens provided by these patients were found to be toxin-positive . These three stool specimens were from three different patients with Crohn's disease of moderate severity who had no recent hospitalization or antibiotic exposure . None received vancomycin therapy and their stools became toxin-negative over 3 weeks with no apparent change in the patients' clinical condition . No patient with severe disease or recent exposure to antibiotics or sulfasalazine was found to have toxin-positive stools . Routine screening for C . difficile toxin in children with inflammatory bowel disease appears unwarranted.

Eur J Clin Microbiol, 1985 Oct, 4(5), 505 - 7
Identification of Clostridium difficile using the API ZYM system; Levett PN; The use of the API ZYM system for the identification of Clostridium difficile was investigated . The enzyme profiles generated by this system readily distinguished strains of Clostridium difficile from other clostridia commonly isolated from faeces . Enzyme activity of Clostridium difficile was influenced by the composition of the culture medium but appeared to be independent of the age of the culture . Given careful standardisation of techniques the API ZYM system is a suitable alternative to conventional techniques for identification of Clostridium difficile.

Biol Chem Hoppe Seyler, 1985 Oct, 366(10), 953 - 61
Purification and some properties of an acryloyl-CoA reductase of Clostridium kluyveri; Sedlmeier H et al.; Acryloyl-CoA reductase, a presumably previously unknown soluble enzyme, is present in Clostridium kluyveri . It catalyses the reduction of the carbon-carbon double bond of acryloyl-CoA or ethyl vinyl ketone and other alpha, beta-unsaturated carbonyl compounds at the expense of reduced methylviologen . On the basis of a Vmax/Km ratio, which is at least 18 times higher than that for the next best substrate (E)-2-butenoyl-CoA, the enzyme is called acryloyl-CoA reductase . A purity of over 90% was achieved . The apparent molecular mass, as determined by gel chromatography, is 28.4 kDa . Dodecyl sulfate gel electrophoresis shows subunits with a molecular mass of 14.2 kDa . Based on a molecular mass of 28.4 kDa about 1.5 mol FMN have been observed . Less than 0.2 g-atom iron per mol protein were determined . Ferredoxin or flavodoxin seem to be able to carry electrons from hydrogenase to the acryloyl-CoA reductase . The addition of hydrogen to the alpha-carbon of ethyl vinyl ketone occurs from the re-side.

Br J Ophthalmol, 1985 Oct, 69(10), 774 - 7
Clostridium septicum panophthalmitis with systemic complications; Insler MS et al.; A fulminant case of endophthalmitis due to Clostridium septicum is described . The patient presented with spontaneous gas gangrene panophthalmitis, with early visual loss and an air bubble in the anterior chamber . Death ensued, and necropsy revealed changes consistent with severe arterosclerotic cardiovascular disease, a relationship not uncommon in patients with clostridium sepsis . This association as well as the histopathology of the globe are discussed.

Cancer Res, 1985 Oct, 45(10), 4986 - 9
Effects of the antitumor drug Adriamycin on human red blood cell discocyte-echinocyte transitions; Chahwala SB et al.; The antitumor drug Adriamycin, when preincubated with human red blood cells (discocytes) for 10 min, prevented the formation of echinocytes induced by the calcium ionophore A23187 in the presence of 0.2 mM calcium . The degree of protection was concentration dependent and was greater than 90% at 10 microM Adriamycin . Adriamycin did not interfere with the accumulation of calcium induced by a 5 microM concentration of the ionophore . Adriamycin reversed echinocyte morphology to the discocyte form in echinocytes which had been formed by adenosine triphosphate depletion but not those formed after treatment with A23187 and Ca2+ . Its ability to protect against Ca2+-induced echinocyte formation contrasts with the failure of the local anesthetic procaine to exert such an effect, even at 45 mM (J . Palek et al., Blood, 50: 155-164, 1977), and this difference suggests that Adriamycin may not be acting simply as a chaotropic agent . This hypothesis was supported by the observation that Adriamycin alone did not induce a cup-form morphology in discocytes (stomatocytosis) . Wheat germ agglutinin protection of echinocyte formation induced by calcium loading was reversed by 30 mM N-acetylglucosamine, which partially reversed the Adriamycin protection of echinocyte formation . However, desialylation of human red blood cells with Clostridium perfringens type V neuraminidase, while preventing the protection of echinocyte formation by wheat germ agglutinin, had no effect on the protection afforded by Adriamycin . This suggests that Adriamycin does not prevent echinocyte formation via binding to the sialic acid residues of the transmembrane protein glycophorin and that another mechanism or mechanisms are involved in its action to modulate morphological transitions of the red blood cell membrane.

Appl Environ Microbiol, 1985 Oct, 50(4), 795 - 800
Germination of spores from Clostridium botulinum B-aphis and Ba410; Montville TJ et al.; The germination of spores from Clostridium botulinum B-aphis and Ba410 was examined . In a complex medium, heat activation of spores from both strains doubled the germination rates and was required for germination in the presence of 2% NaCl . In a defined medium (CTB {D . B . Rowley and F . Feeherry, J . Bacteriol . 104:1151-1157, 1970}), the parent strain B-aphis germinated at a rate of 0.77% min-1 in the absence of NaCl and was not affected by 2% NaCl . A salt-tolerant derivative, strain Ba410, germinated at rates of 0.16% min-1 in CTB and 0.04% min-1 in CTB containing 2% NaCl . L-Alanine-triggered spores germinated faster than did L-cysteine-triggered spores from both strains . When both amino acids were present, B-aphis germinated rapidly in the absence of NaCl and had biphasic kinetics in the presence of NaCl . Strain Ba410 had biphasic kinetics in the absence of NaCl and germinated slowly with single-phase kinetics in the presence of NaCl . L-Alanine- and L-cysteine-triggered germinations were each inhibited by both D-alanine and D-cysteine, indicating a common germinant-binding site for both alanine and cysteine . Attempts to select for variants with amino acid-specific germinant-binding sites were unsuccessful . Differences in the germination kinetics of both strains could not be explained by ultrastructural differences . Transmission electron micrographs revealed striking similarities between the strains.

J Antibiot (Tokyo), 1985 Oct, 38(10), 1322 - 6
Luminamicin, a new antibiotic . Production, isolation and physico-chemical and biological properties; Omura S et al.; A new antibiotic, luminamicin, was isolated from the culture broth of an actinomycete strain OMR-59 . It exhibits antibacterial activity against anaerobic bacteria, especially against Clostridium sp . The molecular formula of the antibiotic was determined as C32H38O12 on the basis of high resolution mass spectrum, elemental analysis and NMR spectrum.

Proc Natl Acad Sci U S A, 1985 Oct, 82(20), 6811 - 4
Evidence that an iron-nickel-carbon complex is formed by reaction of CO with the CO dehydrogenase from Clostridium thermoaceticum; Ragsdale SW et al.; The interaction between carbon monoxide and the CO dehydrogenase from Clostridium thermoaceticum was studied by electron spin resonance (ESR) techniques . When the enzyme reacts with CO, a paramagnetic complex is formed which previously was shown, by isotope substitution, to be due to a nickel-carbon species . In this paper, we demonstrate that iron is also a component of this ESR-detectable complex . When the iron in the enzyme is replaced with 57Fe, a broadening of 18 G in the g parallel and 7 G in the g perpendicular region is seen . This hyperfine interaction is probably due to more than one iron atom in the complex . Coenzyme A influences this ESR spectrum . In the absence of CoA, the ESR spectrum consists of two superimposed signals, which were simulated using the following ESR parameters: signal 1, with g = 2.074 and g = 2.028, and signal 2 with gx = 2.062, gy = 2.047, and gz = 2.028 . CoA converts signal 2 into signal 1 . Since iron, nickel, and carbon all are part of this ESR-detectable complex, we propose that these atoms exist in a spin-coupled complex with net spin = 1/2, analogous to other iron-sulfur centers in which the metals are bridged by acid-labile sulfide.

Zh Mikrobiol Epidemiol Immunobiol, 1985 Oct, (10), 22 - 5
{Differentiation of the main species of Clostridium by gas chromatography}; Bychenko BD et al.; For the first time in the USSR the properties of microorganisms of the genus Clostridium have been studied with the use of the gas-chromatographic techniques . The analysis of the quantitative and qualitative composition of extracellular alcohols and carboxylic acids in 99 museum and newly isolated strains of 18 Clostridium species has made it possible to classify these microorganisms with 7 sharply differing groups . The above techniques permit the classification of clostridia with one of the groups within 2 hours if the microbial cultures have been grown in glucose-containing peptone yeast medium.

Appl Environ Microbiol, 1985 Oct, 50(4), 1110 - 1
Selective and differential medium for detecting Clostridium botulinum; Silas JC et al.; A selective and differential growth medium was developed for detection of Clostridium botulinum types A, B, and F . The medium consisted of peptone-glucose-yeast extract agar supplemented with cycloserine, 250 micrograms/ml; sulfamethoxazole, 76 micrograms/ml; and trimethoprim, 4 micrograms/ml as selective inhibitors and various types and levels of botulinal antibodies for type differentiation in the immunodiffusion reaction . Growth of proteolytic types of C . botulinum were not affected by the incorporation of the selective agents, but some nonproteolytic types were suppressed . Cross-reactions between types A and B were visually distinguishable, whereas cross-reactions between type F and Clostridium sporogenes did not occur at the optimum antibody titer . Optimum antibody titer varied with toxin type . The proposed selective differential medium should be valuable in isolating and typing of proteolytic C . botulinum types A, B, and F from samples containing mixed microbial populations.

Appl Environ Microbiol, 1985 Oct, 50(4), 1097 - 9
Protoplast formation and cell wall regeneration in Clostridium perfringens; Stal MH et al.; A protocol was developed for the efficient production and regeneration of Clostridium perfringens protoplasts . Cell wall regeneration frequencies of up to 5% were obtained.

Am J Vet Res, 1985 Oct, 46(10), 2147 - 8
Enteropathogenicity of purified Clostridium perfringens enterotoxin in the pig; Popoff MR et al.; In an assay procedure, purified Clostridium perfringens enterotoxin induced the accumulation of fluid in the ileal loop of the axenic pig . The smallest amount of enterotoxin causing a positive response was 25 micrograms (1:32 titer by counterimmunoelectrophoresis {CIEP} ), and 100 micrograms (1:128 CIEP titer) caused a marked response . Possibly, diarrhea in pigs with 1:32 CIEP titer or more of enterotoxin in the feces may be associated with enterotoxigenic C perfringens.

Biochemistry, 1985 Sep 24, 24(20), 5647 - 52
Kinetics of reduction of high redox potential ferredoxins by the semiquinones of Clostridium pasteurianum flavodoxin and exogenous flavin mononucleotide . Electrostatic and redox potential effects; Przysiecki CT et al.; We have measured the ionic strength dependence of the rate constants for the electron-transfer reactions of flavin mononucleotide (FMN) and flavodoxin semiquinones with 10 high redox potential ferredoxins (HiPIP's) . The rate constants were extrapolated to infinite ionic strength by using a theoretical model of electrostatic interactions developed in our laboratory . In all cases, the sign of the electrostatic interaction was the same as the protein net charge, but the magnitudes were much smaller . The results are consistent with a model in which the electrical charges are approximately uniformly distributed over the HiPIP surface and in which there are both short- and long-range electrostatic interactions . An electrostatic field calculation for Chromatium vinosum HiPIP is consistent with this . The presumed site of electron transfer includes that region of the protein surface to which the iron-sulfur cluster is nearest and appears to be relatively hydrophobic . The principal short-range electrostatic interaction would involve the negative charge on the iron-sulfur cluster . For some net negatively charged proteins, this effect is magnified, and for net positively charged HiPIP's, it is counterbalanced . The rate constants extrapolated to infinite ionic strength can be correlated with redox potential differences between the reactants, as has previously been shown for cytochrome-flavin semiquinone reactions . Both electrostatic and redox potential effects are magnified for the flavodoxin semiquinone as compared to the FMN semiquinone-HiPIP reactions . This was also observed previously for the flavin semiquinone-cytochrome reactions.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochim Biophys Acta, 1985 Sep 20, 831(1), 159 - 60
Evidence for a pH-dependent isomerization of Clostridium oroticum dihydroorotase; Bidigare RR et al.; Analytical gel permeation chromatography on both Sephadex and polyacrylamide columns shows that Clostridium oroticum dihydroorotase (L-5,6-dihydroorotate amidohydrolase, EC 3.5.2.3) undergoes a large decrease in molecular size when the pH is decreased from 8 to 6 . The Stokes radius decreases from about 40 A to 36 A . Neither the molecular size nor kinetic properties are dependent on protein concentration . Thus, the decreased molecular size reflects a pH dependent isomerization of the enzyme.

Biochim Biophys Acta, 1985 Sep 11, 836(2), 255 - 61
Characterization of delta 4-3-ketosteroid-5 beta-reductase and 3 beta-hydroxysteroid dehydrogenase in cell extracts of Clostridium innocuum; Stokes NA et al.; Cell extracts prepared anaerobically from Clostridium innocuum and Clostridium paraputrificum reduced delta 4-3-ketosteroids to 3 beta 5 beta and 3 alpha 5 beta derivatives, respectively . delta 4-3-Ketosteroid-5 beta-reductase (5 beta-reductase) from both organisms required NADH for activity . 5 beta-Reductase from C . innocuum had a pH optimum of 5.0 . The substrate concentration at half-maximal reaction velocity was 4.2 microM, and a specific activity of 17 nmol product formed/h per mg protein was determined using 4-pregnen-3,20-dione (progesterone) as a substrate . delta 4-3-Ketosteroid-5 beta-reductase from C . innocuum reduced progesterone and testosterone, but not 4-cholesten-3-one, to corresponding 3-keto-5 beta derivatives . A relative molecular (Mr) weight of 80 000 was estimated for 5 beta-reductase using HPLC-gel filtration chromatography . 3 beta-Hydroxysteroid dehydrogenase in cell extracts of C . innocuum was oxygen sensitive and required NADH for activity . An Mr of 80 000 was estimated for 3 beta-hydroxysteroid dehydrogenase . However, 5 beta-reductase and 3 beta-hydroxysteroid dehydrogenase activities were separated using an HPLC-DEAE chromatography technique.

Biochim Biophys Acta, 1985 Sep 6, 841(3), 306 - 17
Amidation of, and (R)-1-amino-2-propanol attachment to, the corrin ring during vitamin B-12 biosynthesis by Clostridium tetanomorphum extracts; Ford SH; Two intermediate stages in cobalamin biosynthesis, amidation of carboxylic acid groups in the corrin ring and (R)-1-amino-2-propanol attachment at propionic acid position f, have been studied using cell-free extracts from the obligate anaerobe Clostridium tetanomorphum . The preparation of an incomplete corrinoid, probably cobinic acid-a,c,d,e,g-pentaamide, as an in vitro amidation substrate was accomplished via mild acid hydrolysis of cobinamide . Weak, but reproducible activities for both amidation and (R)-1-amino-2-propanol attachment were found in crude, nucleic acid-free and DE-52 column-purified protein fractions . The amidation reaction was glutamine-dependent in crude fractions, but became ammonium ion-dependent in more purified fractions . Significant problems encountered were (a) the weak and unstable character of both enzyme activities, and (b) the irreversible changes in the visible spectra of the incomplete corrinoids employed as substrates caused by use of thiol-reducing agents in the buffers and assays.

J Biol Chem, 1985 Sep 5, 260(19), 10461 - 6
Separation, purification, partial characterization and comparison of the heavy and light chains of botulinum neurotoxin types A, B, and E; Sathyamoorthy V et al.; Clostridium botulinum produces botulinum neurotoxin (NT) in antigenically distinct forms . When isolated from bacterial cultures type E is a single chain, type B is a mixture of single and two-chain molecules, and type A is essentially a two-chain molecule (Mr approximately 150,000) . Protease(s) in the cultures or trypsin nick single-chain NT to the two-chain form . The heavy (Mr approximately 100,000) and light (Mr approximately 50,000) chains of the two-chain molecule remain held together by -S-S-bond(s) . The two chains are presumed to have different functions . NT binds to nerve cells via the heavy chain and then light chain enters the cell and blocks release of acetylcholine (Simpson, L . L . (1981) Pharmacol . Rev . 33, 155-188) . We nicked single-chain NT to form the two-chain form with trypsin, minimizing secondary cleavages, then separated and purified the heavy and light chains using ion-exchange chromatography . The technique, with minor modifications, is a generalized method for types A, B, and E . These subunit chains (each a single band in sodium dodecyl sulfatepolyacrylamide gel electrophoresis) were analyzed for their complete amino acid compositions . The amino acid contents of the heavy and light chains agreed well with the parent two-chain molecule . This affirms that NT is composed of two chains . The two subunit chains are now usable for amino acid sequence and other studies . Comparison of the amino acid contents indicates more similarity among the light chains than the heavy chains of the three NT types, a similarity that agrees with our published partial amino acid sequences (first 13-18 residues) of these chains . Several (up to 9) different amino acid residues of the heavy chain (which is twice the size of the light chain) are present in double the number of corresponding residues in the light chain.

Arch Microbiol, 1985 Sep, 142(4), 375 - 82
Metabolic and energetic aspects of the growth of Clostridium butyricum on glucose in chemostat culture; Crabbendam PM et al.; The influence of a number of environmental parameters on the fermentation of glucose, and on the energetics of growth of Clostridium butyricum in chemostat culture, have been studied . With cultures that were continuously sparged with nitrogen gas, glucose was fermented primarily to acetate and butyrate with a fixed stoichiometry . Thus, irrespective of the growth rate, input glucose concentration, specific nutrient limitation and, within limits, the culture pH value, the acetate/butyrate molar ratio in the culture extracellular fluids was uniformly 0.74 +/- 0.07 . Thus, the efficiency with which ATP was generated from glucose catabolism also was constant at 3.27 +/- 0.02 mol ATP/mol glucose fermented . However, the rate of glucose fermentation at a fixed growth rate, and hence the rate of ATP generation, varied markedly under some conditions, leading to changes in the Y glucose and YATP values . In general, glucose-sufficient cultures expressed lower yield values than a corresponding glucose-limited culture, and this was particularly marked with a potassium-limited culture . However, with a glucose-limited culture increasing the input glucose concentration above 40 g glucose X 1(-1) also led to a significant decrease in the yield values that could be partially reversed by increasing the sparging rate of the nitrogen gas . Finally glucose-limited cultures immediately expressed an increased rate of glucose fermentation when relieved of their growth limitation . Since the rate of cell synthesis did not increase instantaneously, again the yield values with respect to glucose consumed and ATP generated transiently decreased . Two conditions were found to effect a change in the fermentation pattern with a lowering of the acetate/butyrate molar ratio . First, a significant decrease in this ratio was observed when a glucose-limited culture was not sparged with nitrogen gas; and second, a substantial (and progressive) decrease was observed to follow addition of increasing amounts of mannitol to a glucose-limited culture . In both cases, however, there was no apparent change in the YATP value.(ABSTRACT TRUNCATED AT 250 WORDS)

Leber Magen Darm, 1985 Sep, 15(5), 192 - 7
{Disorders of intestinal flora in intensive care patients}; Graninger W; The intestinal flora under normal conditions prevents colonisation of the intestinal mucosa with pathogenic bacteria . Various diseases as well as antibiotics may disturb the host/bacteria balance . If patients are in addition immunocompromised, otherwise commensal bacteria may cause life threatening infections . Treatment of intensive care patients with antibiotics thus should account for preservation of resistance against colonization . Antibiotics active against anaerobes or poorly absorbed from the gastrointestinal tract, or excreted in the bile should be avoided . In patients with colitis induced by antibiotics the number of clostridium difficile with subsequent toxin production are greatly increased as a consequence of the killing of the normal anaerobic colon bacterial flora; in these patients vancomycin has to be applied . Mostly "dysbiosis" caused by antibiotics does not need any treatment . Therapeutic adjuncts like the administration of bacterial preparations e.g . lactobacilli are of no value.

Can J Surg, 1985 Sep, 28(5), 432 - 3
Pseudomembranous colitis and wound infection following perioperative use of multiple antibiotics; Bohnen JM et al.; The prophylactic use of antibiotics in elective surgery of the colon is accepted practice, but it has inherent risks . The authors report the case of a 70-year-old woman who had wound infection and severe, relapsing pseudomembranous colitis due to Clostridium difficile after a short course of antibiotics given orally and parenterally at the time of elective resection of the colon . Perioperatively, she received erythromycin base and neomycin orally, plus netilmicin and metronidazole intravenously . Although the concomitant administration of parenteral antibiotics may enhance the benefit of antibiotics given orally before operation, this does not entirely prevent wound infection . Until the relation between the number of drugs and risk of antibiotic-associated colitis is more clearly defined, caution should be exercised in the use of multiple antibiotics in elective colonic surgery.

J Antimicrob Chemother, 1985 Sep, 16(3), 305 - 13
The in-vitro activity of a novel penem FCE 22101 compared to other beta-lactam antibiotics; Neu HC et al.; FCE 22101 is a penem antibiotic which inhibits the majority of Enterobacteriaceae, Haemophilus influenzae, and Neisseria gonorrhoeae at concentrations of 0.5-4 mg/l . It inhibits staphylococci, haemolytic streptococci and Streptococcus pneumoniae at less than or equal to 0.25 mg/l . Pseudomonas aeruginosa and other Pseudomonas species are resistant . Bacteroides fragilis and Clostridium species are inhibited by less than or equal to 1 mg/l . FCE 22101 is not hydrolyzed by the common plasmid and chrosmosomal beta-lactamases . It shows minimal discrepancy between MIC and MBC values and there is minimal effect of inoculum size . Although FCE 22101 is generally less active against Enterobacteriaceae than are cefotaxime and ceftazidime, it does inhibit some Enterobacter spp . resistant to these agents . FCE 22101 and imipenem are similar in activity against Gram-positive and anaerobic species.

Vet Res Commun, 1985 Sep, 9(4), 269 - 87
Etiology and pathogenesis of necrotic enteritis; Shane SM et al.; Sporulated oocysts of Eimeria acervulina were administered orally to cage-housed broilers at a dose of 3.5 X 10(5) resulted in mild subclinical coccidiosis . Clostridium perfringens incorporated in feed at a level of 2.5 X 10(8) organisms/g . produced lesions characteristic of necrotic enteritis . Mortality of 8% (7/80) occurred in birds fed a ration inoculated with Cl . perfringens alone . Mortality of 35% (28/80) was observed in birds which received an oral dose of E . acervulina and which were fed simultaneously with a ration containing Cl . perfringens . Birds which were fed an inoculated ration two days after an oral dose of E . acervulina showed 41% (33/80) mortality . Birds which received an inoculated ration for two days before administration of an oral dose of E . acervulina demonstrated 18% mortality (15/80) . Birds which were fed an inoculated ration four days after an oral dose of E . acervulina showed 10% mortality . Infection with E . acervulina reduced the pH of intestinal contents with a simultaneous depression in serum protein . A 39% increase in intestinal passage time from 178 to 248 minutes occurred on the fifth day after infection with E . acervulina . These experiments suggest that necrotic enteritis, attributed to proliferation of a toxigenic strain of Cl . perfringens, followed intestinal stasis and minimal lesions induced by mild intestinal coccidiosis.

J Hosp Infect, 1985 Sep, 6(3), 312 - 22
A hospital outbreak of Clostridium difficile?
Hall SM, Calver GP, Williams M.
An increase in numbers of patients with Clostridium difficile and its toxin in their stools at a hospital in South-west London led to closure of a ward to admissions and to an investigation of a possible nosocomial outbreak . The findings suggested that the increase was not due to an outbreak of related cases but to increased investigation . The cost of the episode both in financial terms and in the effect on patient care, was considerable . This study highlights the need for caution in interpreting the significance of Cl . difficile in stool specimens . Laboratory data can only alert clinicians to the possibility of colitis; decisions about treatment and control of spread of infection should also be based on clinical criteria.

J Assoc Off Anal Chem, 1985 Sep-Oct, 68(5), 881 - 3
Rapid detection of Clostridium perfringens: comparison of lactose sulfite broth with tryptose-sulfite-cycloserine agar; Neut C et al.; The lactose sulfite (LS) medium recommended for the detection and identification of Clostridium perfringens in foods was compared with a reference method using tryptose-sulfite-cycloserine (TSC) agar for the enumeration of this organism in a variety of foods and food ingredients . C . perfringens was detected and enumerated in 17 of the 54 samples examined with LS broth, but its presence could be confirmed in only 9 of the samples with TSC agar . In only 2 instances, C . perfringens was detected on TSC agar but not in LS broth . A positive response (FeS + and gas +) in LS broth incubated at 46 degrees C always corresponded to the presence of C . perfringens; whereas the black colonies formed on TSC agar incubated at 37 degrees C were frequently found to be Clostridium species other than C . perfringens . Thus, because of its highly selective nature, LS broth was superior to TSC agar for enumerating and confirming the small numbers of C . perfringens that were present in a majority of the samples . This was especially true when other clostridia were also present . Besides its greater selectivity and sensitivity, LS broth had the additional advantages of requiring less work and giving confirmed results within 24-48 h compared with 3 days for the TSC agar method.

Biochem J, 1985 Sep 1, 230(2), 451 - 5
A study on the mechanism of the epimerization at C-3 of chenodeoxycholic acid by Clostridium perfringens; Aragozzini F et al.; The mechanism of 3-hydroxy epimerization of chenodeoxycholic acid by Clostridium perfringens was investigated in 3 alpha, 7 alpha-dihydroxy-{2,2,4,4-2H4}-, 3 alpha, 7 alpha-dihydroxy-{3 beta-2H}- and 3 beta, 7 alpha-dihydroxy-{3 alpha-2H}-5 beta-cholanoic acid transformations . Our findings rule out a dehydration-rehydration pathway and agree with a redox mechanism involving 3-oxochenodeoxycholic acid as intermediate.

Age Ageing, 1985 Sep, 14(5), 296 - 302
Diarrhoea due to enterotoxigenic Clostridium perfringens: clinical features and management of a cluster of ten cases; Williams R et al.; Clostridium perfringens has recently been shown to be associated with antibiotic-associated diarrhoea . We describe here the clinical features and management of an outbreak of diarrhoea in a Geriatric Unit . Ten cases were due to enterotoxigenic C . perfringens and in these cases there was a highly significant correlation with recent antibiotic administration (P = 0.0001) . The importance of early recognition of C . perfringens as a cause of infective diarrhoea in the elderly is stressed.

Eur J Biochem, 1985 Aug 15, 151(1), 75 - 82
Inactivation of Clostridium botulinum type A neurotoxin by trypsin and purification of two tryptic fragments . Proteolytic action near the COOH-terminus of the heavy subunit destroys toxin-binding activity; Shone CC et al.; Limited treatment of Clostridium botulinum type A neurotoxin with trypsin resulted in the cleavage of the heavy (95000 Da) subunit at approximately the mid-position and a loss of toxic activity . The rate of toxicity loss was considerably faster than that of mid-chain cleavage; thus a loss of toxicity in excess of 90% was accompanied by only 30-35% mid-chain cleavage of the heavy subunit . A study of the binding of 125I-labelled neurotoxin to rat brain synaptosomes showed the loss of toxicity on trypsin treatment to be paralleled by a loss of toxin binding to rat brain synaptosomes suggesting the presence of at least two sites of tryptic action on the 95000-Da binding subunit . Prolonged treatment of the neurotoxin with trypsin resulted in the complete digestion of a 46000-Da fragment of the heavy subunit, leaving intact a soluble fragment of approximately 105000 Da containing the light subunit linked to the remaining (49000-Da) portion of the heavy subunit . This fragment exhibited less than 0.01% of the original toxicity and gave immunoprecipitation reactions indistinguishable from the native toxin . The 49000-Da portion of the heavy chain was purified from the 105000-Da fragment of the toxin and the sequence of the first 35 amino acids determined . The sequence of the first 10 residues was found to be identical to that previously reported for the heavy subunit showing that the 49000-Da fragment represents the NH2-terminal portion of the heavy chain and that this region is resistant to tryptic action . It is suggested that the primary site(s) of tryptic action on the heavy subunit of botulinum type A neurotoxin is close to the COOH terminus and that cleavage of the polypeptide chain in this region results in a loss of toxic activity mediated by the destruction of the neurotoxin-binding site.

Biochem J, 1985 Aug 15, 230(1), 101 - 8
Dihydro-orotase from Clostridium oroticum . Purification and reversible removal of essential zinc; Pettigrew DW et al.; A new purification procedure involving five column-chromatography steps is described for dihydro-orotase (L-5,6-dihydro-orotate amidohydrolase, EC 3.5.2.3) from Clostridium oroticum (A.T.C.C . 25750) . The native purified enzyme is a dimer of Mr 102 000 and contains 4.0 +/- 0.3 g-atoms of zinc/mol of dimer . These observations agree with those reported previously {Taylor, Taylor, Balch & Gilchrist (1976) J . Bacteriol . 127, 863-873} . It is conclusively demonstrated that dihydro-orotase is a zinc metalloenzyme . Zinc is reversibly removed by treatment with chelators in phosphate buffer at pH 6.5, as demonstrated by atomic absorption spectrophotometry and decrease of enzyme activity . The specific activity is linearly dependent on zinc content . Addition of ZnSO4 to the chelator-treated enzyme results in regain of the normal complement of zinc and enzyme activity . Kinetic properties of the reconstituted enzyme are indistinguishable from those of the native enzyme . The amino acid composition of the homogeneous enzyme suggests that the zinc atoms occupy different environments.

Angew Parasitol, 1985 Aug, 26(3), 151 - 5
Connections between Ascaridia galli and the bacterial flora in the intestine of hens; Okulewicz A et al.; Parasitological dissections of 502 intestinal tracts of hens deriving from big private chicken-farms have been done . In the jejunum of 146 hosts (ext . 29.1%) from 1 to 21 individuals of A . galli were detected . Using bacterial selective media and biochemical tests, the microorganisms from the hen's intestinal tracts as well as from the cuticle surface of the nematodes were identified . Among them were: grampositive (+) Lactobacillus, Bacillus, Staphylococcus, Streptococcus, Micrococcus, Sarcina, Clostridium, Corynebacterium; gramnegative (-) Enterobacteriaceae, Pseudomonas, Pasteurella, and fungi Candida and others . The lower frequency of microorganisms and the smaller amount of bacteria in the intestinal content in infected hens than in uninfected show that A . galli has antibacterial properties.

J Pediatr Gastroenterol Nutr, 1985 Aug, 4(4), 563 - 7
Differential effect of botulinal toxin on esophageal motor function in infants; Cannon RA; The effect of Clostridium botulinum toxin on esophageal motor function was studied in four infants (ages 5-9 months) with confirmed infant botulism . Esophageal motility studies using a perfused catheter assembly were performed during the acute phase in all patients, and during the recovery phase in one patient . Motor function of the proximal esophagus and upper esophageal sphincter was abnormal in each, while motor function of the distal esophagus and lower esophageal sphincter was normal . Mean lower esophageal sphincter pressure was 24 mm Hg for the group (normal: 15-30 mm Hg) . Sequential studies of proximal esophageal motility in one infant revealed a return of normal motor function which correlated with recovery of peripheral muscle strength and gag reflexes . C . botulinum toxin impairs proximal, but not distal, esophageal motor function in the infant botulism model . This effect appears to be consistent with the known action of the toxin on synaptic acetylcholine release and current concepts regarding distribution of cholinergic and noncholinergic neurotransmitter receptors in the esophagus.

Arch Otolaryngol, 1985 Aug, 111(8), 550 - 3
Clostridium difficile colitis following head and neck surgery . Report of cases; Griebie M et al.; Clostridium difficile, a toxin-producing, gram-positive anaerobe, has been implicated as the causative agent of pseudomembranous colitis, an acute inflammatory bowel disease that generally occurs in association with antimicrobial therapy . This subject has received extensive review in the general surgical, medical, and pediatric literature but has not been specifically addressed in the literature of our specialty . We present the report of four recent cases, including one that progressed to the clinical picture of peritonitis and toxin megacolon . The literature is reviewed regarding presentation, diagnosis, and treatment of C difficile colitis . Familiarity with this disease process may minimize morbidity and prevent disastrous complications following major head and neck surgery.

Am J Med, 1985 Aug, 79(2), 256 - 8
Clostridium septicum septicemia with identical metastatic myonecroses in a granulocytopenic patient . Infectious disease emergency; Tikko SK et al.; Clostridium septicum is a gram-positive, sporulating spindle-shaped rod . Gas gangrene secondary to trauma is not uncommon . However, nontraumatic clostridial infection causing myonecrosis is quite unusual . This is a unique case report of Clostridium septicum bacteremia with two simultaneously evolving metastatic foci of myonecrosis of the left arm and right thigh that developed in a patient with lymphoma when he became granulocytopenic during his hospital course.

J Med Microbiol, 1985 Aug, 20(1), 17 - 26
The antagonism of tetracycline and ferric iron in vivo; Miles AA et al.; To test the hypothesis that the in-vivo antibiotic action of tetracycline might be affected by ferric iron and the enhancement of infection by ferric iron by tetracycline, the actions of intraperitoneal antibiotic and local ferric ammonium citrate, given separately and together, were measured in the dorsal skin of guinea-pigs bearing lesions due to staphylococci, streptococci, a Proteus sp., an Erysipelothrix sp., Clostridium perfringens, Pseudomonas aeruginosa, Aeromonas hydrophila and Klebsiella pneumoniae . Tetracycline, given in two intraperitoneal doses of 25 mg/kg at 0 and 2 h after intracutaneous challenge, maintained plasma concentrations of 4-6 micrograms/ml for more than the first 4 h of infection, after which the local lesions had become largely insusceptible to the antibiotic . The intracutaneous injection of Fe 10 micrograms in a volume of 0.1 ml containing the bacteria was sufficient to enhance infection by those strains susceptible to this effect . The in-vivo efficacy of tetracycline was not always related to low MIC; a low MIC was sometimes associated with little action and a high MIC with moderate action . Sixteen organisms were tested . The iron diminished the tetracycline effect only feebly with one staphylococcal strain and the strain of E . rhusiopathiae . In only one case, with a strain of Proteus sp., was the tetracycline action grossly diminished . On the other hand, tetracycline diminished the enhancement effect of iron moderately with three strains of staphylococci and one strain each of K . pneumoniae, P . aeruginosa and C . perfringens, and strongly with two strains of staphylococci, a group-C streptococcus and one strain each of K . pneumoniae, E . rhusiopathiae and A . hydrophila . It is evident that the diminution of tetracycline action by moderate excess of readily available Fe , whether endogenous or administered, is an unlikely event (three instances among the 16 tested) whereas the diminution of the infection-enhancing effect of iron by tetracycline is much more likely (12 instances among the 16) . Insofar as a decrease in iron available for enhancement of infection is valid evidence of a diminution of the iron available for necessary physiological processes of the subject treated, our results suggest that these processes might be affected by tetracycline.

J Bacteriol, 1985 Aug, 163(2), 552 - 9
Organization and distribution of the cellulosome in Clostridium thermocellum; Bayer EA et al.; The properties of the cellulosome (the cellulose-binding, multicellulase-containing protein complex) in Clostridium thermocellum were examined by comparing the cellulase systems derived from the wild type and an adherence-defective mutant . The growth conditions--specifically, growth either on cellulose (Avicel) or on cellobiose as insoluble or soluble carbon sources, respectively--were found to be critical to the distribution of the cellulosome in the mutant system: the cellobiose-grown mutant (in contrast to the wild type) lacked the cellulosome on its surface and produced only minor quantities of the extracellular cellulosome accompanied by other relatively low-molecular-weight cellulases . The polypeptide composition of the respective purified cellulosome was dependent on the nature of the carbon source and was similar for both wild-type and mutant cells . Ultrastructural analysis revealed the presence of novel polycellulosomal protuberances on the cell surface of the cellobiose-grown wild type which were absent in the mutant.

Appl Environ Microbiol, 1985 Aug, 50(2), 274 - 9
Inhibition of germinant binding by bacterial spores in acidic environments; Blocher JC et al.; Commitment to germinate occurred in both Clostridium botulinum and Bacillus cereus spores during 0.5 min of exposure to 100 mM L-alanine or L-cysteine, measured by the inability of germination inhibitors (D form of amino acid) to inhibit germination . Spore germination at pH 4.5 was inhibited because the germinant did not bind to the trigger sites . C . botulinum spores exposed to 100 mM L-alanine or L-cysteine at pH 4.5 remained sensitive to D-amino acid inhibition at pH 7, indicating that no germinants had bound to the trigger site at pH 4.5 . Inhibition of germinant binding at pH 4.5 was reversible but lagged in commitment to germinate upon transfer to pH 7 . Spores sequentially exposed to pH 4.5 buffer and pH 7 buffer with the germinant also demonstrated a lag in commitment to germinate . The pH at which binding was inhibited was not significantly affected by composition of the buffer or by reduced germinant concentrations (10 mM) . Nonspecific uptake of L-{3H}alanine by C . botulinum spores was not inhibited at pH 4.5 . Inhibition of germinant binding in acidic environments appeared to be due to protonation of a functional group in or near the trigger site . This may represent a general mechanism for inhibition of spore germination in acidic environments.

Infect Immun, 1985 Aug, 49(2), 452 - 4
Kinetics of growth and toxigenicity of Clostridium botulinum in experimental wound botulism; Dezfulian M et al.; An animal model of wound botulism was developed in mice using an inoculum of Clostridium botulinum type A spores . The number of C . botulinum in infected wounds was quantitated by culturing on egg yolk agar, and the level of C . botulinum toxin in infected wound tissue was measured by a bioassay in mice and by an enzyme-linked immunosorbent assay . All infected mice receiving no further treatment developed neuroparalytic symptoms consistent with botulism after an incubation period of ca . 48 h, and all of these animals died . Serotherapy with C . botulinum type A antitoxin initiated 24 h postchallenge reduced the mortality rate to 5% . Treatment with metronidazole 2 to 24 h postchallenge resulted in recovery rates of 40 to 91%.

J Gen Microbiol, 1985 Aug, 131 ( Pt 8), 2097 - 105
Identification of restriction fragments from two cryptic Clostridium butyricum plasmids that promote the establishment of a replication-defective plasmid in Bacillus subtilis; Collins ME et al.; Clostridium butyricum NCIB 7423 carries two cryptic plasmids, pCB101 (6.05 kbp) and pCB102 (7.8 kbp) . Sites for the restriction enzymes EcoRI, EcoRV, HindIII, ClaI and PstI have been found in one or both of these plasmids and their relative positions determined . Restriction fragments from both plasmids have been inserted into a vector plasmid (pJAB1) that is able to replicate in Escherichia coli but not in Bacillus subtilis and the recombinant plasmids have been established in E . coli . A 3.3 kbp Sau3A fragment of pCB101 conferred upon the vector the ability to transform both Rec+ and Rec- strains of B . subtilis . Plasmid pRB1, a representative chimaera carrying only the 3.3 kbp Sau3A fragment of pCB101, was successfully transferred from B . subtilis back to E . coli . Plasmid pRB1 was readily lost from B . subtilis in the absence of selection . This evidence, together with the results of hybridization experiments, suggests that pRB1 is present as a weakly replicating autonomous element in B . subtilis . A recombinant plasmid carrying a 2.0 kbp Sau3A fragment of pCB102 underwent integration into the B . subtilis chromosome.

Biomed Mass Spectrom, 1985 Aug, 12(8), 359 - 63
Gas chromatography/mass spectrometry of bacterial amines; Tavakkol A et al.; Bacterial amines were examined by gas chromatography/mass spectrometry . Under electron impact all trifluoroacetamides exhibited peaks at m/z 69 due to {CF3}+ . Many trifluoroacetamides also showed peaks at m/z 97 corresponding to the {COCF3}+ ion fragment . The spectra of n-alkyl and aralkyl trifluoroacetamides were consistent with the spectra and their interpretations in the earlier literature . Molecular ions were of low abundance for all alkyl trifluoroacetamides having alkyl chains longer than two carbon atoms . Chemical ionization gave molecular weight information in all cases . Most peaks observed were molecular addition products, e.g . {M + H}+ and {M + NH4}+ . Application of chemical ionization mass spectrometry to analysis of bacterial amines revealed the production of beta-phenylethylamine, n-decylamine, 1,4-diaminobutane and 1,5-diaminopentane by Clostridium histolyticum; whereas both Clostridium bifermentans and Clostridium oedematiens produced beta-phenylethylamine . The latter organism also produced a peak with a retention time similar to that of an authentic amylamine derivative.

Appl Environ Microbiol, 1985 Aug, 50(2), 249 - 56
Activation and injury of Clostridium perfringens spores by alcohols; Craven SE et al.; The activation properties of Clostridium perfringens NCTC 8679 spores were demonstrated by increases in CFU after heating in water or aqueous alcohols . The temperature range for maximum activation, which was 70 to 80 degrees C in water, was lowered by the addition of alcohols . The response at a given temperature was dependent on the time of exposure and the alcohol concentration . The monohydric alcohols and some, but not all, of the polyhydric alcohols could activate spores at 37 degrees C . The concentration of a monohydric alcohol that produced optimal spore activation was inversely related to its lipophilic character . Spore injury, which was manifested as a dependence on lysozyme for germination and colony formation, occurred under some conditions of alcohol treatment that exceeded those for optimal spore activation . Treatment with aqueous solutions of monohydric alcohols effectively activated C . perfringens spores and suggests a hydrophobic site for spore activation.

Appl Environ Microbiol, 1985 Aug, 50(2), 202 - 6
Development of a minimal medium for Clostridium perfringens by using an anaerobic chemostat; Goldner SB et al.; A minimal medium was developed for the cultivation of Clostridium perfringens in an anaerobic chemostat . Cultures of C . perfringens ATCC 3624 and NCTC 10240 were grown at 46 and 43 degrees C, respectively, in a glucose-limited, chemically defined medium at pH 7.2 . The concentrations of amino acids, minerals, nucleotides, and vitamins, initially present in excess, were varied independently . The minimum concentration of each nutrient which would support 3 X 10(8) CFU/ml with a generation time of less than 40 min was determined and used to develop a reformulated defined medium . Atomic absorption spectroscopy and amino acid analyses of the reformulated medium indicated additional adjustments in nutrient content which led to the development of a minimal medium for each strain . The nutritional profile for each strain was similar . A decrease in the concentration of arginine, histidine, and tyrosine for strain 3624 and of arginine, histidine, and isoleucine for strain 10240 resulted in an increase in the optical density of each culture.

Biochem Biophys Res Commun, 1985 Jul 31, 130(2), 904 - 9
The cellulolytic enzyme complex of Clostridium thermocellum is very large; Coughlan MP et al.; The cellulolytic enzyme system bound to cellulose during the early stages of growth of C . thermocellum on this substrate was resolved into two major complexes . These complexes, as viewed by electron microscopy, are spherical particles with diameters of 210 A and 610 A and calculated molecular weights of 4.2 million and 102 million daltons, respectively.

Vet Rec, 1985 Jul 20, 117(3), 58 - 60
Diagnosis and treatment of botulism in lions; Greenwood AG; Six circus lions (Panthera leo) showed neurological and gastrointestinal signs after consuming casualty broiler chickens . Signs included ataxia, hindlimb paralysis and recumbency . Neurological examination of two affected males showed paralysis of extraocular muscles, fixed dilated pupils and inability to swallow . Replacement fluids and antibiotics were given and Clostridium botulinum type C antitoxin was found in serum samples . Type C antitoxin was not then available and therapy was started in one lioness with guanidine hydrochloride . Convulsions were controlled by diazepam but this animal died . One of the two males was given type C antitoxin; both were given anabolic steroids . All the remaining animals made slow recoveries over varying periods; one lion was recumbent for 41 days . No lion developed respiratory paralysis; other animals which had consumed the chickens remained healthy . Aspects of the treatment of botulism in animals are discussed.

Biochim Biophys Acta, 1985 Jul 9, 835(2), 304 - 14
Loss of stereospecificity of phospholipases C and D upon introduction of a 2-alkyl group into rac-1,2-diacylglycero-3-phosphocholine; Bugaut M et al.; rac-1-{1-14C}Lauroyl-2-oleylglycero-3-phospho{methyl-3H}choline and rac-1-lauroyl-2-{1-14C}oleoylglycero-3-phospho{methyl-3H}choline along with rac-1-palmitoyl-2-oleylglycero-3-phosphocholine and sn-1-palmitoyl-2-oleylglycero-3-phosphocholine were synthesized and subjected to hydrolysis with phospholipase C (EC 3.1.4.3) from Clostridium perfringens and phospholipase D (EC 3.1.4.4) from cabbage . Kinetics of hydrolysis of the radioactive substrates were determined by measuring the 3H radioactivity retained in the aqueous phase due to free choline and phosphocholine and the 3H and 14C radioactivity recovered in the organic phase due to the released diacylglycerols and phosphatidic acids and the residual phosphatidylcholines . The rate of hydrolysis of the unlabelled substrates by phospholipase C was determined by thin-layer chromatography and gas-liquid chromatography of the methanolysis products . The relative initial rates of hydrolysis of sn-1,2,- and sn-2,3-enantiomers were 100-200:1 for phospholipase C and 40-50:1 for phospholipase D using rac-1-lauroyl-2-oleoylglycero-3-phosphocholine as the substrate . The substitution of the 2-acyl group by an alkyl group resulted in a loss of stereospecificity, which was partial for phospholipase C (relative rates equal to 8-13:1) and total for phospholipase D . There was a parallel dramatic decrease (500-1000-fold) in the initial rate of hydrolysis with phospholipase C but the activity of phospholipase D was only moderately reduced (18-fold) . These findings are consistent with the earlier observed loss of the stereospecificity of lipoprotein lipase following introduction of a 2-alkyl group into triacylycerols, and point to a general unsuitability of 2-alkyl-linked acylglycerols as substrates for the assay of the stereospecificity of lipases, as well as for the isolation of enantiomeric 2-alkylacylglycerols by means of stereospecific lipases.

J Gen Microbiol, 1985 Jul, 131 ( Pt 7), 1697 - 703
Taxonomic position of lecithinase-negative strains of Clostridium sordellii; Popoff MR et al.; Eleven out of 43 strains of Clostridium sordellii from clinical sources did not produce lecithinase activity and were not toxic to mice . However, these strains did belong to the C . sordellii group and could readily be differentiated from C . bifermentans and C . difficile on the basis of DNA-DNA homologies, carbohydrate fermentation patterns, enzyme activities, GLC analysis of fatty acid fermentation products and the electrophoretic analysis of whole cell protein extracts.

Arch Microbiol, 1985 Jul, 142(2), 128 - 35
A sodium ion gradient as energy source for Peptostreptococcus asaccharolyticus; Wohlfarth G et al.; The determination of enzymatic activities in cell-free extracts of Acidaminococcus fermentans and Peptostreptococcus asaccharolyticus led to a refined scheme for the pathway of glutamate fermentation via (R)-2-hydroxyglutarate to acetate and butyrate . From the ratio of these products the amount of ATP generated by substrate level phosphorylation was calculated . Growth experiments with the organisms including Clostridium symbiosum and Clostridium tetanomorphum indicated that a sodium gradient contributed additional energy for growth . The high growth yields found in organisms containing the biotin dependent sodium pump glutaconyl-CoA decarboxylase could be reduced by the sodium ionophor monensin . In P . asaccharolyticus energy equivalent up to 0.6 mol ATP per mol of glutaconyl-CoA decarboxylated was conserved via the Na+ gradient . The data may explain the growth promoting effects of monensin in cattle.

J Assoc Off Anal Chem, 1985 Jul-Aug, 68(4), 626 - 31
Gas chromatographic detection of D-(-)-2,3-butanediol and butyric acid produced by sporeformers in cream-style corn and canned beef noodle soup: collaborative study; Schafer ML et al.; A gas chromatographic method that identifies sporeformers as the cause of spoilage in swollen cans of low-acid foods was collaboratively studied in 2 stages . Two organic compounds produced by sporeformers, D-(-)-2,3-butanediol and butyric acid, are measured in the upper phase after centrifugation of the liquid portion of the can contents . Each sample is assayed on 2 packed columns designed for the assay of aqueous solutions of volatile fatty acids, using flame ionization detectors . For study 1, 16 duplicate inoculated cans of cream-style corn and beef noodle soup were sent to 9 collaborators . For study 2, 7 collaborators received 11 duplicate inoculated cans of the 2 foods . Duplicate uninoculated cans of each food served as negative controls . The inocula were 6 sporeforming organisms (4 Clostridium and 2 gas-forming Bacillus species) and 2 nonsporeformers . After the deletion of marginal samples, the percentages of correctly identified sporeformers and nonsporeformers in beef noodle soup were 83 (110/132) and 90 (54/60), respectively; corresponding percentages for cream-style corn were 80 (98/123) and 100 (35/35) . The method has been adopted official first action.

J Antimicrob Chemother, 1985 Jul, 16 Suppl A, 137 - 49
Evolution and epidemiology of MLS resistance; Duval J; Within the framework of this symposium, it is not feasible to present an exhaustive description of the present state of knowledge regarding the sensitivity and resistance of bacterial species to macrolides, lincosamides and streptogramins (MLS) . This paper is limited to a description of the evolution of different types of resistance in the light of decisive factors described in previous papers, in order to deduce, if at all possible, trends in future strategy in therapeutics . Only acquired resistance lends itself to epidemiological study, in contrast to natural resistance which is, by definition, characteristic of a species or a genus, and not liable to change . Three groups will therefore be studied in turn: Staphylococcus aureus, streptococci and Bacteroides fragilis . There is as yet insufficient accumulated data to draw conclusions regarding the epidemiology and evolution of MLSB resistance observed in Clostridium perfringens and Corynebacterium diphtheriae, or regarding the high-level resistance to erythromycin due to enzymatic inactivation recently described in Escherichia coli.

Appl Environ Microbiol, 1985 Jul, 50(1), 63 - 7
Monoclonal antibody-based immunoassay for type A Clostridium botulinum toxin is comparable to the mouse bioassay; Shone C et al.; A monoclonal antibody (BA11) has been produced against Clostridium botulinum type A neurotoxin by the fusion of myeloma cells (P3 NS1/1-Ag4-1) with spleen cells from BALB/c mice immunized with botulinum type A neurotoxoid . The antibody bound specifically to botulinum type A neurotoxin, showing no cross-reactivity with types B and E botulinum toxins or with any of several other bacterial toxins tested . The monoclonal antibody did not bind to botulinum type A neurotoxin which had been denatured with sodium dodecyl sulfate and bound only weakly to each of the separated heavy and light subunits of the neurotoxin, suggesting a conformational requirement for the antigenic determinant of the antibody . A sensitive immunoassay for C . botulinum type A toxin with monoclonal antibody BA11 in conjunction with an enzyme amplication system has been developed which allows detection of 5 to 10 mouse 50% lethal doses ml-1 of purified neurotoxin . The assay was equally sensitive when applied to the detection of crude toxin in food stuffs; the average value for the minimum level of detectable toxin in extracts of tinned salmon or corned beef was 9 +/- 3.1 mouse 50% lethal doses ml-1.

Rev Infect Dis, 1985 Jul-Aug, 7 Suppl 3, S417 - 25
In vitro activity of imipenem against anaerobic bacteria; Wexler HM et al.; The in vitro activity of imipenem, metronidazole, clindamycin, moxalactam, and cefoxitin against 203 strains of anaerobic bacteria isolated from patients at the Veterans Administration Wadsworth Medical Center in Los Angeles was studied . Imipenem and metronidazole were the most active agents overall, inhibiting 98% and 99%, respectively, of all anaerobes tested . At breakpoint levels all of the agents tested were very active against anaerobic cocci . Clostridium perfringens, and Bacteroides species other than those of the Bacteroides fragilis group . Imipenem, metronidazole, and clindamycin were the most active agents against the B . fragilis group in this study, although more recent experience with clindamycin indicates less potency . Marked variation among the susceptibility results obtained at various centers may be due to differences in technique, including inoculum size, media, and incubation time . In all instances, however, imipenem has clearly been the most active of the beta-lactam agents.

Curr Eye Res, 1985 Jul, 4(7), 803 - 6
Inhibition of collagenase activity by extracts of bovine ocular tissues; Harper J et al.; Bovine eyes were dissected and separate pools of lens, lens capsule, cornea and vitreous were extracted in guanidine, subjected to ultrafiltration, and examined for their effects on collagenolytic activity . Although lens extract was not inhibitory, the cornea and vitreous both contained inhibitors of collagenase . More inhibition was present in the filtrate of the vitreous than in the retentate, whereas the total amount of inhibition in the cornea was distributed almost equally between the two fractions . The inhibition observed was dose dependent . The partially purified inhibitors from cornea and vitreous blocked the activity of human skin and tadpole back skin collagenases, but they failed to inhibit the bacterial (Clostridium histolyticum) collagenase . The inhibitor was stable to heating to 60 degrees for 30 minutes and to trypsinization.

Appl Environ Microbiol, 1985 Jul, 50(1), 16 - 20
Inhibition of Clostridium botulinum 52A toxicity and protease activity by sodium acid pyrophosphate in media systems; Wagner MK et al.; The effects of two pH levels (5.55 or 5.85) in combination with 0.4% sodium acid pyrophosphate (SAPP), NaH2PO4 X H2O, Na2HPO4 X 7H2O, or NaCl on the growth and toxicity of Clostridium botulinum 52A were studied . Absorbancy measurements at 630 nm, microscopic observations, and the mouse bioassay procedure were used to observe the effects . At pH 5.55 and 5.85 most control cultures exhibited toxicity when cell lysis began . Vegetative cell development was normal (4 micron long; 1 micron wide) . SAPP-containing (0.4%) treatment cultures displayed similar growth and lysis but no or delayed (48 h) toxicity . Cells grown in the SAPP treatment culture were longer and wider (6 micron long; 1.5 micron wide) than in most other treatment cultures . Trypsinization of nontoxic supernatants from 0.4% SAPP resulted in toxicity . Addition of 0.4% SAPP to toxic C . botulinum supernatant delayed but did not prevent death of mice . The addition of various levels of SAPP to toxic supernatants resulted in a decrease in zone size with an increase in the level of SAPP (9 mm with 0.4% SAPP to 7 mm with 1.0% SAPP), using a dual substrate protease assay . A decrease in the zone size also occurred with the supernatant from cultures grown in the presence of SAPP and with Bacillus polymyxa protease dilutions containing 0.4% SAPP . Results suggest that the actual production or function of the protease responsible for toxin activation may have been inhibited by the presence of SAPP.

Plasmid, 1985 Jul, 14(1), 37 - 46
Worldwide distribution of the conjugative Clostridium perfringens tetracycline resistance plasmid, pCW3; Abraham LJ et al.; The aim of this study was to test the hypothesis that all conjugative R-plasmids of Clostridium perfringens are closely related to the previously characterized tetracycline resistance plasmid, pCW3 . Fourteen conjugative R-plasmids derived from 11 C . perfringens strains isolated in Australia, the United States, France, Belgium, and Japan were analyzed . Eleven of the plasmids encoded tetracycline resistance while three carried both tetracycline and chloramphenicol resistance . Each of these plasmids was compared, by restriction analysis, to the reference plasmid, pCW3 . Seven of the tetracycline resistance plasmids had EcoRI, XbaI, and ClaI restriction profiles that were identical to those of the corresponding pCW3 digests . The seven remaining R-plasmids were different from pCW3 . Comparison of partial restriction maps of these plasmids with a complete map of pCW3 indicated that they contained at least 17 kb of DNA that also was present in pCW3 . Hybridization analysis confirmed that these plasmids shared substantial homology with pCW3 . The three tetracycline and chloramphenicol resistance plasmids frequently lost a 6-kb chloramphenicol resistance segment during conjugation . Cloning experiments showed that the chloramphenicol resistance determinant was expressed in Escherichia coli and that the chloramphenicol resistance gene of one of these plasmids, pIP401, was contained within a 1.5-kb region of the 6-kb deletion segment . Hybridization analysis indicated that the deletion segment of pIP401 was related to those of the other two chloramphenicol resistance plasmids . During the course of this study, conjugative R-plasmids which appear to be identical to pCW3 or closely related to pCW3 were identified from C . perfringens strains from human, animal and environmental sources in five countries . It is concluded that C . perfringens strains in humans and animals throughout the world have overlapping gene pools and that all the conjugative C . perfringens R-plasmids examined probably evolved from a pCW3-like element.

J Assoc Off Anal Chem, 1985 Jul-Aug, 68(4), 807 - 8
Comparison of rapid perfringens medium and lactose sulfite medium for detection of Clostridium perfringens; Smith M; Two tubed media, lactose sulfite (LS) and rapid perfringens medium (RPM), were evaluated and compared for their ability to detect and enumerate Clostridium perfringens in inoculated and naturally contaminated samples . In a 3-tube most probable number system, the number of organisms recovered from spiked samples was several-fold higher with RPM than with LS and was close to the numbers inoculated . Levels detected in 15 naturally contaminated samples were also higher with RPM . Only 13 of the samples were positive for C . perfringens with LS medium . These results indicate that RPM is a more effective medium than LS; therefore, its use is recommended.

J Clin Microbiol, 1985 Jul, 22(1), 52 - 5
Comparative evaluation of three identification systems for anaerobes; Murray PR et al.; The accuracy of two new 4-h identification systems for anaerobes, the AN-IDENT (Analytab Products, Plainview, N.Y.) and the RapID ANA (Innovative Diagnostic Systems, Inc., Atlanta, Ga.) was compared with that of the API 20A system (Analytab Products) . A total of 132 clinical isolates were tested in each of the three systems . The overall accuracies at the genus and species level for the three systems were: API 20A, 68.9 and 56.8%, respectively; AN-IDENT, 90.2 and 73.5%; and RapID ANA, 93.9 and 81.8% . Improved identification of anaerobes with the AN-IDENT and the RapID ANA systems was observed for isolates of the genus Fusobacterium, Clostridium species other than Clostridium perfringens, non-spore-forming bacilli, and isolates of the genus Peptostreptococcus . Reproducibility studies demonstrated that the results of the individual test reactions in all three identification systems were reproducible when the interpretive guidelines of the manufacturer were followed precisely.

J Clin Microbiol, 1985 Jul, 22(1), 129 - 31
Neuraminidase activity is not the cause of influenza virus-induced neutrophil dysfunction; Abramson JS et al.; Influenza viruses have been shown to decrease the ability of polymorphonuclear leukocytes (PMN) to respond to a variety of stimuli . This study was done to determine if viral neuraminidase was responsible for decreased PMN function . Treatment of human PMN with purified neuraminidases from influenza virus, Vibrio cholerae, or Clostridium perfringens did not significantly affect the ability of human PMN to respond to stimulation . Occasional virus preparations that lacked the ability to depress PMN function did not differ in neuraminidase activity from viruses capable of causing depression . These results demonstrate that neuraminidase activity is not the cause of influenza virus-induced PMN dysfunction.

Ann Inst Pasteur Microbiol, 1985 Jul-Aug, 136B(1), 75 - 91
{Demonstration of 2 main serological groups in Clostridium tyrobutyricum}; Bergere JL; Antisera prepared against heated cells of 10 strains of Clostridium tyrobutyricum gave cross-reactions with several other clostridial species . Such reactions could be completely eliminated by absorption with C . beijerinckii, C . fallax and C . tetanomorphum . Both agglutination and immunofluorescence tests with these antisera showed that 85 strains of C . tyrobutyricum were divided into two main serological groups, A and B . The 56 strains of group A possessed the same thermostable species-specific antigen, whereas those of group B were lacking in it . Antisera prepared against either formol- or ethanol-treated cells or low-heated cells of C . tyrobutyricum showed only low titre cross-reactions with other clostridial species . When these were removed by absorption, only the homologous strains and a few other strains of C . tyrobutyricum group B were still agglutinated at a full titre by these antisera . Therefore, only strains of group A of C . tyrobutyricum can be easily identified by a single type-specific antiserum.

Vet Rec, 1985 Jun 29, 116(26), 687 - 9
Gastritis in Lake Tanganyika cichlids (Tropheus duboisii); Ferguson HW et al.; Necrotic and granulomatous gastritis is described in Lake Tanganyika cichlids . Clostridium hastiforme and flagellated protozoa were both associated with the reaction but the significance of either is unknown . Nevertheless, treatment of surviving fish with ampicillin was carried out and mortalities ceased . The possible involvement of an unsuitable diet as a predisposing factor is discussed.

J Biol Chem, 1985 Jun 25, 260(12), 7442 - 51
31P NMR studies of Clostridium thermocellum . Mechanism of end product inhibition by ethanol; Herrero AA et al.; 31P NMR studies of intact cells and perchloric acid extracts are used to investigate the effect of ethanol on the bioenergetics and glycolysis of Clostridium thermocellum, an anaerobic bacterium potentially useful for the single step conversion of biomass to ethanol . Whole cells suspended in phosphate buffer and given a carbon source (cellobiose) at 60 degrees C rapidly establish a pH gradient across the membrane that can be monitored by the chemical shifts of inorganic phosphate in the exterior buffer and in the cytoplasm . Peak intensities can be related to phosphate active transport rates . Wild type bacteria and cells grown in inhibiting concentrations of ethanol establish similar pH gradients, but with slower kinetics and slower phosphate transport rates for the cells adapted to growth in ethanol . Direct addition of ethanol does not affect the rate of pH gradient formation or phosphate transport . Thus, while ethanol does not directly affect processes for energy conservation carried out by the membrane, adaptation to ethanol does alter membrane functions such as phosphate transport . 31P NMR spectra of perchloric acid extracts show that when wild type cells are adapted to grow in inhibiting concentrations of ethanol and then energized with cellobiose, sugar phosphate content is increased and the steady state distribution of glycolytic intermediates is altered . Nucleotide triphosphate/nucleotide diphosphate ratios are unaltered in these cells . These results strongly indicate that in C . thermocellum growth inhibition by ethanol is related to a blockage in glycolysis.

Biochemistry, 1985 Jun 18, 24(13), 3149 - 57
Clostridium histolyticum collagenase: development of new thio ester, fluorogenic, and depsipeptide substrates and new inhibitors; Vencill CF et al.; A new series of thio ester, depsipeptide, and peptide substrates have been synthesized for the bacterial enzyme Clostridium histolyticum collagenase . The hydrolysis of the depsipeptide substrate was followed on a pH stat, and thio ester hydrolysis was measured by inclusion of the chromogenic thiol reagent 4,4'-dithiopyridine in the assay mixture . The best thio ester substrate, Boc-Abz-Gly-Pro-Leu-SCH2CO-Pro-Nba, had a kcat/KM of 63 000 M-1 s-1, while several shorter thio ester sequences were inactive as substrates . In general, the peptide analogues of all the reactive thio ester substrates were shown to be hydrolyzed 5-10 times faster by collagenase . In one case (Z-Gly-Pro-Leu-Gly-Pro-NH2) where a comparison was made, the peptide substrate was respectively 8- and 106-fold more readily hydrolyzed than the corresponding thio ester and ester substrates . Cleavages of the two fluorescence-quench substrates Abz-Gly-Pro-Leu-Gly-Pro-Nba and Abz-Gly-Pro-Leu-SCH2CO-Pro-Nba could be easily followed fluorogenically since a 5-10-fold increase in fluorescence occurred upon hydrolysis . The fluorescent peptide substrate is the best synthetic substrate known for C . histolyticum collagenase with a kcat/KM value of 490 000 M-1 s-1 . A series of new reversible inhibitors were developed by the attachment of zinc ligating groups (hydroxamic acid, carboxymethyl, and thiol) to various peptide sequences specific for C . histolyticum collagenase . The shorter peptides designed to bind to either the P3-P1 or P1'-P3' subsites were poor to moderate inhibitors . The thiol HSCH2CH2CO-Pro-Nba had the lowest K1 (0.02 mM).(ABSTRACT TRUNCATED AT 250 WORDS)

FEBS Lett, 1985 Jun 17, 185(2), 253 - 6
Identification of acrylate, the product of the dehydration of (R)-lactate catalysed by cell-free extracts from Clostridium propionicum; Schweiger G et al.; Cell extracts from Clostridium propionicum harvested in the late log-phase catalysed the dehydration of (R)-lactate to acrylate at a maximum rate of 0.06 U/mg protein . The unsaturated acid was identified by high-performance liquid chromatography and as p-bromophenacyl ester by gas chromatography combined with mass spectroscopy . The amount of acrylate formed was dependent on protein and (R)-lactate concentrations . However, due to product inhibition the yield of acrylate did not exceed 0.5% . Like the dehydration of (R)-2-hydroxyglutarate to glutaconate the dehydration of (R)-lactate to acrylate was inhibited by 1 mM hydroxylamine, 1mM azide, 0.1 mM dinitrophenol, 10 mM EDTA or by exposure to air . A radical mechanism is postulated.

FEBS Lett, 1985 Jun 17, 185(2), 267 - 71
Clostridium pasteurianum glutamine synthetase mechanism . Evidence for active site tyrosine residues; Krishnan IS et al.; Preliminary chemical modification studies indicated the presence of tyrosine, carboxyl, arginine, histidine and the absence of serine and sulfhydryl residues at or near the active site of Clostridium pasteurianum glutamine synthetase . The conditions for tyrosine modification with tetranitromethane were optimized . The inactivation kinetics follow pseudo-first-order kinetics with respect to enzyme and second order with respect to modifier per active site . There was no inactivation at pH 6.5 suggesting the absence of thiol oxidation . The synthetase and transferase reactions followed the same pattern of inactivation on enzyme modification and both were equally protected by glutamate plus ATP . Thus tyrosine residues are present at the active site of the enzyme and are essential for both transferase and synthetase activities.

Am J Med, 1985 Jun 7, 78(6A), 95 - 9
Imipenem/cilastatin therapy of bacteremia; Eron LJ; Imipenem/cilastatin was used to treat 135 patients with bacteremia and signs of infection . Ninety percent responded favorably . The bacteriologic eradication rate was 98 percent among the 153 isolates . Only one patient had breakthrough bacteremia and this was a susceptible Clostridium septicum . In two isolates of Pseudomonas aeruginosa, emergence of resistance to imipenem during therapy was noted and appeared to be responsible for clinical failure . Superinfection occurred in 13 patients and was responsible for two treatment failures.

Am J Med, 1985 Jun 7, 78(6A), 134 - 9
Role for newer beta-lactam antibiotics in treatment of osteomyelitis; Gentry LO; Monotherapy of osteomyelitis with the newer broad-spectrum beta-lactam antibiotics has become attractive because of the efficacy, safety, and cost of these antibiotics when compared with conventional combination therapy . Imipenem/cilastatin is a recent and promising addition to this antibiotic family . Experience with imipenem/cilastatin and that reported for cefotaxime, ceftazidime, and ceftizoxime in the treatment of biopsy-proved osteomyelitis was compared, using data from published reports from five centers . Two hundred forty-three patients were evaluable: 34 were treated with imipenem/cilastatin, 84 with cefotaxime, 122 with ceftazidime, and 33 with ceftizoxime . Staphylococcus aureus was isolated by 80 bone cultures and was the most common single species encountered . There were 75 isolates of Pseudomonas aeruginosa, 113 mixed Enterobacteriaceae species, 115 mixed gram-positive and -negative isolates of miscellaneous species, and 30 anerobic isolates . Polymicrobial infection was present in 101 cases (41.6 percent) . Failure rates were similarly low in all groups (10 to 30 percent) . However, resistance developed during therapy in all groups with P . aeruginosa . Side effects were predictably few, but reversible neutropenia, pseudomembranous colitis due to Clostridium difficile, and nausea required therapy to be discontinued in seven patients . Imipenem/cilastatin should prove to be a very effective and relatively safe single agent for treatment of osteomyelitis.

Eur J Biochem, 1985 Jun 3, 149(2), 287 - 93
Purification and properties of an enterotoxin from a coatless spore mutant of Clostridium perfringens type A; Lindsay JA et al.; A method is described for isolating an enterotoxin from a coatless spore mutant (8-6) of Clostridium perfringens type A . The characteristics of this enterotoxin only slightly resembled those of previously isolated enterotoxins of C . perfringens . The type A (8-6) enterotoxin was found to be composed of two subunits of Mr 18 000 with isoelectric points of 3.8 and 4.3 . The LD50 for mice was 39 micrograms/kg with 0.10 micrograms corresponding to one erythemal unit . The type A (8-6) enterotoxin was inactivated by heating for 10 min at 60 degrees C . The amino acid composition data of type A (8-6) and delta toxins was similar, but type A (8-6) and type A enterotoxins showed less similarity . This lack of similarity between type A and type A (8-6) enterotoxins was confirmed by the failure of anti-sera to type A enterotoxin to neutralize the type A (8-6) enterotoxin, in both the mouse and erythemal tests.

Vet Microbiol, 1985 Jun, 10(4), 315 - 24
An indirect hemagglutination test for the detection of antibodies to Clostridium chauvoei; Tamura Y et al.; An indirect hemagglutination (IHA) test using sonicated extract as the antigen was developed for the detection of antibodies to Clostridium chauvoei . This antigen can be adsorbed onto glutaraldehyde-fixed sheep red blood cells treated with tannic acid and can be destroyed by trypsin and heat treatment . It corresponded well with the flagella of the organism, when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the gel diffusion test . No serological cross-reactivity was found in the IHA test when the antigen was tested against 4 species of clostridial antibodies . Our results suggest that the IHA test mainly detects antibodies against the flagella of C . chauvoei.

J Antimicrob Chemother, 1985 Jun, 15 Suppl C, 193 - 8
Susceptibility of anaerobic bacteria to Sch 34343 and other antibiotics; Glupczynski Y et al.; The in-vitro activity of Sch 34343 was evaluated against 137 strains of anaerobic bacteria by the agar dilution technique . Sch 34343 was compared with imipenem, cefoxitin, latamoxef (moxalactam), clindamycin and metronidazole . Organisms studied included the Bacteroides fragilis group, other Bacteroides spp., Clostridium perfringens, Cl . difficile, other Clostridium spp . and anaerobic cocci . Overall, Sch 34343 and imipenem were significantly more active than the other antibiotics against most organisms tested, especially the Bact . fragilis group, including clindamycin-resistant strains . Apart from Cl . difficile, which required up to 8 mg/l of Sch 34343 and imipenem for inhibition, all the strains were inhibited by 1 mg/l of Sch 34343 and by 2 mg/l of imipenem . Of the remaining agents tested, against the Bact . fragilis group metronidazole (2 mg/l to inhibit 90% of the strains) was the most active, followed by cefoxitin (16 mg/l), latamoxef (32 mg/l) and clindamycin (32 mg/l) . On the basis of its activity in vitro, Sch 34343 appears to be one of the most promising new antimicrobial agents for the treatment of infections involving anaerobic bacteria.

J Antimicrob Chemother, 1985 Jun, 15 Suppl C, 189 - 91
Susceptibility of anaerobic bacteria to Sch 34343 and other beta-lactam antibiotics; Jacobus NV et al.; The in-vitro activity of Sch 34343 was evaluated against 381 anaerobic clinical isolates and compared with that of imipenem, cefoxitin, latamoxef (moxalactam), cefotaxime and cefoperazone . Test isolates included strains with unknown resistance to the beta-lactam drugs, and two Bacteroides fragilis strains that could inactivate cefoxitin . Sch 34343 and imipenem demonstrated superior activity against all the anaerobic bacteria tested, with MIC90s less than or equal to 1 mg/l . Sch 34343 was generally one- to two-fold more active than imipenem . Resistance to both was demonstrated by the two Bact . fragilis strains that inactivated cefoxitin and by one strain of Fusobacterium; these three strains were also resistant to the other drugs studied . There were no other resistant strains to either Sch 34343 or imipenem . Cefotaxime and cefoperazone were the least active against the Bact . fragilis group, with resistance rates of 33 and 47%, respectively . Strains resistant to cefoxitin, latamoxef, cefotaxime and cefoperazone were found among some Clostridium spp . The Gram-positive cocci and Gram-positive bacilli were sensitive to all the drugs evaluated.

J Antimicrob Chemother, 1985 Jun, 15 Suppl C, 177 - 82
Comparative in-vitro activity of Sch 34343 for a wide spectrum of clinically significant anaerobic bacteria; Wells CL et al.; One hundred and fifty strains of anaerobic bacteria including 45 bacteroides, 19 fusobacteria, 41 cocci, 34 clostridia, and 11 Gram-positive non-sporeforming rods were tested by agar dilution for their susceptibilities to cefoxitin, cefuroxime, latamoxef (moxalactam), penicillin G, chloramphenicol, clindamycin, metronidazole and Sch 34343 . Excluding the 34 clostridia, 115 of the 116 remaining strains were inhibited by less than or equal to 1 mg/l of Sch 34343 . One isolate of Bacteroides fragilis required 32 mg/l for inhibition . All of the 34 clostridia were inhibited by less than or equal to 8 mg/l of Sch 34343: 14 isolates of Clostridium difficile had an MIC50 and an MIC90 of 4 mg/l, whereas the remaining 20 species of clostridia had an MIC50 of 0.125 mg/l and an MIC90 of 2 mg/l . On a weight basis, Sch 34343 was generally more active than any of the seven other antimicrobial agents tested.

Antimicrob Agents Chemother, 1985 Jun, 27(6), 961 - 3
Activity of a peptidyl prodrug, alafosfalin, against anaerobic bacteria; Grappel SF et al.; Alafosfalin, an antibacterial phosphonodipeptide requiring peptide transport for activity, was tested for activity against clinical strains of anaerobic bacteria in peptide-free Roche Sensitivity Test Medium no . 5 agar . It was active against Bacteroides spp., Fusobacterium nucleatum, and Clostridium perfringens but not against Clostridium difficile . Alafosfalin activity was antagonized by appropriate peptides . Synergy was obtained with other cell wall-active antibiotics.

J Med Microbiol, 1985 Jun, 19(3), 351 - 7
Lesions produced by Clostridium butyricum strain CB 1002 in ligated intestinal loops in guinea pigs; Popoff MR et al.; Heated spores (80 degrees C, 10 min) of Clostridium butyricum strain CB 1002 isolated from a fatal case of necrotising enterocolitis in a human neonate were inoculated into ligated intestinal loops prepared in young conventional guinea pigs . Necropsy findings 18 h later included congestion, patchy haemorrhage of the intestinal mucosa and bacteraemia . No abnormalities were observed in control loops given inocula of inactivated spores (heated at 100 degrees C for 10 min) or TYG 6 medium . The results suggest that vascular lesions are produced by C . butyricum in the intestine of young conventional guinea pigs.

J Med Microbiol, 1985 Jun, 19(3), 339 - 50
Protection of hamsters against Clostridium difficile ileocaecitis by prior colonisation with non-pathogenic strains; Borriello SP et al.; Prior colonisation of clindamycin-treated hamsters with non-toxigenic strains of C . difficile protected them from subsequent colonisation with a toxigenic pathogenic strain . In total, 13 of 18 'protected' hamsters survived for up to 27 days whereas all 27 animals challenged with the toxigenic strain alone died within 48 h . Protection was not evident if a heat-killed suspension was used or if the colonising non-toxigenic strain was first removed with vancomycin . No antitoxic activity could be detected in the faeces of animals colonised with the non-toxigenic strains . Other species of clostridia did not protect against the lethal effects of subsequent exposure to the toxigenic strain . Conversely, non-toxigenic strains would not protect the animals from the lethal effects of a different clostridial pathogen, C . spiroforme . In most cases, even in the protected animals, the toxigenic strain eventually became dominant and caused disease, with translocation across the gut wall occurring early in the disease process . It was also shown that a non-toxigenic strain of C . difficile can adhere to gut mucosa . It is proposed that the protection afforded by the non-toxigenic strains may be due to competition for ecological niches.

Arch Phys Med Rehabil, 1985 Jun, 66(6), 394 - 6
Pseudomembranous colitis in spinal cord injury; Johnson DK et al.; Pseudomembranous colitis is a well-known disease associated with antibiotic administration and caused by the Clostridium difficile toxin . Clinical presentation is usually marked by watery diarrhea, crampy abdominal pain, and fever . Since early appropriate therapy can reduce morbidity and mortality, it is important for health care professionals to be aware of this disease . Patients with spinal cord injury have a relatively high incidence of respiratory and urinary tract infections that are treated with antibiotics . Therefore, these patients theoretically have a higher risk of contracting pseudomembranous colitis . This article presents a case report of a spinal cord injured patient with this disease who has several of the common difficulties encountered in the diagnosis and treatment, such as indeterminate assays and relapses . The clinical presentation, diagnosis, and treatment of pseudomembranous colitis are described.

J Appl Bacteriol, 1985 Jun, 58(6), 577 - 84
Mutagenesis of Clostridium acetobutylicum; Bowring SN et al.; Mutagenesis of the obligate anaerobe Clostridium acetobutylicum was best accomplished using agents (e.g . ethyl methane sulphonate or N-methyl-N'-nitro-N-nitrosoguanidine) which are believed to act by a direct mutagenic mechanism . Other agents (e.g . u.v . radiation) whose effectiveness relies on misrepair of damaged DNA via an error-prone pathway, were poor mutagens of this organism . Procedures are described which readily yielded a variety of auxotrophic and other useful mutant strains of Cl . acetobutylicum and related saccharolytic clostridia.

Zh Mikrobiol Epidemiol Immunobiol, 1985 Jun, (6), 3 - 6
{Production of tetanolysin preparations and characteristics of their properties}; Shabarov IA et al.; As the result of the study of tetanolysin-producing Clostridium tetani strains, their populations have been found to be markedly heterogeneous with respect to the hemolytic activity of clone cultures . On the basis of normal and dialyzed cultures of selected variants with maximum activity the preparations of tetanolysin have been obtained, and their hemolytic activity and antigenic properties have been studied . Antihemolytic rabbit sera have also been obtained and characterized . Partially purified preparations of tetanolysin with high hemolytic activity have been obtained by the fractionation of C . tetani dialyzed cultures with ammonium sulfate.

Infect Immun, 1985 Jun, 48(3), 769 - 75
Binding of the two components of C2 toxin to epithelial cells and brush borders of mouse intestine; Ohishi I et al.; C2 toxin elaborated by Clostridium botulinum types C and D is composed of two nonlinked protein components and has enterotoxic activity, for which the cooperation of these two components is necessary . In the present study, the binding of components I and II, the two components of C2 toxin, to isolated epithelial cells and brush borders of mouse intestine was examined . Immunofluorescence studies showed that component II, either trypsinized (T-II) or untrypsinized (UT-II), bound to the cells and the brush borders of mouse intestine, whereas component I alone did not . The binding of I was observed only when the cells and the brush borders were reacted with T-II, but not when they were reacted with UT-II . These results are consistent with the fact that the biological activities of C2 toxin are elicited by the combination of I and T-II, but not of I and UT-II . The in vitro binding of I and II to isolated brush borders of mouse intestinal cells also showed similar binding characteristics . The binding of I and II to brush borders was rapid and not temperature dependent . Ultracentrifugal analysis revealed that both I and T-II bound to microvillous membranes of the intestinal cells . The data from the present study indicate that the enterotoxic activity of C2 toxin is initiated by the binding of T-II to the microvillous membrane of intestinal cells followed by that of I, for which the site of the cell membrane is induced by the binding of T-II, but not of UT-II.

J Antimicrob Chemother, 1985 Jun, 15 Suppl C, 39 - 56
Evaluation of the in-vitro antibacterial activity of Sch 34343; Adam C et al.; The in-vitro activity (as measured by geometric mean MICs, mg/l) of Sch 34343 against aerobic and anaerobic bacteria was compared with that of 14 other selected beta-lactam antibiotics including aztreonam, latamoxef (moxalactam), ceftazidime and imipenem . Sch 34343 had good activity (less than 2 mg/l) against most Gram-negative aerobic bacteria whether or not they contained high levels of plasmid-mediated or chromosomally-mediated beta-lactamases . It was slightly less potent against strains of Morganella and Serratia (less than 4 mg/l) and inactive against Pseudomonas (greater than 64 mg/l) . A very small inoculum effect was observed against strains containing beta-lactamases indicating stability . Unlike the third-generation cephalosporins, Sch 34343 had excellent activity (less than or equal to 0.18 mg/l) against staphylococci, comparable to that of imipenem and ampicillin . While Sch 34343 had equally good potency (0.17 mg/l) against penicillinase-positive staphylococci, it was inactive against methicillin-resistant staphylococci (greater than or equal to 35 mg/l) . Sch 34343 also had good activity against streptococci . The most unusual aspect of the in-vitro activity was its activity against Bacteroides (including Bact . fragilis) and other anaerobes . Sch 34343 had mean MICs less than or equal to mg/l for all Bacteroides and Clostridium spp . tested except CI . difficile (3.4 mg/l).

Antimicrob Agents Chemother, 1985 Jun, 27(6), 980 - 1
Studies with temocillin in a hamster model of antibiotic-associated colitis; Boon RJ et al.; Hamsters given the new penicillin temocillin, either orally or by injection, did not develop antibiotic-associated colitis, whereas animals given the control antibiotics cefoxitin or clindamycin developed the disease, which is characterized by marked hemorrhagic cecitis and high cecal levels of Clostridium difficile cytotoxin.

J Antimicrob Chemother, 1985 Jun, 15(6), 671 - 7
The comparative in-vitro activity of cefotetan against anaerobic bacteria; Watt B et al.; The in-vitro activity of cefotetan, a new cephamycin, was assessed against a total of 336 strains of anaerobic bacteria by means of an agar dilution procedure and compared with that of cefoxitin, mezlocillin, piperacillin, clindamycin and metronidazole . Overall clindamycin and metronidazole were the most active of the test compounds . Cefotetan showed good activity against anaerobic cocci and clostridia, except for Clostridium difficile (MIC90 = 16 mg/l), although it was comparatively less active than the other beta-lactams against anaerobic cocci . In the case of the Gram-negative anaerobes, cefotetan showed moderate activity comparable to that of cefoxitin; against the 120 test strains of Bacteroides fragilis both cefotetan and cefoxitin were markedly more active than the penicillins . In studies with antibiotic combinations, cefotetan + cefsulodin showed marked synergy (FIC index less than 0.3) against the majority of strains of Bact . fragilis tested.

Jpn J Antibiot, 1985 Jun, 38(6), 1516 - 28
Effect of aspoxicillin on anaerobic bacteria; Ueno K et al.; Aspoxicillin (ASPC), a semisynthetic penicillin has a broad spectrum of antibacterial activities against Gram-positive and Gram-negative anaerobic bacteria . Its in vitro antibacterial activity was less than those of cefoxitin against Peptostreptococcus and Veillonella, but was significantly high against Bacteroides fragilis, one of the most clinically important anaerobe . The therapeutic and/or protective effect of ASPC in experimental subcutaneous abscess or experimental intraabdominal mixed infection due to beta-lactamase producing B . fragilis and non-producing Escherichia coli were much stronger than those of ticarcillin . In order to account the superiority of ASPC in vivo, the effects of ASPC and other beta-lactams on B . fragilis were compared and the results were analyzed in relation to their in vitro bactericidal activities, stability against the beta-lactamase, binding properties with penicillin-binding proteins and pharmacokinetic properties . Interestingly, administration of ASPC did not increase the bacterial counts of Clostridium difficile in caecal contents, but piperacillin, ticalcillin, carbenicillin, ampicillin and cefotaxime increased the counts.

Exp Mol Pathol, 1985 Jun, 42(3), 401 - 10
A comparison of the effects of cytotoxic cerebrospinal fluid on cell cultures with other cytopathogenic agents; Taylor GR et al.; Protein synthesis, antigen synthesis, and cell membrane permeability were analyzed after inoculating human diploid fibroblasts with control or cytotoxic CSF, herpes simplex virus type 1 (HSV 1), poliovirus 3, or Clostridium difficile toxin . Whereas protein synthesis and membrane permeability were affected by the viruses, and virus antigens detectable by pooled human serum were synthesized, the bacterial toxin and cytotoxic CSF did not induce any new proteins or antigens, although the cytotoxic CSF reduced cellular protein synthesis levels and caused an increase in the permeability of the cell membranes . The effect of the cytotoxic CSF in cell culture resembles that of a toxin rather than a replicating virus.

Rev Esp Fisiol, 1985 Jun, 41(2), 195 - 9
Utilization of cellobiose and D-glucose by Clostridium thermocellum ATCC-27405; Hernandez PE et al.; Cultures of Clostridium thermocellum ATCC-27405, maintained on cellulose and not adapted to grow on glucose utilize cellobiose preferentially over D-glucose, and are only able to initiate growth on D-glucose when the cellobiose has been exhausted from the growth medium . However, D-glucose is the carbon source preferentially utilized when cultures of this microorganism, previously adapted for growth on glucose, are transferred to a medium with equivalent concentrations of both sugars . One reason for the preferential utilization of glucose over that of cellobiose might be the competitive inhibition of cellobiose phosphorylase by intracellular glucose accumulation . When in the glucose-adapted cultures the pressure to grow on glucose as the sole carbon source is again released, both sugars can be simultaneously utilized.

Eur J Epidemiol, 1985 Jun, 1(2), 131 - 8
Purification by high performance liquid chromatography of Clostridium perfringens type A enterotoxin prepared from high toxin producers selected by a toxin-antitoxin halo; Sugimoto N et al.; High enterotoxin-producing substrains of Clostridium perfringens type A were selected reproducibly as colonies having toxin-antitoxin haloes on agar plates of Duncan-Strong medium containing antitoxin serum . Enterotoxin from these substrains was subjected to rapid purification by high performance liquid chromatography (HPLC) . For this, the toxin was extracted by sonication from sporulating bacteria grown in Duncan-Strong sporulation medium, fractionated by ammonium sulfate (40% saturation) precipitation and differential solubilization and then purified by HPLC: gel permeation chromatography through a G2000SW column and ion-exchange chromatography on a Mono Q column . Purified toxin preparations had a similar specific activity (4.2 X 10(2) mouse MLD/mg protein) and homogeneity on polyacrylamide gel-electrophoresis to preparations obtained by conventional gel permeation through a Sephadex-G200 column . By further HPLC on a Mono Q column, minor nontoxin proteins were separated from the toxin without loss of the toxicity on a protein basis . The final yield of the purified toxin was about 15% of that in the bacterial extract . The two HPLC procedures each took only one hour.

P N G Med J, 1985 Jun, 28(2), 75 - 82
A review of pigbel (necrotising enteritis) in Papua New Guinea, 1961-1984; Davis MW; Pigbel has been recognised as a major cause of mortality and morbidity in the Papua New Guinea Highlands for over 20 years . The clinical features, epidemiology and pathogenesis of this disease have been elucidated, leading to the development of an effective vaccine (Clostridium perfringens type C beta toxoid) for the prevention of pigbel.

Aust Vet J, 1985 Jun, 62(6), 194 - 6
Haemorrhagic necrotising enteritis in foals associated with Clostridium perfringens; Sims LD et al.; Two foals aged 35 and 48 h from 2 Thoroughbred studs died several hours after developing clinical signs of depression, severe haemorrhagic diarrhoea and dehydration . Both foals had an acute haemorrhagic enteritis extending from the anterior jejunum to the terminal ileum which was characterised histologically by villus necrosis . Necrotic villi were surrounded by large numbers of rod-shaped Gram positive bacteria . Clostridium perfringens was recovered from the intestines of both foals and the isolates were considered to be C . perfringens type C . Other cases of diarrhoea were also observed in foals of the same age on these 2 studs, but the aetiology of these was not determined.

J Gen Microbiol, 1985 Jun, 131 ( Pt 6), 1487 - 96
Purification and properties of spore-lytic enzymes from Clostridium perfringens type A spores; Gombas DE et al.; Spores of Clostridium perfringens contain at least two spore-lytic enzymes active in hydrolysing cortical peptidoglycan . One enzyme has been purified 1800-fold and has a molecular weight of 17 400 determined from chromatography on Sephadex G-75 . Two protein bands were apparent after SDS-PAGE . The isolated enzyme was investigated for response to temperature, pH, ionic strength and enzyme inhibitors, and for mode of action . A second enzyme activity, differing from the first in apparent molecular weight (29 800) as determined by gel exclusion chromatography, and also in its pH optimum and activity on cortical substrate, was also isolated, although not purified to the same extent.

J Clin Gastroenterol, 1985 Jun, 7(3), 269 - 72
Crohn's disease with persistence of Clostridium difficile, surgical elimination; DeRidder PH et al.; Clostridium difficile has been associated with increased activity of Crohn's disease in some patients, and in them its eradication has proved beneficial . We have seen a patient unresponsive to two courses of vancomycin with persistence of C . difficile colonization and toxin production in whom surgical intervention eliminated the C . difficile cytotoxin and organism.

Am Surg, 1985 Jun, 51(6), 301 - 3
Clostridium perfringens bacteremia . Opportunist or killer?
Nelson RM, Wilson RF, Osmer RL.
Clostridia septicemia has traditionally been associated with severe histocytotoxic infections . Recently, due to better anaerobic culture techniques, clostridia bacteremia is seen with increasing frequency in patients without an obvious source of infection . The authors reviewed their experience with 29 patients with clostridia bacteremia . The overall mortality rate was 45 per cent . No obvious source for the clostridia bacteremia was identified in 21 patients (72%) . Eighty-three per cent of the patients had polymicrobial infections . The patients were generally debilitated with significant associated systemic disease . Antibiotic therapy did not appear to have any effect on mortality . Patients with asymptomatic clostridial bacteremia are, in general, patients with advanced malignancies and chronic illnesses . Antibiotic therapy appears to have little effect on the patient's outcome.

Can J Microbiol, 1985 Jun, 31(6), 575 - 8
Development of a cell wash buffer that minimizes nucleic acid loss from Clostridium perfringens 10543 A; Blaschek HP et al.; Autolytic activity and nucleic loss from Clostridium perfringens 10543 A was demonstrated during successive cell washes in hypotonic TES buffer . Autolysis increased nearly sixfold and nucleic acid loss nearly twofold when 10 mM EDTA was added to 0.3 M Tris-sucrose buffer . Attempts to minimize both autolysis and nucleic acid loss from C . perfringens during routine washing steps were unsuccessful when the effects of sucrose concentration, pH, CaCl2 addition, or wash temperature were examined independently . However, autolytic activity was eliminated and nucleic acid loss reduced to less than 5% when C . perfringens cells were washed at 4 or 25 degrees C in 1.0 M sucrose, 50 mM Tris--HCl, and 25 mM CaCl2 at pH 5.7.

Arch Biochem Biophys, 1985 Jun, 239(2), 523 - 30
Purification and properties of a quinone-dependent p-nitrophenylphosphatase from Clostridium sticklandii; Davis JN et al.; A highly specialized phosphatase that depends on both a quinone (e.g., 2-methyl-1,4-napthoquinone) and a sulfhydryl compound for activity was purified to homogeneity from extracts of Clostridium sticklandii . Selective adsorption to Cibacron Blue-Sepharose 4B followed by elution with p-nitrophenylphosphate was an effective enrichment procedure . An affinity matrix containing vitamin K5 (4-amino-2-methyl-1-naphthol) covalently attached to Sepharose 4B selectively retained the enzyme and was also used in its purification . The only known substate for the enzyme, p-nitrophenylphosphate, is hydrolyzed to equivalent amounts of orthophosphate and p-nitrophenol . Although a protein phosphotyrosine residue seemed a likely candidate as the natural substrate, the enzyme failed to hydrolyze 32P-labeled phosphotyrosine residues in casein, in vinculin, or in denatured glutamine synthetase . Also, free O-phosphotyrosine and numerous phosphate esters that serve as substrates for common phosphomonoesterases were not hydrolyzed . The molecular weight of the native enzyme, estimated by Sephacryl-S-200 gel chromatography, is 27,600 . Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis showed a single component with a molecular weight of 28,600 . From the amino acid composition, a minimum molecular weight of 28,000 was calculated.

Zentralbl Bakteriol Mikrobiol Hyg {A}, 1985 May, 259(3), 307 - 16
{Differentiation of clostridia}; Niculescu ER et al.; 162 Clostridium strains (20 species), isolated from clinical specimens, were identified by their morphological, biochemical, and gaschromatographic characteristics . The reliability of the Api 20A system in identifying Clostridium species was studied by performing biochemical tests both in the microsystem and according to the VPI Anaerobic Handbook . 66% of the Clostridium strains could be identified by the Api 20A system supplemented, in accordance with the Api Handbook by other tests such as morphology, lipase and lecithinase production . The following Clostridium species, known as clinically most significant, could be identified by the Api 20A system: Clostridium sordellii and C . sporogenes (92%), C . bifermentans (60%) and C . tertium (78%) . Methods which can be used in the clinical laboratory for the identification of Clostridium species are described, i.e . isolation of Clostridium strains from mixed cultures, examination of morphological characteristics, performance of biochemical tests in the Api 20 A system as a micromethod, verification by conventional biochemical test systems and gaschromatographic analysis in special laboratories if necessary.

J Infect, 1985 May, 10(3), 252 - 5
Spontaneous pseudomembranous colitis not associated with Clostridium difficile; Dickinson RJ et al.; This paper describes two patients who developed macroscopically and microscopically typical pseudomembranous colitis without prior exposure to antimicrobial agents and without detectable Clostridium difficile or its toxin in the faeces.

Infection, 1985 May-Jun, 13(3), 97 - 101
Serum antibody response to Clostridium difficile toxins in patients with Clostridium difficile diarrhoea; Aronsson B et al.; Consecutive serum samples from 61 patients with Clostridium difficile diarrhoea were investigated for antibody response to C . difficile toxins A and B in an indirect enzyme immunoassay (ELISA) and in a neutralization assay against C . difficile cytotoxin . Sera from 64 blood donors, elderly healthy females and patients with other known intestinal enteropathogens served as controls . An immune response was detected by ELISA in approximately half of the patients with C . difficile diarrhoea . The specificity of the ELISA was 94% or 97%, depending on the control material used . Furthermore, a correlation was found between clinical recovery without relapse of C . difficile diarrhoea and high IgG titers to toxin B in the ELISA, and/or appearance of neutralizing antibodies . It is concluded that the ELISA for detection of serum antibodies to C . difficile toxins may be of diagnostic value in combination with the conventional tissue culture assay for cytotoxin in stool . High ELISA IgG titres to toxin B and/or the appearance of neutralizing antibodies may also be a positive prognostic sign in patients with C . difficile diarrhoea.

J Lipid Res, 1985 May, 26(5), 610 - 6
Phosphatidylserine decarboxylase from Clostridium butyricum; Verma JN et al.; Phosphatidylserine decarboxylase activity has been characterized in membrane preparations from Clostridium butyricum ATCC 19398 . A particulate fraction was shown to catalyze the formation of phosphatidylethanolamine and plasmenylethanolamine when vesicles containing phosphatidylserine and plasmenylserine were used as substrate . No plasmenylethanolamine was formed when phosphatidylserine alone was used as substrate . The activity with phosphatidylserine was activated by divalent cations and was optimal under anaerobic conditions . Ionic detergents inhibited phosphatidylethanolamine formation strongly and nonionic detergents inhibited partially . In the presence of Triton X-100, phosphate from {32P}phosphatidylserine appeared in three unidentified lipid products, in addition to phosphatidylethanolamine . The formation of these products was time- and Triton X-100 concentration-dependent . Hydroxylamine inhibited phosphatidylserine decarboxylase, but did not prevent the reactions stimulated by Triton X-100.

J Antibiot (Tokyo), 1985 May, 38(5), 555 - 60
Aridicins, novel glycopeptide antibiotics . I . Taxonomy, production and biological activity; Shearer MC et al.; A new species of a new genus of the Actinomycetales was discovered, Kibdelosporangium aridum . This strain produces a new family of glycopeptide antibiotics designated aridicins, that contain an unusual glycolipid constituent . They inhibit Gram-positive bacteria, including staphylococci, enterococci and Clostridium sp.

Antimicrob Agents Chemother, 1985 May, 27(5), 863 - 7
Role of the phosphoroclastic reaction of Clostridium pasteurianum in the reduction of metronidazole; Lockerby DL et al.; To demonstrate the importance of electron siphoning by the metronidazole reductase system from reduced ferredoxin to the mechanism of action of the drug in Clostridium pasteurianum, the effects of the reduction of metronidazole on the phosphoroclastic reaction were studied . Metronidazole concentrations between 0.5 and 5 mM caused a significant increase in acetyl phosphate production by the phosphoroclastic reaction compared to the control system without metronidazole . When this enzymatic reaction was assayed by standard manometric techniques under nitrogen gas, two simultaneous effects of electron siphoning were demonstrated: (i) the electrons from reduced ferredoxin were initially consumed for the reduction of metronidazole instead of being evolved as H2 via the ferredoxin-linked hydrogenase and (ii) phosphoroclastic activity was stimulated, with augmented production of CO2 and acetyl phosphate . This work further supports the notion of preferential scavenging of electrons away from ferredoxin-linked enzymatic reactions by metronidazole reductase(s) in C . pasteurianum.

Biol Chem Hoppe Seyler, 1985 May, 366(5), 463 - 72
Structure of enoate reductase from a Clostridium tyrobutyricum (C . spec . La1); Kuno S et al.; Enoate reductase from Clostridium tyrobutyricum was purified by a rapid novel procedure . Chromatography on DEAE-Sepharose and on hydroxyapatite resulted in a high yield of about 90% pure enzyme in less than 10 h . A purity greater than 98% could be obtained by additional chromatography on Sephacryl S-300 . The enzyme sediments in the analytical ultracentrifuge as a single, symmetrical boundary with a velocity of S(0)20,w = 24.9 S . Equilibrium ultracentrifugation yielded a molecular mass of 940 000 +/- 20 000 Da . The enzyme contains one type of subunit as shown by dodecyl sulfate electrophoresis and partial sequence determination . A subunit molecular mass of about 73 000 Da was established by dodecyl sulfate electrophoresis and by sedimentation equilibrium analysis in guanidine hydrochloride . In addition to FAD, iron and labile sulfur, the enzyme purified by the new method showed approximately 0.7 mol of FMN per mol of subunit . A dissociation product sedimenting at a velocity of S(0)20,w = 9.8 S can be obtained by various experimental protocols . The fragment was obtained in pure form by gel permeation chromatography . The molecular mass was 230 000 +/- 10 000 Da as shown by sedimentation equilibrium analysis . Thus it appears that the dissociation product is a trimer of the 73 000-Da subunit . The formation of the 10-S fragment by dissociation of the native enzyme is accompanied by the loss of most of the FMN, whereas the FAD content is not changed . The fragment catalysed the reduction of acetylpyridine adenine dinucleotide by NADH . However, enoate reductase activity with NADH or methylviologen as cosubstrate was low . Electron micrographs of negatively stained enoate reductase show trigonal symmetry . The data suggest that enoate reductase is a dodecamer (tetramer of trimers) with tetrahedral symmetry.

Am Fam Physician, 1985 May, 31(5), 115 - 20
Antibiotic-associated pseudomembranous colitis; Amin NM; All antibiotics, except parenteral aminoglycosides, sulfonamides and vancomycin, can induce pseudomembranous colitis . The worst offenders are clindamycin, ampicillin, amoxicillin and the cephalosporins . The cytotoxin produced by Clostridium difficile has been identified as the cause of pseudomembranous colitis . Treatment includes an antimicrobial to eradicate the organism and cholestyramine or colestipol to bind the toxin.

Infect Immun, 1985 May, 48(2), 312 - 7
Purification and characterization of neurotoxin produced by Clostridium botulinum type C 6813; Terajima J et al.; The toxin produced by Clostridium botulinum type C 6813 (C-6813) was purified 1,009-fold from the culture supernatant in an overall yield of 30% . The specific toxicity was 1.1 X 10(7) mouse minimum lethal doses per mg of protein . The toxin had a molecular weight of 144,000, composed of the light and heavy chains with molecular weights of 52,000 and 92,000, respectively, linked by one or two disulfide bond(s) . The purified C-6813 toxin heavy and light chains reacted strongly with anti-type D heavy chain immunoglobulin G and anti-type C1 light chain immunoglobulin G, respectively . The amino acid compositions of C-6813 toxin heavy and light chains were more similar to those of type D heavy chain and type C1 light chain than to those of type C1 heavy chain and type D light chain, respectively . These results suggest that in the toxin produced by the type C strain at least two subtypes exist.

Ann Intern Med, 1985 May, 102(5), 616 - 8
Botulism and botulism-like illness in chronic drug abusers; MacDonald KL et al.; From 1982 to 1983 we received reports of a neurologic illness characterized by a symmetric descending paralysis in six drug abusers from widely separated geographic areas . Botulism was confirmed in two patients; type B botulinal toxin was found and Clostridium botulinum was isolated from a small abscess in one, and type A botulinal toxin was found in the serum of the other . The clinical illness in the remaining four patients, although not laboratory confirmed, was also compatible with botulism . None of the patients had histories suggestive of foodborne botulism, and wound botulism was suspected as the cause of illness . There are several reports of tetanus associated with parenteral drug abuse; wound botulism is another toxin-mediated clostridial infection that may occur as a complication of chronic drug abuse.

Clin Pharm, 1985 May-Jun, 4(3), 304 - 10
Management of antibiotic-associated pseudomembranous colitis; Gross MH; The diagnosis, etiology, epidemiology, and drug therapy of antibiotic-associated pseudomembranous colitis (AAPMC) are reviewed . AAPMC is an uncommon but potentially serious adverse reaction to therapy with almost any oral or injectable antibiotic and certain antineoplastic agents that alter intestinal flora . Proliferation of Clostridium difficile and subsequent release of clostridial cytotoxins cause pseudomembranous lesions and symptoms such as watery diarrhea, cramping abdominal pain, and low-grade fever . Symptoms can appear from four days after the start of antibiotic or antineoplastic therapy to 10 weeks after therapy has been discontinued . Drug therapy of AAPMC is directed at reducing the amount of Cl . difficile in the colon and promoting normalization of intestinal flora . Mild cases of AAPMC may respond to discontinuation of the etiologic agent and replacement of fluid and electrolytes . Therapy with an anticlostridial antibiotic is indicated in severe cases; although a seven- to 10-day course of oral vancomycin hydrochloride is the most widely recognized therapy, the drug is expensive and unpalatable . Good results have been reported with oral metronidazole and with bacitracin, both of which are less expensive than vancomycin . For all of these therapies, relapse rates are 20-39% . Anion exchange resins may be useful in mild cases of AAPMC . Successful management of AAPMC depends on a complex and ill-defined interrelationship between normal intestinal flora, patient immune response, antibiotic therapy, and the infecting clostridium strain . For moderate or severe cases of AAPMC, therapy should begin with metronidazole or bacitracin and vancomycin should be reserved for refractory cases, relapses, or patients with allergies to the other agents.

Pediatr Infect Dis, 1985 May-Jun, 4(3), 246 - 9
Increased risk of illness among nursery staff caring for neonates with necrotizing enterocolitis; Gerber AR et al.; In 1983 an outbreak of necrotizing enterocolitis (NEC) and hemorrhagic gastroenteritis occurred in our newborn nurseries . Eleven children were ill and three required bowel resections . During the outbreak many of the medical and nursing staff in the nurseries also were ill, prompting a microbiologic and epidemiologic investigation . Bacterial and viral cultures, Clostridium difficile toxin assays, enzyme-linked immunosorbent assays for viral antigens and immunoelectron microscopy of stools identified no associated pathogen . However, using a method of calculating relative risk as an incidence density ratio, we found that nurses who had cared for ill infants were at higher risk for sick call within the 9 days following exposure than nurses who had cared for babies without NEC (relative risk, 1.96; P = 0.05) . These results provide additional evidence that a transmissible agent may be responsible for some cases of NEC and support the recommendation for infection control measures during outbreaks . The epidemiologic methods used in this study may be useful in prospective studies of NEC and may help to provide further clues to the cause of this disease.

Diagn Microbiol Infect Dis, 1985 May, 3(3), 193 - 200
Postmortem bacteriology of cadaver tissue donors: an evaluation of blood cultures as an index of tissue sterility; Martinez OV et al.; Microbiological cultures were performed on the blood and bone marrow of 239 cadaver bone donors and 58 "beating heart cadaver" organ donors who had been asymptomatic of sepsis . The incidence of positive blood cultures was significantly lower among the "beating heart cadaver" donors (8.6%) as compared to other donors from whom tissues were excised up to 30 hr postmortem (38%) . Microorganisms were isolated from the bones of 82 of 148 (55.4%) bone donors as well as from 36 of 53 (67.9%) "beating heart cadaver" donors who had negative blood cultures . The majority of microbial species recovered from the blood and bone marrow belonged to species normal to the skin microflora (coagulase-negative staphylococci, Bacillus and Propionibacterium species) . Species of Clostridium were the second most common organisms isolated from the blood . Blood cultures alone were not useful as indicators of sepsis in cadaver tissue donors or as an index of the sterility of the tissues excised for transplantation.

Arch Biochem Biophys, 1985 May 1, 238(2), 544 - 8
Partial amino acid sequences of botulinum neurotoxins types B and E; Schmidt JJ et al.; Clostridium botulinum type E neurotoxin, a single-chain protein of Mr 147,000, was purified and subjected to amino acid sequencing . The same was done for single-chain botulinum type B neurotoxin (Mr 152,000), and for the heavy and light chains (Mr 104,000 and 51,000 respectively) derived from type B by limited trypsin digestion . Twelve to eighteen residues were identified and the following conclusions were drawn: The light chain of the nicked (dichain) type B is derived from the N-terminal one-third of the single-chain (unnicked) parent neurotoxin; sequence homologies are present between single-chain types B and E and the light chain of the nicked type A {J . J . Schmidt, V . Sathyamoorthy, and B . R . DasGupta (1984) Biochem . Biophys . Res . Commun . 119, 900-904}; the N-terminal regions of the heavy chains of types A and B have some structural similarity; and activation of type B neurotoxin cannot involve removal of amino acids or peptides from the N terminus.

J Antibiot (Tokyo), 1985 May, 38(5), 649 - 60
Antibacterial activity of cefminox against anaerobes; Watanabe K et al.; The antibacterial activity of cefminox (CMNX) against anaerobic bacteria was studied in vitro . The results are as follows: 1 . CMNX exerted antibacterial activity against a wide range of anaerobes, excluding Clostridium innocuum . The antibacterial activity of CMNX against Bacteroides fragilis was comparable to that of latamoxef and superior to cefoxitin, but CMNX's activity against anaerobic cocci was slightly inferior to cefoxitin's; 2 . A comparison of the MICs and MBCs of CMNX indicated that this drug exerts a complete bactericidal effect at a concentration which inhibits the growth of bacteria; 3 . CMNX was found to be stable to the beta-lactamases produced by B . fragilis; 4 . CMNX exerted an antibacterial activity against C . difficile.

Antimicrob Agents Chemother, 1985 May, 27(5), 745 - 8
Safety and efficacy of high-dose treatment with imipenem-cilastatin in seriously ill patients; Zajac BA et al.; Imipenem-cilastatin was given in doses of 1 g intravenously every 6 h to 31 patients . Twenty-five patients, with 27 infections, were clinically evaluable and received 20 to 210 g of imipenem for a duration of 5 to 56 days (average 16.3 days) . Infections included seven cases of osteomyelitis, seven of bacteremia, five of cellulitis, two of pneumonia, three of pelvic cellulitis, two of intraabdominal abscess, and one each of empyema, mediastinitis, and endometritis . Fifty-five percent of the infections were caused by gram-negative bacilli, 33% were due to gram-positive organisms, and 10% were caused by anaerobes . Twenty-two patients (81%) were cured, three improved, one relapsed, and one became superinfected with a resistant organism . In 5 of 11 cases with Pseudomonas aeruginosa, the imipenem MIC for organisms isolated by the end of treatment was higher than it was initially, raising concern that imipenem should not be used alone to treat Pseudomonas aeruginosa infections . Twenty-one patients had no adverse reaction; of the remaining 10 patients, 4 had nausea, 1 had urticaria, and 6 had mild abnormalities in hepatic function; three episodes of diarrhea included two with Clostridium difficile toxin in stool and one with pseudomembranous colitis, as determined by sigmoidoscopy . Levels of creatinine, hemoglobin, leukocytes, platelets, prothrombin, and urine components were unchanged . Imipenem-cilastatin is a clinically effective antibiotic with freedom from nephrotoxicity and hematological abnormalities in the large doses used in this study.

J Bacteriol, 1985 May, 162(2), 485 - 93
Regulation and order of involvement of molybdoproteins during synthesis of molybdoenzymes in Clostridium pasteurianum; Hinton SM et al.; The accumulation of 99Mo (from 99MoO4(2-) into molybdenum-containing species in Clostridium pasteurianum was investigated to identify the molybdoprotein(s) involved in Mo metabolism . Mo accumulation by clostridial cells during the derepression of the nitrogenase system increased substantially beginning 1.5 h before nitrogenase activity was detected . The increase in Mo accumulation by the cells is a result of the incorporation of Mo into a high-molecular-weight molybdenum species (suspected membrane fragments), a low-molecular-weight molybdenum species, a Mo binding-storage protein, a 30-kilodalton molybdoprotein, and formate dehydrogenase . Mo incorporation into the MoFe protein was detected 1 h after the onset of metal uptake . Kinetics of Mo accumulation into the molybdoproteins during the derepression of nitrogenase suggests that Mo incorporation or uptake or both occur in the following sequence: (i) membranes and MoO4(2-), (ii) a low-molecular-weight molybdenum species, (iii) Mo binding-storage protein and a 30-kilodalton molybdoprotein, (iv) formate dehydrogenase, and (v) the MoFe protein . The intracellular level of all molybdenum components except the MoFe protein appears to be influenced by the availability of Mo . Clostridial cells grown in the presence of a limiting amount of Mo became Mo deficient as a result of growth and a MoO4(2-) supplement added to such cells rapidly accumulated within the cells to levels five times that found in steady-state nitrogen-fixing cells . The Mo accumulated by the Mo-deficient cells was rapidly incorporated into preformed demolybdoproteins in the absence of de novo protein synthesis . The increase in Mo accumulation by Mo-deficient cells was a result of an increase in all molybdoproteins except the MoFe protein.

J Bacteriol, 1985 May, 162(2), 477 - 84
Identification of molybdoproteins in Clostridium pasteurianum; Hinton SM et al.; Cells of Clostridium pasteurianum whose N source is switched from NH3 to N2 accumulate large amounts of molybdenum beginning 1.5 h before the detection of nitrogenase activity . Anaerobic multiphasic gel electrophoresis and anion-exchange chromatography were used to identify the molybdoproteins and molybdenum-containing components present in N2-fixing cells . In addition to molybdate, six distinct 99Mo-labeled species were detected, i.e., a membrane fragment, the MoFe protein of nitrogenase, formate dehydrogenase, a Mo "binding-storage" protein, a 30-kilodalton molybdoprotein, and a low-molecular-weight molybdenum species . Of these, the MoFe protein, formate dehydrogenase, and the Mo binding-storage protein were present in more than one zone because of complex formation with other proteins, partial denaturation, and variation in the amount of Mo bound to the protein, respectively . In addition to the six proteins, a soluble "free" Mo cofactor in the cytosol was detected by showing that it reconstituted nitrate reductase activity in crude extracts of the Neurospora crassa mutant nit-1.

Clin Pediatr (Phila), 1985 May, 24(5), 252 - 5
Clostridium difficile isolation in leukemic children on maintenance cancer chemotherapy . A preliminary study; Chiesa C et al.; Between December 1982 and November 1983, stool specimens from 15 children with acute lymphoblastic leukemia, who were on maintenance cancer chemotherapy, were examined weekly for the presence of Clostridium difficile and its toxin . Four out of 15 patients were positive for C . difficile: three patients had stool specimens that did not contain toxin, but cultures yielded growth of toxigenic C . difficile on only one occasion . The fourth patient, who had a recent history of hospitalization, particularly aggressive cancer chemotherapy, neutropenia, and antibiotic therapy, excreted both C . difficile and its toxin for at least 1 month . All children were asymptomatic at the time of positive cultures . This preliminary study reveals a low rate of C . difficile colonization in leukemic children on maintenance cancer chemotherapy.

Diagn Microbiol Infect Dis, 1985 May, 3(3), 263 - 8
BMY28142, cefbuperazone (T-1982), and Sch 34343 . Antimicrobial activity against 94 anaerobes compared to seven other antimicrobial agents; Jones RN et al.; Three new beta-lactams were evaluated against 94 anaerobic strains representing 15 species using a Wilkins-Chalgren broth microdilution method . The penems, Sch 29482 and Sch 34343, were most active with all minimum inhibitory concentrations (MICs) at less than or equal to 4.0 micrograms/ml and MIC90s of less than or equal to 0.25 micrograms/ml . BMY 28142 had a more limited antianaerobic activity against Bacteroides fragilis with a MIC50 and MIC90 of 32 and 128 micrograms/ml, respectively . Cefbuperazone (T-1982) had low B . fragilis MICs (MIC90, 8.0 micrograms/ml), but potentially resistant range MIC90 results for the other species in the B . fragilis group and Clostridium species.

Pathol Biol (Paris), 1985 May, 33(5), 421 - 5
{Sensitivity of obligate anaerobes to ofloxacin, pefloxacin, enoxacin and norfloxacin}; Dubreuil L et al.; The in vitro activity of four new quinolones against 355 obligate anaerobes was investigated . MICs were determined using the reference method of Sutter et al . {14} . The four fluorinated quinolones tested differ from nalidixic acid which is inactive on most anaerobes except for some Clostridium perfringens strains . All strains tested were inhibited by 64 and 16 mg/l pefloxacin and ofloxacin respectively . Except for Bacteroides, all strains were inhibited by 64 mg/l norfloxacin or enoxacin . Clostridium strains other than C . perfringens exhibited heterogeneity; C . difficile and C . ramosum had the highest MICs . Bacteroides fragilis proved less susceptible to fluorinated quinolones than the other obligate anaerobes . Ofloxacin showed the greatest activity, with 4 mg/l inhibiting 100% of C . perfringens and 83% of all anaerobes investigated.

Infect Immun, 1985 May, 48(2), 331 - 5
Tracheobronchial mucin receptor for Pseudomonas aeruginosa: predominance of amino sugars in binding sites; Vishwanath S et al.; Pseudomonas aeruginosa, a common respiratory tract colonizer and pathogen, adheres to injured tracheal cells and to tracheobronchial mucin . These phenomena suggest that there are specific receptors for this organism in the respiratory tract . The receptor on injured tracheal cells contains n-acetylneuraminic acid as the principal sugar, but the structure of the receptor in mucin has not been described . Using a microtiter plate assay to study bacterial adherence to mucin, we have partially characterized the mucin receptor for P . aeruginosa . The receptor for both nonmucoid and mucoid strains is sensitive to periodate oxidation, suggesting that it is carbohydrate in nature, and the amino sugars n-acetylglucosamine and n-acetylneuraminic acid inhibited the adherence of both types of strains . Nonmucoid strains were more sensitive to inhibition by n-acetylneuraminic acid than to inhibition by n-acetylglucosamine, but the mucoid strains varied in their sensitivities to inhibition by each amino sugar . Preincubation of mucin with heat-inactivated influenza A virus (which binds to neuraminic acid) significantly reduced the adherence of P . aeruginosa . Treatment of mucin with Clostridium perfringens neuraminidase also reduced bacterial adherence significantly . Treatment of mucin with pronase did not affect adherence . Our results suggest that n-acetylglucosamine and n-acetylneuraminic acid are important constituents of the binding sites for P . aeruginosa on human tracheobronchial mucin.

Prikl Biokhim Mikrobiol, 1985 May-Jun, 21(3), 365 - 71
{Synthesis and properties of an affinity adsorbent for purifying neuraminidases of varying origins}; Poltorak AN et al.; Neuraminidases (NA) from Clostridium perfringens, noncholera vibrios and influenza virus were purified by affinity chromatography on Sepharose coupled to para-aminophenyl oxamic acid . Adsorption was carried out at pH 5.5 . The effect of elution conditions on purification of NA was studied . The use of the pH gradient enhances 10-fold the purification degree as compared with the pH shift-elution process, 90% of the activity being eluted from the column within a pH range from 6.0 to 6.6 . According to the described procedure, electrophoretically homogeneous preparations of NA were obtained from noncholera vibrios and influenza virus.

Plasmid, 1985 May, 13(3), 155 - 62
Cloning and analysis of the Clostridium perfringens tetracycline resistance plasmid, pCW3; Abraham LJ et al.; Clostridium perfringens strain CW92 carries pCW3, a conjugative 47-kb plasmid that confers inducible resistance to tetracycline . The plasmid was examined by restriction endonuclease analysis and by cloning each of the five ClaI fragments of pCW3 in Escherichia coli, using pBR322 . Analysis of the recombinant plasmids allowed the deduction of a detailed restriction map of pCW3 . The tetracycline resistance determinant of pCW3 was mapped by examining the phenotype of recombinant E . coli clones derived from the cloning, into pUC vector plasmids, of EcoRI fragments from pCW3 . The C . perfringens tetracycline resistance determinant was expressed in E . coli and was shown to be located on two juxtaposed EcoRI fragments which together encompass a 4-kb region of pCW3 . Deletion experiments showed that the tetracycline resistance gene, and/or its control regions, contained internal EcoRI and SphI sites . E . coli strains that carried recombinant plasmids with only the 4-kb region were found to express tetracycline resistance constitutively . In contrast, recombinant plasmids harboring a 10.5-kb ClaI fragment of pCW3, that included the 4-kb region, coded for an inducible tetracycline resistance phenotype . The existence of a negatively regulated resistance gene, similar to that proposed for several other bacteria is postulated.

Biochem Biophys Res Commun, 1985 Apr 30, 128(2), 760 - 6
Role of one tryptophan residue in the lethal activity of Clostridium perfringens epsilon toxin; Sakurai J et al.; The lethal activity of Clostridium perfringens epsilon toxin was inactivated by N-bromosuccinimide and N-chlorosuccinimide . Amino acid analysis of N-bromosuccinimide-treated prototoxin indicated that the one tryptophan residue present in the protein was abolished, and methionine and tyrosine reduced markedly . N-chloro-succinimide-treated prototoxin lost completely both tryptophan and methionine residues . The toxin was not inactivated by chloramine T, but all of methionine residues present in the protein was found to be oxidized by the agent . The data suggest that the one tryptophan residue present in the toxin is important for the lethal activity.

Vet Rec, 1985 Apr 27, 116(17), 467 - 9
Vaccination of cows with clostridial antigens and passive transfer of clostridial antibodies from bovine colostrum to lambs; Clarkson MJ et al.; Two Friesian cows were used to attempt to produce colostrum containing a high concentration of clostridial antibodies which could be fed to newborn lambs in order to passively transfer immunity to diseases caused by clostridia . One cow was given a commercial multicomponent clostridial sheep vaccine in two successive pregnancies and the second cow in one pregnancy . The first cow produced a low concentration of epsilon antitoxin (Clostridium perfringens, type D) in its blood and colostrum after the first course of three injections of vaccine . A higher concentration was produced by cow 2 after a course of six injections and by cow 1 after a further course of four injections in its next pregnancy . Two hundred ml of colostrum from cow 1 (after the second course of vaccine) was given to 12 newborn colostrum-deprived lambs . All showed a high concentration of antitoxin 48 hours later . The lambs were actively immunised by injections of the same clostridial vaccine at three and nine weeks or six and 12 weeks old and all produced sufficient antitoxin to protect up to slaughter at 24 weeks . It is concluded that colostrum from cows vaccinated with sheep clostridial antigens can be fed to protect lambs passively.

J Biol Chem, 1985 Apr 10, 260(7), 3970 - 7
Acetate biosynthesis by acetogenic bacteria . Evidence that carbon monoxide dehydrogenase is the condensing enzyme that catalyzes the final steps of the synthesis; Ragsdale SW et al.; The purified carbon monoxide dehydrogenase from Clostridium thermoaceticum is the only protein required to catalyze an exchange reaction between carbon monoxide and the carbonyl group of acetyl-CoA . This exchange requires that the CO dehydrogenase bind the methyl, the carbonyl, and the CoA groups of acetyl-CoA, then equilibrate the carbonyl with CO in the solution and re-form acetyl-CoA . CoA is not necessary for the exchange and, in fact, inhibits the reaction . These studies support the view that CO dehydrogenase is the condensing enzyme that forms acetyl-CoA from its component parts . Carbon dioxide also exchanges with the C-1 of acetyl-CoA, but at a much lower rate than does CO . At 50 degrees C and pH 5.3, the optimal pH, the turnover number is 70 mol of CO exchanged per min/mol of enzyme . Low potential electron carriers are stimulatory . The Km app for stimulation by ferredoxin is 50-fold less than the value for flavodoxin . Neither ATP or Pi stimulate the exchange . The EPR spectrum of the CO-reacted enzyme is markedly changed by binding of CoA or acetyl-CoA . Arginine residues of the CO dehydrogenase appear to be involved in the active site, possibly by binding acetyl-CoA . Mersalyl acid, methyl iodide, 5,5-dithiobis-(2-nitrobenzoate), and sodium dithionite inhibit the exchange reaction . A scheme is presented to account for the role of CO dehydrogenase in the exchange reaction and in the synthesis of acetate.

Can J Comp Med, 1985 Apr, 49(2), 145 - 8
Clinical and antibody responses to Clostridium perfringens type A enterotoxin in experimental sheep and calves; Niilo L et al.; Clostridium perfringens type A live cultures or sonicated sporulating cells, all containing enterotoxin, were repeatedly inoculated into sheep and calves by the intraduodenal route over periods of 30 to 35 days . Serum antibody to C . perfringens enterotoxin, tested by ELISA, developed in four of seven sheep and in two of four calves . The titers ranged from 400 to 1600 . The live organism introduced into the duodenum did not become established in the bacterial flora of the intestinal tract.

Eur J Clin Microbiol, 1985 Apr, 4(2), 102 - 7
Enzyme immunoassay for detection of Clostridium difficile toxins A and B in patients with antibiotic-associated diarrhoea and colitis; Aronsson B et al.; A sandwich enzyme immunoassay (ELISA) was developed to detect Clostridium difficile toxins A and B in stools from patients with antibiotic associated diarrhoea and colitis . Immune serum to crude Clostridium difficile toxin and non-immune serum were coated onto polystyrene microtiter plates to act as capture antibodies; toxins A and B in human stools were detected by antibodies from rabbits immunized with purified toxins A and B . The ELISA for toxin B showed cross-reactions with Clostridium bifermentans and Clostridium sordellii and lacked diagnostic sensitivity in clinical samples . The ELISA for toxin A showed no cross-reactions with other clostridiae investigated and was positive in 33% (62/189) of patients with antibiotic associated diarrhoea . This compared with 38% (71/189) positive in the tissue culture assay for Clostridium difficile cytotoxin . With a predictive value of 96% in clinical specimens, the ELISA for toxin A constitutes a sensitive and specific tool for diagnosis of Clostridium difficile associated diarrhoea and colitis.

Appl Environ Microbiol, 1985 Apr, 49(4), 874 - 8
Regulation and butanol inhibition of D-xylose and D-glucose uptake in Clostridium acetobutylicum; Ounine K et al.; Clostridium acetobutylicum exhibited diauxie growth in the presence of mixtures of glucose and xylose . Both glucose- and xylose-grown cells had a glucose uptake activity . On the other hand, growth on xylose was associated with the induction of a xylose permease activity, which was repressed by glucose in xylose-induced cells . The rate of sugar uptake with increasing sugar concentrations showed saturation kinetics with an apparent Km of 1.25 X 10(-5) M for glucose and 5 X 10(-3) M for xylose . Concomitant with the production of solvents, the activities of the glucose and xylose transport systems decreased . Among the main products of fermentation, butanol was shown to be a potent inhibitor of the growth of the organism and of the rate of sugar uptake as well as of sugar incorporation into cell materials . These inhibitory effects of butanol were more pronounced in xylose-grown cells than in glucose-grown cells . Butanol completely inhibited growth at a concentration of 14 g/liter for cultures growing on glucose and 8 g/liter for cultures growing on xylose . Concentrations of 7 and 10.5 g/liter of butanol caused a 50% inhibition of the xylose and glucose incorporations into cell materials . These inhibitory levels of butanol were found in typical glucose or xylose fermentation.

Appl Environ Microbiol, 1985 Apr, 49(4), 733 - 6
Influence of pH on organic acid production by Clostridium sporogenes in test tube and fermentor cultures; Montville TJ et al.; The influence of pH on the growth parameters of and the organic acids produced by Clostridium sporogenes 3121 cultured in test tubes and fermentors at 35 degrees C was examined . Specific growth rates in the fermentor maintained at a constant pH ranged from 0.20 h-1 at pH 5.00 to 0.86 h-1 at pH 6.50 . Acetic acid was the primary organic acid in supernatants of 24-h cultures; total organic acid levels were 2.0 to 22.0 mumol/ml . Supernatants from pH 5.00 and 5.50 cultures had total organic acid levels less than one-third of those found at pH 6.00 to 7.00 . The specific growth rates of the test tube cultures ranged from 0.51 h-1 at pH 5.00 to 0.95 h-1 at pH 6.50 . The pH of the medium did not affect the average total organic acid content (51.5 mumol/ml) but did affect the distribution of the organic acids, which included formic, acetic, propionic, butyric, 3-(p-hydroxyphenyl)propionic, and 3-phenylpropionic acids . Butyric acid levels were lower, but formic and propionic acid levels were higher, at pH 5.00 than at other pHs.

Antimicrob Agents Chemother, 1985 Apr, 27(4), 640 - 2
In vitro activity of cefbuperazone, a new cephamycin, against anaerobic bacteria; Prabhala RH et al.; The 90% MIC of cefbuperazone (BMY 25182) was 32 micrograms/ml for Bacteroides fragilis and Bacteroides spp., 128 micrograms/ml for Fusobacterium and Clostridium spp., 64 micrograms/ml for Eubacterium and Peptococcus spp., 8 micrograms/ml for Actinomyces spp., and 32 micrograms/ml for Peptostreptococcus spp . The level of activity of cefbuperazone was higher against B . fragilis and lower against anaerobic cocci than those of related cephalosporins, i.e., cefoxitin, cefoperazone, cefotaxime, ceftizoxime, and cefmenoxime . However, the activity of cefbuperazone was comparable to that of moxalactam against all groups tested . Size of inoculum and type of media used did not alter the MICs of cefbuperazone for B . fragilis . Cefbuperazone showed synergistic activity when combined with cefoxitin against resistant strains of B . fragilis.

J Appl Bacteriol, 1985 Apr, 58(4), 363 - 9
Nitrogen metabolism by the microbial flora of the rabbit caecum; Forsythe SJ et al.; The dense microbial flora of the rabbit caecum consisted chiefly of bacteria (10(11)/g) with small numbers of yeast cells (10(6)/g) . Using strictly anaerobic technique, 23% of the direct microscopic cell count was cultivated and 55% of the cultivatable bacteria utilized ammonia as the sole source of nitrogen . Ureolytic bacteria were isolated from the caecal lumen and mucosa and were identified as Bacteroides vulgatus, Clostridium clostridiiforme, Bacillus spp . and Staphylococcus spp . Ammonia assimilation by the bacterial flora of the caecum was by incorporation into alpha-oxoglutarate catalysed by NADPH-linked glutamate dehydrogenase.

J Clin Microbiol, 1985 Apr, 21(4), 654 - 5
Isolation of an organism resembling Clostridium barati which produces type F botulinal toxin from an infant with botulism; Hall JD et al.; All reported cases of infant botulism except one have been caused by proteolytic strains (group I) of Clostridium botulinum, toxin types A or B . We describe the cultural and biochemical characteristics of the causative organism of this singular case of infant botulism, caused by type F botulinal toxin . Although this organism produces type F botulinal toxin, it is quite different from proteolytic (group I) C . botulinum, being more closely related to Clostridium barati.

J Clin Microbiol, 1985 Apr, 21(4), 636 - 7
Effect of adding sodium taurocholate to selective media on the recovery of Clostridium difficile from environmental surfaces; Buggy BP et al.; The recovery of Clostridium difficile on a medium containing cefoxitin, cycloserine, fructose, and egg yolk was compared with that on media containing one of three preparations of sodium taurocholate . In aerobic environments contaminated with C . difficile, media containing either crude taurocholate from Mann Research Laboratories, New York, N.Y., or pure taurocholate from Sigma Chemical Co., St . Louis, Mo., recovered organisms significantly more often than did cefoxitin-cycloserine-fructose-egg yolk agar.

Aust Vet J, 1985 Apr, 62(4), 112 - 4
Comparison of immune response stimulated in sheep, rabbits and guinea pigs by the administration of multi-component clostridial vaccines; Webster AC et al.; Over 3 years, the immunogenic responses of various batches of multi-component clostridial vaccines in sheep, rabbits and guinea pigs were compared . Fully susceptible healthy sheep were found to be more suitable than rabbits or guinea pigs for testing the potency of multi-component clostridial vaccines containing Clostridium novyi type B, C . perfringens type D, C . septicum and C . tetani, and recommendations are made that sheep are the preferred species for testing the potency of clostridial vaccines.

Appl Environ Microbiol, 1985 Apr, 49(4), 939 - 43
Rapid, simplified method for production and purification of tetanus toxin; Ozutsumi K et al.; A rapid, simplified method for production and purification of tetanus toxin from bacterial extracts was described . The extracts were prepared by stirring young cells (ca . 45-h culture) of Clostridium tetani in 1 M NaCl-0.1 M sodium citrate, pH 7.5, overnight at 0 to 4 degrees C . The toxin was purified by a combination of (i) ammonium sulfate fractionation (0 to 40% saturation), (ii) ultracentrifugation for removal of particulate materials, and (iii) gel filtration by high-pressure liquid chromatography on a TSK G3000 SW-type column . This method required 6 days as follows: (i) overnight incubation of the seed culture, (ii) 2 days for growing the bacteria for toxin production, (iii) overnight extraction of the toxin from the bacteria, (iv) overnight precipitation of the toxin with ammonium sulfate, (v) 2 h for ultracentrifugation of the ammonium sulfate concentrate of the bacterial extract, and (vi) 1 h for high-pressure liquid chromatography . The minimum lethal dose of the purified toxin preparations for mice was 1.4 X 10(7) to 1.5 X 10(7) per mg of protein and they showed 360 to 390 Lf (flocculating activity) per mg protein and a 280/260 nm absorbance ratio of 2.0 to 2.1 . The final recovery of the toxin from bacterial extracts was 90 to 93% . The purified preparations gave a single band of toxin protein with a molecular weight of 150,000 +/- 5,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis . On crossed immunoelectrophoresis, the purified toxin preparations gave a single precipitation arc against anti-crude toxin serum.

Scand J Gastroenterol, 1985 Apr, 20(3), 373 - 80
Short-chain fatty acids in intestinal content of germfree mice monocontaminated with Escherichia coli or Clostridium difficile; Hoverstad T et al.; The short-chain fatty acids (SCFAs) have been analysed in coecal and small-intestinal content of conventional (CONV) and germfree (GF) mice, in germfree mice monocontaminated with Escherichia coli (MEC) or Clostridium difficile (MCD), and in germfree mice conventionalized by the visitor technique (EXG) . The total concentrations of SCFAs in coecal content, measured by gas chromatography, were (mean (SD), mmol/kg): CONV, 125.2 (32.9); GF, 1.02 (0.39); MEC, 6.88 (0.76); MCD, 4.50 (0.12); and EXG, 115.6 (17.4) . The concentrations of SCFAs were 3- to 25-fold higher in the coecum than in the small intestine in CONV, MEC, and EXG mice (p less than 0.05) . The fermentation patterns (that is, the relative composition of the acids) of E . coli and Cl . difficile were distinctively different under defined in vitro conditions and similar to those found in intestinal content of monocontaminated animals . Combined gas chromatography and mass spectrometry showed that Cl . difficile produced an unusual metabolite, 2-methylbutyric acid, in vitro and in vivo . The findings indicate that the fermentation patterns are closely related to the bacteria . Variations in the bacterial flora may be more important in determining the concentrations and patterns of SCFAs in intestinal content than variations in the intake of substrate for SCFAs formation as dietary fibre.

J Clin Microbiol, 1985 Apr, 21(4), 599 - 606
Studies of metabolites in diarrheal stool specimens containing Shigella species by frequency-pulsed electron capture gas-liquid chromatography; Brooks JB et al.; Eleven diarrheal stool specimens and 10 control stool specimens from Cairo, Egypt, were studied by frequency-pulsed electron capture gas-liquid chromatography (FPEC-GLC) . Four cases involving Shigella sonnei, three cases involving Shigella boydii, and four cases involving Shigella flexneri were studied . The aqueous stools were centrifuged, extracted with organic solvents, and derivatized to form specific electron-capturing derivatives of carboxylic acids, alcohols, hydroxy acids, and amines . Analyses were performed on high-resolution glass columns with an instrument equipped with an extremely sensitive electron capture detector that is specific for the detection of electron-capturing compounds . The diarrheal stools studied had specific FPEC-GLC profiles and contained metabolic markers that readily distinguished between the Shigella spp . studied and Escherichia coli producing heat-stable or heat-labile enterotoxins . S . sonnei stools contained hexanoic acid, 2-hydroxy-4-methylmethiobutyric acid, and some unidentified alcohols that distinguished this organism from other enteric pathogens . S . boydii produced an acid that was unique for this species, and S . flexneri produced alcohols that could be used to distinguish between it and other enteric organisms . The FPEC-GLC profiles obtained during this study were also very different from those reported earlier for Clostridium difficile and rotavirus . This study presents further evidence that the selectivity and sensitivity of FPEC-GLC techniques can be used to rapidly identify causative agents of diarrhea and detect physiological changes that occur in the gut during the course of diarrheal illness.

Antimicrob Agents Chemother, 1985 Apr, 27(4), 674 - 6
In vitro activity of cefbuperazone against anaerobic bacteria; Wexler H et al.; The in vitro activity of cefbuperazone was compared with that of cefoxitin, moxalactam, and piperacillin against 305 strains of anaerobic bacteria . Piperacillin was the most active overall, inhibiting 97% of all anaerobes tested at 128 micrograms/ml . Cefbuperazone had poor activity against the Bacteroides fragilis group and Clostridium difficile (43 and 0% susceptible, respectively) but good activity (90.5%) against all other anaerobic bacterial species tested.

Antimicrob Agents Chemother, 1985 Apr, 27(4), 657 - 9
Comparative activity of the quinolones against anaerobic bacteria isolated at community hospitals; Goldstein EJ et al.; The in vitro activity of five quinolone compounds, amoxicillin, and clindamycin against 118 strains of anaerobic bacteria isolated at community hospitals was determined by an agar dilution method . Nalidixic acid and cinoxacin had poor activity, and norfloxacin and enoxacin showed relatively poor activity . Ciprofloxacin was active against Bacteroides fragilis, Fusobacterium species, Clostridium perfringens, and gram-positive cocci . At peak levels achievable in the feces, norfloxacin and enoxacin had moderate activity.

Mol Biol Rep, 1985 Apr, 10(3), 159 - 61
A site-specific endonuclease from Pseudomonas aeruginosa; Sokolov NN et al.; PaeI, a new restriction endonuclease from Pseudomonas aeruginosa clinical strain was isolated and characterized . It recognizes and cleaves the sequence 5'-GCATG reduced C-3' generating DNA fragments with 3'-tetranucleotide sticky ends . DNAs of pBR322, SV40 and bacteriophage lambda have one, two and six PaeI recognition sites, respectively . Seventy-two strains of Pseudomonas, Clostridium, Escherichia coli, Shigella, Proteus and Saccharomyces were screened for the presence of site-specific endonucleases . Here we describe the PaeI restriction enzyme found in Pseudomonas aeruginosa; other data will be published elsewhere . Earlier Hinkle and Miller isolated from P . aeruginosa a PaeR7 restriction endonuclease recognizing and cleaving a sequence 5'-C reduced TCGAG-3' (1) . Sequence analysis of DNAs cleaved by PaeI shows that the enzyme is the isoschizomer of SphI (2).

Eur J Biochem, 1985 Apr 1, 148(1), 67 - 73
Enzymic synthesis of the 4Fe-4S clusters of Clostridium pasteurianum ferredoxin; Bonomi F et al.; Ex novo enzymic synthesis of the two 4Fe-4S clusters of Clostridium pasteurianum ferredoxin has been achieved by incubation of the apoprotein with catalytic amounts of the sulfurtransferase rhodanese in the presence of thiosulfate, DL-dihydrolipoate and ferric ammonium citrate . This enzymic reconstitution procedure was compared to a chemical one, in which the enzyme was replaced by sodium sulfide . A further comparison was made with the results previously obtained in the enzymic synthesis of the 2Fe-2S cluster of spinach ferredoxin, allowing the following conclusions to be drawn . The nature of the cluster to be inserted into the reconstituted iron-sulfur protein is determined by the apoprotein itself . The refolding of the structure of the iron-sulfur proteins around the newly inserted cluster is the rate-limiting step in both chemical and enzymic reconstitution . Rhodanese appears to play a role in the recovery of the native architecture of the reconstituted iron-sulfur protein(s) . The extension to the 4Fe-4S centers of the rhodanese-based biosynthetic system allows this enzymic route to be proposed as a general way to the in vivo synthesis of iron-sulfur structures.

Gastroenterology, 1985 Apr, 88(4), 964 - 70
Antibiotic depression of evoked and spontaneous responses of opossum distal colonic muscularis mucosae in vitro: a factor in antibiotic-associated colitis?
Percy WH, Christensen J.
Certain antibiotics depress both skeletal neuromuscular transmission and intestinal neuroeffector transmission . Impaired intestinal motility may facilitate the proliferation of the bacterium Clostridium difficle and thus lead to the development of antibiotic-associated pseudomembranous colitis . Many antibiotics accumulate in the colonic lumen at concentrations several times their associated blood levels . This study examined whether certain of these could interfere with colonic muscularis mucosal movement in vitro, using tissue from opossum distal colon as a model . At concentrations approximating those in the colonic lumen, ampicillin, clindamycin, erythromycin, and lincomycin depressed tone and spontaneous contractions of the muscularis mucosae . Clindamycin, gentamicin, kanamycin, and lincomycin abolished electrically evoked contractions but only gentamicin and kanamycin could abolish the ensuing relaxation . Vancomycin potentiated the response of the muscularis mucosae to acetylcholine; erythromycin and clindamycin depressed it . Antibiotic-induced depression of colonic muscularis mucosal movement may contribute to the development of antibiotic-associated pseudomembranous colitis.

Br J Exp Pathol, 1985 Apr, 66(2), 243 - 9
Antitoxin activity of human polymorphonuclear leucocytes; Larson HE et al.; Human polymorphonuclear leucocytes (PMNL) inactivate Clostridium difficile cytotoxin and C . perfringens phospholipase C, but not C . perfringens enterotoxin . Both whole cells and sonicated suspensions possess activity, but mononuclear cell fractions of peripheral blood do not . Antitoxin activity closely correlates with cell concentration . The highest cell concentrations tested completely inactivated C . difficile cytotoxin by 2 min . Sucrose density gradient fractionation of PMNL showed antitoxin activity to be associated with myeloperoxidase, locating it in the primary or azurophil granules . Toxin inactivation was prevented by protease inhibitors suggesting that it is due to one of the neutral proteases present in these granules . PMNL are more active against C . difficile cytotoxin than purified chymotrypsin . PMNL may be a primary defence against certain bacterial exotoxins.

South Med J, 1985 Apr, 78(4), 440 - 4
Clostridial sepsis: unusual clinical presentations; Moustoukas NM et al.; We present four cases exhibiting the widely diverse nature of clinical infections due to anaerobic Clostridium perfringens . These cases exemplify the need for a thorough initial physical examination, immediate Gram staining of fluid from involved tissue, and recognition of the severity of the disease in any patient who has early septic deterioration after elective or emergency surgical procedures . Management of these infections includes both high-dose parenteral penicillin therapy and aggressive initial surgical debridement of all involved tissues.

FEBS Lett, 1985 Mar 25, 182(2), 479 - 84
The amino acid sequence of the enterotoxin from Clostridium perfringens type A; Richardson M et al.; The amino acid sequence of the enterotoxin from Clostridium perfringens type A was determined by analysis of peptides derived from the protein by digestion with trypsin chymotrypsin, thermolysin, pepsin, a lysine-specific protease . S . aureus V8 protease and a proline-specific protease, and fragments generated by cleavage with cyanogen bromide or by dilute acetic acid in 7 M guanidine HCl . The sequence which is complete except for the definite order of 3 small peptides between residues 88 and 103 consists of 309 amino acids and contains a correction to our preliminary announcement {(1984) FEMS Symp . 24, 329-330}.

Clin Chim Acta, 1985 Mar 15, 146(2-3), 119 - 27
Sialidase activity in the sera of patients and rabbits with clostridial myonecrosis; Schauer R et al.; The origin and nature of gas gangrene can be diagnosed exactly only by time-consuming bacteriological tests . In order to improve the diagnostic procedures, rabbits were infected with strains of Clostridium perfringens, Clostridium septicum or Clostridium sordellii . Sialidase activity was found to increase rapidly in serum; elevated creatine kinase activities were observed, too . High sialidase concentrations were found in sera (up to 1.6 mU/ml) and in tissues of wounded regions (up to 110 mU/g) of patients diagnosed to be infected with C . perfringens . By inhibition of enzyme activity with antibodies specific for the sialidase from this Clostridium species, it was possible to identify the clostridial origin of the sialidase activities . In the same material from other patients supposed to suffer from gas gangrene, but where no Clostridia could be detected, significant sialidase activity was not found . Thus, sialidase may be a useful tool for the diagnosis of myonecrosis due to clostridial infection.

J Biol Chem, 1985 Mar 10, 260(5), 2771 - 6
Substrate specificity of beta-collagenase from Clostridium histolyticum; Steinbrink DR et al.; The substrate specificity of beta-collagenase from Clostridium histolyticum has been investigated by measuring the rate of hydrolysis of more than 50 tri-, tetra-, penta-, and hexapeptides covering the P3 to P3' subsites of the substrate . The choice of peptides was patterned after sequences found in the alpha 1 and alpha 2 chains of type I collagen . Each peptide contained either a 2-furanacryloyl (FA) or cinnamoyl (CN) group in subsite P2 or the 4-nitrophenylalanine (Nph) residue in subsite P1 . Hydrolysis of the P1-P1' bond produces an absorbance change in these chromophoric peptides that has been used to quantitate the rates of their hydrolysis under first order conditions ({S} much less than KM) from kcat/KM values have been obtained . The identity of the amino acids in all six subsites (P3-P3') markedly influences the hydrolysis rates . In general, the best substrates have Gly in subsites P3 and P1', Pro or Ala in subsite P2', and Hyp, Arg, or Ala in subsite P3' . This corresponds well with the frequency of occurrence of these residues in the Gly-X-Y triplets of collagen . In contrast, the most rapidly hydrolyzed substrates do not have residues from collagen-like sequences in subsites P2 and P1 . For example, CN-Nph-Gly-Pro-Ala is the best known substrate for beta-collagenase with a kcat/KM value of 4.4 X 10(7) M-1 min-1, in spite of the fact that there is neither Pro nor Ala in P2 or Hyp nor Ala in P1 . These results indicate that the previously established rules for the substrate specificity of the enzyme require modification.

J Biol Chem, 1985 Mar 10, 260(5), 3140 - 4
Incorporation and distribution of selenium into thiolase from Clostridium kluyveri; Sliwkowski MX et al.; Clostridium kluyveri incorporates selenium as selenomethionine into its acetoacetyl-CoA thiolase when grown in media containing normal sulfur-to-selenium ratios . Antibodies raised against the purified enzyme permitted quantitative immunoprecipitation of thiolase from crude cell extracts and thus facilitated the systematic analysis of the effects of wide variation in sulfur-to-selenium ratios on selenium incorporation into the enzyme . The extent of incorporation of selenium into thiolase was found to be dependent on the form of selenium supplied . When {75Se}selenomethionine was the source of selenium, the incorporation of selenium into thiolase was inversely proportional to the level of added methionine . However, similar levels of methionine failed to decrease the incorporation of selenium from selenite . To study the location of selenomethionine and methionine residues in the polypeptide chain of the enzyme, thiolase was prepared from cells cultured in the presence of H2 35SO4 or Na2 75SeO3 . The 35S- or 75Se-labeled protein was treated with trypsin and the resulting peptides were isolated by reverse phase high performance liquid chromatography . The peptide maps of the enzyme indicated that selenium was distributed throughout the primary structure in a manner that paralleled methionine . From these studies, it is concluded that selenium occurs in thiolase adventitiously and is not required for any biological function.

J Biol Chem, 1985 Mar 10, 260(5), 2798 - 803
Pyruvoyl-dependent histidine decarboxylases . Preparation and amino acid sequences of the beta chains of histidine decarboxylase from Clostridium perfringens and Lactobacillus buchneri; Huynh QK et al.; Histidine decarboxylase (HisDCase) from Lactobacillus buchneri was purified to homogeneity . Its subunit structure, (alpha beta)6, and enzymatic properties resemble closely those of the immunologically cross-reactive HisDCase of Lactobacillus 30a (Recsei, P . A., and Snell, E . E . (1984) Annu . Rev . Biochem . 53, 357-387) . The complete amino acid sequences of the beta chains of the HisDCase from L . buchneri (81 residues) and Clostridium perfringens (86 residues) were then determined to be a and b, respectively . (a) SEFDKKLNTLGVDRISVSPYKKWSRGYMEPGNIGNGYVSGLKVDAG VVDKTDDMVLDGIGSYDRAETKNAYIGQINMTTAS . (b) TLSEGIHKNIKNIKVRAP KIDKTAISPYDRYCDGYGMPGAYGDGYVSVLKVSVGTVKK TDDILLDGIVSYDRAEINDAYVGQINMLTAS . SEFDKKLNTLGVDRISVSPYKKWSRGYMEPGNIGNGYVSGLKVDAGVV . Although these sequences differ substantially near the NH2-terminal ends, there is striking homology near the COOH termini and also near the NH2 terminus of the two alpha chains (pyruvoyl-Phe-X-Gly-Val-, where X is Ser or Cys) . If the four known pyruvoyl-dependent HisDCases arise from inactive proenzymes by the mechanism previously demonstrated for the HisDCase of Lactobacillus 30a (Recsei, P . A., Huynh, Q . K . and Snell, E . E . (1983) Proc . Natl . Acad . Sci . U.S.A . 80, 973-977), then each of these proenzymes has the sequence -Thr-Ala-Ser-Ser-Phe- at the activation site (where -Ser- becomes the COOH terminus of the beta chain and -Ser- becomes the pyruvoyl group blocking the NH2 terminus of the alpha chain), and the sequences around this activation site are highly conserved in all four enzymes . These facts support the assumptions that the four enzymes have evolved from a common ancestral protein, are formed from inactive pyruvate-free proenzymes by similar mechanisms, and have similar catalytic mechanisms.

J Biol Chem, 1985 Mar 10, 260(5), 2794 - 7
Pyruvoyl-dependent histidine decarboxylases . Comparative sequences of cysteinyl peptides of the enzymes from Lactobacillus 30a, Lactobacillus buchneri, and Clostridium perfringens; Huynh QK et al.; The two cysteinyl residues present in histidine decarboxylase from Lactobacillus 30a differ greatly in reactivity . One (class 1) reacts readily in the native state with dithiobis-(2-nitrobenzoate) with complete loss of enzyme activity; the other (class 2) reacts only after denaturation of the enzyme (Lane, R . S., and Snell, E . E . (1976) Biochemistry 15, 4175-4179) . These differences in reactivity permitted use of covalent (disulfide) chromatography to isolate separate peptides that contain these two residues . Sequence analysis showed that the class 1 cysteinyl residue is at position 147 in a hydrophilic portion of the alpha chain (Huynh, Q . K., Recsei, P . A., Vaaler, G . L., and Snell, E . E . (1984) J . Biol . Chem . 259, 2833-2839), while the class 2 cysteinyl residue is present at position 71, adjacent to a hydrophobic portion of the same chain . Cysteinyl peptides identical with or homologous to the class 2 cysteinyl peptide of the Lactobacillus 30a enzyme were isolated from the alpha subunits of histidine decarboxylases from Lactobacillus buchneri and Clostridium perfringens, respectively . The L . buchneri enzyme also contained a peptide homologous to the class 1 cysteinyl peptide from Lactobacillus 30a . However, no corresponding peptide was present in the enzyme from C . perfringens, in which the second cysteinyl residue of the alpha chain occupies position 3, very near the essential pyruvoyl residue . This enzyme, unlike those from Lactobacillus 30a or L . buchneri, also contains one cysteinyl residue in its beta chain . Although Cys 147 is an active site residue in histidine decarboxylase from Lactobacillus 30a, the absence of a corresponding residue in the C . perfringens enzyme confirms previous indications (Recsei, P . A., and Snell, E . E . (1982) J . Biol . Chem . 257, 7196-7202) that this SH group is not essential for decarboxylase action.

Anal Biochem, 1985 Mar, 145(2), 222 - 9
Anaerobic multiphasic gel electrophoresis of the molybdoproteins in extracts of Clostridium pasteurianum; Hinton SM et al.; Simple procedures using multiphasic buffer systems for anaerobic electrophoresis have been devised to identify oxygen-labile metalloproteins . An anaerobic slab gel apparatus was developed with cooling and design for anaerobic conditions . Included is a procedure to remove sample wells after stacking proteins in a crude extract, to prevent streaking (background) caused by continuous leakage of "nonstacked protein" from the sample wells . Identification of eleven Mo zones in extracts of Clostridium pasteurianum demonstrates the usefulness of the technique in identifying radiolabeled oxygen-labile proteins in cell-free crude lysates.

Mikrobiologiia, 1985 Mar-Apr, 54(2), 322 - 4
{Superoxide dismutase in Clostridium butyricum spores}; Gaenko GP et al.; Superoxide dismutase was found for the first time in the spores of the anaerobic bacterium Clostridium butyricum . The prosthetic group of the enzyme was shown to contain iron . The enzyme was demonstrated to be highly thermostable.

Can J Microbiol, 1985 Mar, 31(3), 276 - 81
Determination of mercury and organomercurial resistance in obligate anaerobic bacteria; Rudrik JT et al.; A methodology for determining the minimum inhibitory concentration of inorganic and organomercurial compounds for obligate anaerobic bacteria is described . A wide variation in the susceptibility of anaerobic clinical and sewage isolates was observed . Isolates of Bacteroides ruminicola and Clostridium perfringens resistant to mercury were examined for their plasmid content and ability to demonstrate inducible resistance . None of the resistant anaerobes contained any plasmids, while resistant facultative isolates from the same source contained several plasmids . In 24 h, resistant strains of clostridia and Bacteroides volatilized 20 and 43% of the 203Hg2+ added to cultures, while Escherichia coli R100 and a sewage isolate of Enterobacter cloacae volatilized 63 and 27%, respectively, of the added 203Hg2+ . Attempts to induce mercury resistance in the aerobic isolates were successful, but no induction was seen in the anaerobes . Thus, mercury resistance in these anaerobic isolates was neither inducible nor plasmid mediated.

Appl Environ Microbiol, 1985 Mar, 49(3), 682 - 5
beta-Glucuronidase activities of intestinal bacteria determined both in vitro and in vivo in gnotobiotic rats; Gadelle D et al.; The beta-glucuronidase activities of bacterial strains isolated from the rat intestinal tract were studied both in vitro in culture media and in vivo in the intestinal contents of gnotobiotic rats . Only 50 of 407 strains tested were found to be positive in vitro . They belonged to the three genera Clostridium, Peptostreptococcus, and Staphylococcus . The in vitro-negative strains were also negative in vivo . The beta-glucuronidase activities of the beta-glucuronidase activities of the positive strains were generally greater in vivo than in vitro . The highest in vivo activities were found in the intact bacterial cells and in the soluble fractions prepared from disrupted pellets . There was a discrepancy between the activities obtained from both conventional and gnotobiotic rats harboring selected positive strains, suggesting that the main beta-glucuronidase-positive strains have not yet been isolated from the intestines of conventional rats.

Appl Environ Microbiol, 1985 Mar, 49(3), 644 - 9
Bovine serum eliminates rapid nonspecific toxic reactions during bioassay of stored fish for Clostridium botulinum toxin; Solberg M et al.; When stored fish or some fish products were tested for the presence of Clostridium botulinum toxin, nonspecific toxic reactions in mice often occurred, rendering the bioassay inconclusive . The nonspecific toxic reactions were mediated by the gram-negative microbiota, inherent to the fish, which were the source of lethal, heat-stable endotoxins . The treatment of assay samples with bovine serum eliminated nonspecific reactions through the interaction of constituent serum immunoglobulin M (IgM) with endotoxic material . Removal of IgM from bovine serum through treatment with protein A or concanavalin A resulted in a loss of protective activity.

Antimicrob Agents Chemother, 1985 Mar, 27(3), 419 - 21
Effect of L-cysteine on the activity of penicillin antibiotics against Clostridium difficile; Markowitz SM et al.; We observed elevated MICs of penicillin antibiotics while performing agar dilution susceptibility testing of strains of Clostridium difficile on supplemented brain heart infusion agar, an effect which was completely eliminated by the exclusion of L-cysteine from the medium . L-Cysteine antagonizes the activity of penicillins against C . difficile, most likely by direct inactivation of the antibiotic.

Mycopathologia, 1985 Mar, 89(3), 129 - 34
Case report: prosthetic valve endocarditis caused by Pseudallescheria boydii and Clostridium limosum; Gordon G et al.; This report describes a patient with a combined infection due to Pseudallescheria boydii and Clostridium limosum on a prosthetic dura mater aortic valve homograft . While this patient had C . limosum only growing in blood cultures, both organisms were isolated from the surgically resected aortic valve . Because P . boydii is generally resistant to amphotericin B but susceptible to miconazole, accurate differentiation of P . boydii from other fungi which may appear similarly in tissue sections (e.g., aspergillus) is important.

Arch Dis Child, 1985 Mar, 60(3), 252 - 4
Detection of Clostridium difficile enterotoxin in neonates by latex agglutination; Ushijima H et al.; Clostridium difficile enterotoxin (D-1) was detected in 13 symptomatic and nine asymptomatic neonates in the neonatal intensive care unit by latex agglutination test but was not found in 18 healthy neonates in two other newborn nurseries . Environmental contamination in the intensive care unit may have been the cause . An association between the presence of enterotoxin and clinical symptoms is discussed.

J Hand Surg {Am}, 1985 Mar, 10(2), 281 - 4
Metastatic nontraumatic Clostridium septicum osteomyelitis; Neimkin RJ et al.; Nontraumatic clostridial infections are rare, but need to be diagnosed and treated early or high morbidity and mortality rates result . We believe that this is the first reported case of metastatic nontraumatic Clostridium septicum osteomyelitis . Early treatment with surgical debridement and parenteral antibiotics without hyperbaric oxygen was used . An associated occult rectal malignancy was discovered and treated.

J Clin Microbiol, 1985 Mar, 21(3), 323 - 7
Serogrouping of Clostridium difficile strains by slide agglutination; Delmee M et al.; Six different agglutinating antisera were obtained by immunizing rabbits with Formol-treated strains of Clostridium difficile . After appropriate absorption, these antisera were used to define six serogroups designated by the letters A, B, C, D, F, and G . Altogether, 315 strains of C . difficile from various origins were tested for slide agglutination by these antisera; 312 (99%) of them were agglutinated by one of these antisera . A and C were the most common serogroups . An excellent correlation, ranging from 85 to 100%, was found between the serogroup and the toxigenicity of the strains . The correlation between serogroup and sorbitol fermentation was higher, ranging from 89 to 100% . The results of this typing were compared with the clinical origin of the strains . Only strains of serogroups A, C, and D were isolated in 153 cases of antibiotic-associated diarrhea . This series included strains from three outbreaks; all the strains in two of the outbreaks belonged to serogroup C, and in the third, all the strains belonged to serogroup A . Strains of serogroups B, F, and G were only found in the stools of asymptomatic neonates or young children . In the latter samples, strains of serogroups A and D were found in the same ratio as in adults with antibiotic-associated diarrhea, but strains of serogroup C were seldom isolated . In patients treated with antineoplastic drugs and suffering from diarrhea, the distribution of the strains was the same as in cases of antibiotic-associated diarrhea.

J Infect Dis, 1985 Mar, 151(3), 476 - 81
Antimicrobial agents and Clostridium difficile in acute enteric disease: epidemiological data from Sweden, 1980-1982; Aronsson B et al.; The carrier rate of Clostridium difficile in an adult Swedish population was found to be 11 (1.9%) of 594 . All isolates were toxigenic in vitro, but no healthy individual harbored free cytotoxin in stool . Of 398 patients with acute diarrhea not associated with antibiotic use, cytotoxin was found in stool filtrates of four (1%) . In 4,793 patients with antibiotic-associated diarrhea from all parts of Sweden during 1980-1982, C . difficile cytotoxin was demonstrated in 873 (18%) . The tissue culture assay was found to be more specific than cultivation for the bacterium . By weighted analysis, in the age group greater than 70 years more women than men were infected . In the age group 21-50 years there was an even greater preponderance of infection in women than in men . Cephalosporins and lincosamides were 10-70 times more often implicated in C . difficile colitis than were narrow-spectrum penicillins.

Infect Immun, 1985 Mar, 47(3), 697 - 703
Experimental cecitis in gnotoxenic chickens monoassociated with Clostridium butyricum strains isolated from patients with neonatal necrotizing enterocolitis; Popoff MR et al.; An animal model for Clostridium butyricum necrotizing cecitis has been developed in axenic chickens inoculated orally between 2 and 50 days of life . Cecitis was obtained with two C . butyricum strains isolated from neonatal necrotizing enterocolitis and not with a Clostridium beijerinckii strain from dairy products; the rate of colonization of the intestinal tract by this strain was lower than that obtained with C . butyricum strains . The clinical findings showed a slow gain in body weight . The cecitis lesions were well developed 3 and 4 weeks after oral inoculation, including enlargement with an increase of the cecum weight-body weight ratio, a marked hyperplasia, congestion, inflammatory infiltrate and pneumatosis of the cecal wall and mesentery, hemorrhage in the lamina propria and submucosa, and ulcerations and necrotic areas in the mucosa . By immunofluorescence and electron microscopy, the bacterial cells were located in the cecal lumen and in necrotic areas of the mucosa . The presence of 4% lactose in the diet seemed to be a prerequisite for the development of cecitis in chickens . A gradual rise of fluorescent antibodies in the sera was observed.

JAMA, 1985 Mar 1, 253(9), 1275 - 8
Type A botulism from sauteed onions . Clinical and epidemiologic observations; MacDonald KL et al.; Twenty-eight persons were hospitalized in Illinois with neurologic signs and symptoms compatible with botulism in October 1983 . Twelve patients required ventilatory support, and 20 patients were treated with trivalent ABE antitoxin; one patient died while still in the hospital six months after onset of illness . Type A toxin and/or type A Clostridium botulinum were subsequently identified in specimens from 18 patients . Case-control studies implicated sauteed onions made from fresh raw onions and served on a patty-melt sandwich in a local restaurant as the vehicle of transmission . Although the original sauteed onions were not available for toxin testing, type A toxin was detected in washings from a wrapper in which a patty-melt sandwich was taken home by one of the ill persons . Also, type A C botulinum was cultured from five of 75 raw onions taken from the restaurant . This outbreak implicated an unusual vehicle for botulinal toxin that was initially not suspected and demonstrates the importance of considering all theoretically possible food items as potential vehicles for toxin until epidemiologic and laboratory data have been collected and analyzed.

Antimicrob Agents Chemother, 1985 Mar, 27(3), 424 - 6
Establishment of MICs of moxalactam for control and reference anaerobic organisms in agar dilution and microdilution techniques; Sutter VL et al.; The MICs of moxalactam were determined for eight National Committee for Clinical Laboratory Standards control and reference strains and for Fusobacterium nucleatum ATCC 10953 by agar and microdilution techniques . The recommended MIC for the control strain Bacteroides fragilis in both agar and microdilution tests is 0.5 micrograms/ml . Recommended MICs for Bacteroides thetaiotaomicron ATCC 29741 and Clostridium perfringens ATCC 13124 by agar dilution are 8 and 0.063 micrograms/ml, respectively . These two strains gave inconsistent results in microdilution tests . Variation in results with microdilution procedures was seen, which illustrates problems in reading endpoints and with modifications of media . Recommended MICs for the reference strains are presented.

Diagn Microbiol Infect Dis, 1985 Mar, 3(2), 105 - 12
Detection of Clostridium botulinum type B toxin in the presence of a lethal substance interfering with toxin neutralization; Dezfulian M et al.; A low molecular weight substance that induced seizures and death in mice was present in the stool of a child with infant botulism . This interfered with mouse bioassay used for detecting Clostridium botulinum toxin . After removal of the interfering substance by a prolonged dialysis with buffered saline, in vivo neutralization of botulinal toxin was readily achieved . Alternatively, the botulinal toxin was demonstrated in stool of the infant by an enzyme-linked immunosorbent assay (ELISA), using antibodies produced in immunologically tolerant rabbits.

J Lipid Res, 1985 Mar, 26(3), 344 - 50
Bile acid induction of 7 alpha- and 7 beta-hydroxysteroid dehydrogenases in Clostridium limosum; Sutherland JD et al.; When grown in the presence of bile acids, two strains of Clostridium limosum were found to contain significant amounts of NADP-dependent 7 alpha/7 beta-hydroxysteroid dehydrogenase and NAD-dependent 7 alpha-hydroxysteroid dehydrogenase which were active against conjugated and unconjugated bile acids . No measurable activity could be found when deoxycholic acid (3 alpha, 12 alpha-dihydroxy-5 beta-cholan-24-oic acid) was used as substrate . No 7 beta-hydroxysteroid dehydrogenase activity and only a trace of 7 alpha-hydroxysteroid dehydrogenase activity could be demonstrated when bile acid was deleted from the growth medium . If bile acid was added after the time of inoculation, the amounts of 7 alpha/7 beta-hydroxysteroid dehydrogenase were greatly reduced . Enzyme enhancement was blocked by addition of rifampicin . The 7 alpha/7 beta-hydroxysteroid dehydrogenase components had pH optima of approximately 10.5 . Both the 7 alpha/7 beta-hydroxysteroid dehydrogenase activities were heat-labile, with the 7 beta-component being the more stable of the two . When ranked according to the level of enzymes induced, the order in increasing bile acid induction power on an equimolar scale (0.4 mM) was: 7-ketodeoxycholic acid, cholic acid, chenodeoxycholic acid, and deoxycholic acid . Both 7-ketolithocholic acid and ursodeoxycholic acid were ineffective as enzyme inducers . Optimal induction was achieved with high concentrations of cholic acid (5 mM) and a harvest time of 24 hr . Addition of ursodeoxycholic acid to medium containing optimal concentrations of deoxycholic acid suppressed enzyme induction.(ABSTRACT TRUNCATED AT 250 WORDS)

Proc Natl Acad Sci U S A, 1985 Mar, 82(6), 1653 - 7
Cloning and nucleotide sequence determination of the Clostridium pasteurianum ferredoxin gene; Graves MC et al.; We have constructed a library of Clostridium pasteurianum DNA cloned in the plasmid pBR322 . Based on the known amino acid sequence for C . pasteurianum ferredoxin, a 64-fold degenerate heptadecanucleotide pool was synthesized . This mixed probe hybridized to two clones which were shown to contain greater than 6 kilobase pairs of the same genomic DNA . Sequence analysis of a common Sau3A1 0.6-kilobase-pair fragment revealed that it contains the information for the apoferredoxin structural gene . According to the DNA sequence, the only post-translational processing of this small apoprotein is the hydrolysis of the initiator methionine . Putative transcription and translation start and stop signals are present within the sequence.

Anal Biochem, 1985 Mar, 145(2), 286 - 91
Purification of nonspecific protease-free collagenase from Clostridium histolyticum; Bicsak TA et al.; The collagenase (EC 3.4.24.3) produced by the bacterium Clostridium histolyticum has been purified free from nonspecific protease contaminants by a two-step procedure . The crude culture medium is chromatographed over heparin-Sepharose and Sephacryl S-200, and the resulting preparation has no activity versus noncollagenous proteins or N alpha-benzoyl-L-arginine ethyl ester, yet cleaves native thermally reconstituted collagen fibrils quite efficiently (specific activity, 3000 units/mg) . The purification described may be useful for those investigators requiring substantially purified collagenase for applications such as cell culture or collagen quantitation in protein mixtures.

Clin Immunol Immunopathol, 1985 Mar, 34(3), 304 - 15
The clinical manifestations of a genetically determined deficiency of the third component of complement in the dog; Blum JR et al.; The clinical manifestations of a genetically determined deficiency of C3 were examined in a closed colony of dogs . One hundred and twelve dogs, including twenty C3-deficient dogs, were studied over a period of 6 years . Five of the C3-deficient dogs developed significant bacterial infections, such as pneumonia, sepsis, and pyometra, which were caused by Clostridium spp., Escherichia coli, and Klebsiella spp . Two of the C3-deficient dogs who had had significant infections also subsequently developed renal disease . Secondary amyloidosis was the predominant renal lesion in one dog . The predominant renal lesion in the second dog was membranoproliferative glomerulonephritis, although some amyloid was also present . The two dogs with renal disease also had positive rheumatoid factors . No other clinical or serological evidence of autoimmune disease or immune complex disease has been found . None of the dogs heterozygous for C3 deficiency, and none of the homozygous normal dogs in the colony has developed significant bacterial infections or renal disease . Thus, dogs deficient in C3, like C3-deficient humans, demonstrate both an increased susceptibility to infection and renal disease.

J Hosp Infect, 1985 Mar, 6(1), 41 - 5
The activity of glutaraldehyde against Clostridium difficile; Dyas A et al.; The sporicidal activity of 2% glutaraldehyde against Bacillus subtilis var . globigii and Bacillus stearothermophilus was compared with that against Clostridium difficile . The aerobic species, normally chosen for test purposes, survived for 2 h but Cl . difficile was killed in under 10 min . In view of the impractical and lengthy immersions required to satisfy standard tests using non-pathogenic spores, a less stringent standard would probably be more appropriate.

Am J Public Health, 1985 Mar, 75(3), 287 - 8
Two successive outbreaks of Clostridium perfringens at a state correctional institution; Tavris DR et al.; An outbreak of acute gastrointestinal illness of short duration involving 100 inmates at a correctional institution followed a similar outbreak among the same population by eight days . Clostridium perfringens was the specific etiology in both outbreaks; the vehicle was roast beef in the first outbreak, ham in the second . Direct observation of food handling practices revealed that the meats were not cooled quickly enough following cooking; not reheated adequately prior to serving, and; held at improper temperatures prior to serving.

Anal Biochem, 1985 Mar, 145(2), 339 - 42
Fluorescent staining of sialidases in polyacrylamide gel electrophoresis and ultrathin-layer isoelectric focusing; Berg W et al.; Polyacrylamide gels were stained with the sialidase substrate 2'-(4-methylumbelliferyl)-alpha-D-N-acetylneuraminic acid showing the activity of Vibrio cholerae and Clostridium sordellii sialidases in the gels after electrophoresis . With this fluorogenic method minimum sialidase activities of 5 microU could be determined . The sensitivity of this staining is about 10,000-fold higher compared to protein-staining with Coomassie brilliant blue . For the visualization of other proteins than sialidases the specific sialidase staining could be followed by a protein-staining method in the same gel.

Biochim Biophys Acta, 1985 Feb 28, 813(1), 10 - 8
Phospholipid aliphatic chain composition modulates lipid class composition, but not lipid asymmetry in Clostridium butyricum; Johnston NC et al.; The phospholipid composition of the butyric acid-producing clostridia is responsive to the degree of enrichment of the lipids with cis-unsaturated fatty acids . When Clostridium butyricum and Clostridium beijerinckii are grown on oleic acid in media devoid of biotin, the acyl and alk-1-enyl chains of the phospholipids become highly enriched with 18:1 and C19-cyclopropane . Under these conditions there is a marked increase in the glycerol acetals of the major plasmalogens of these organisms . We have grown both species on mixtures of palmitate and oleate in the absence of biotin . The alk-1-enyl chains were highly enriched with C18-unsaturated and C19-cyclopropane residues at all but the highest ratios of palmitate to oleate (80:20, w/w) added to the medium . At ratios of palmitate to oleate greater than or equal to 40:60, the saturated acid was incorporated predominantly into the phospholipid acyl chains in both organisms . The effects of increasing unsaturation of the acyl chains as the ratio of oleate to palmitate was increased was examined in C . butyricum . In cells grown on mixtures of palmitate and oleate equal to or exceeding 40% palmitate, the ratio of glycerol acetal lipid to total phosphatidylethanolamine (PE) was relatively constant . As the proportion of oleic acid added to the medium was increased, the ratio of glycerol acetal lipid to PE increased from 0.7 to 2.0 . Thus the ratio of the polar lipids appears to respond to the content of phospholipids that contain two unsaturated chains . The fraction of PE present as plasmalogen remained relatively stable (0.82 +/- 0.05) at varying ratios of medium oleic and palmitic acids . Both the glycerol acetal of ethanolamine plasmalogen, and ethanolamine plasmalogen, are shown to be 80% or more in the outer monolayer of the cell membrane . These two polar lipids represent approx . 50% of the phospholipids in cells grown on exogenous fatty acid . The bulk of the remainder is polyglycerol phosphatides . We suggest that the ability of both species to grow with highly unsaturated membranes is related to their ability to modulate their polar lipid composition.

Biochemistry, 1985 Feb 26, 24(5), 1141 - 7
Effect of high pN2 and high pD2 on NH3 production, H2 evolution, and HD formation by nitrogenases; Jensen BB et al.; We have investigated the effect of the partial pressure of N2 and D2 on HD formation, H2 evolution, and NH3 production by nitrogenase from Klebsiella pneumoniae and Clostridium pasteurianum . By using pressures up to 4 atm, we have been able to extend the concentration range of N2 and D2 in our investigations beyond that used in previous studies . The pN2 dependence of HD formation with constant pD2 ideally shows no HD formation under zero pN2, reaches a peak which depends on the pD2, and then decreases to zero at very high pN2 . K . pneumoniae and C . pasteurianum nitrogenases differ in their Ki(D2) for nitrogen fixation . C . pasteurianum nitrogenase had the lower activity for formation of HD . With K . pneumoniae nitrogenase, D2 enhanced H2 evolution from 31% of the electron flux partitioned to H2 in the absence of D2 to 51% of the electron flux partitioned to H2 at 400 kPa of D2 . With C . pasteurianum nitrogenase, the equivalent values were 33% and 48% of the total electron flux . Our results support the mechanism for nitrogenase-catalyzed reductions proposed by W . W . Cleland {Guth, J., & Burris, R . H . (1983) Biochemistry 22, 5111-5122}.

Eur J Biochem, 1985 Feb 15, 147(1), 27 - 31
Induction of ornithine decarboxylase in guinea-pig lymphocytes . Synergistic effect of diacylglycerol and calcium; Otani S et al.; Calmodulin antagonists, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), N-(6-aminohexyl)-1-naphthalenesulfonamide (W-5) and trifluoperazine inhibited ornithine decarboxylase induction in lymphocytes activated with phytohemagglutinin or inophore A23187 . W-7, a more potent calmodulin antagonist than W-5, suppressed ornithine decarboxylase induction in a higher extent than did W-5 . These results suggest that calmodulin may play an important role in ornithine decarboxylase induction in the activated lymphocytes . However, the extent of ornithine decarboxylase induction was greater in cells pretreated with Clostridium phospholipase C and then incubated with ionophore A23187 than in cells incubated with ionophore A23187 without the pretreatment . Moreover, combined treatment of cells with ionophore A23187 and tumor promotor, phorbol 12-myristate 13-acetate, caused synergistic induction of ornithine decarboxylase activity . These results, taken together, suggest that both activations of Ca2+-activated phospholipid-dependent protein kinase by diacylglycerol and of calmodulin-dependent function resulted from an elevation of cytosolic Ca2+ concentration may operate in the induction of ornithine decarboxylase in the activated lymphocytes.

J Biol Chem, 1985 Feb 10, 260(3), 1452 - 8
Kinetics of reduction of Clostridium pasteurianum rubredoxin by laser photoreduced spinach ferredoxin:NADP+ reductase and free flavins . Electron transfer within a protein-protein complex; Przysiecki CT et al.; The kinetics of reduction of oxidized Clostridium pasteurianum rubredoxin (Rdox) by free flavin semiquinones generated by the laser flash photolysis technique and by spinach ferredoxin:NADP+ reductase (FNR) semiquinone (also produced by flavin semiquinone reduction) have been investigated under anaerobic conditions . 5-Deazariboflavin semiquinone (5-dRf) rapidly reduces oxidized rubredoxin (Rdox) (k = 3.0 X 10(8) M-1 S-1) and oxidized ferredoxin:NADP+ reductase (FNRox) to the semiquinone level (k = 5.5 X 10(8) M-1 S-1) . Lumiflavin semiquinone reduces Rdox more slowly (k = 1.3 X 10(7) M-1 S-1) and is not measurably reactive with FNRox . Absorption difference spectroscopy and difference CD indicate that Rdox and FNRox form a 1:1 complex at low ionic strength (10 mM), which is completely dissociated at higher ionic strength (310 mM) . Apparent second order rate constants for reduction of Rdox in its free and complexed state by lumiflavin semiquinone are the same . Reduction of Rdox (both free and complexed) by free FNR semiquinone and intracomplex electron transfer were investigated using 5-dRf as the reductant . At I = 10 mM, a first order rate constant of 2.0 X 10(3) S-1 was obtained, which corresponds to the processes involved in intracomplex electron transfer from FNR semiquinone to Rdox . A second order reaction between free FNR semiquinone and complexed Rdox was also observed to occur (k = 5 X 10(7) M-1 S-1) . At I = 310 mM, these reactions are not observed and the reaction of FNR semiquinone with free Rdox is second order (k = 4 X 10(6) M-1 S-1).

J Chromatogr, 1985 Feb 8, 337(2), 213 - 21
Gas chromatographic-mass spectrometric analysis of volatile amines produced by several strains of Clostridium; Pons JL et al.; A gas chromatographic--mass spectrometric technique is proposed for the analysis of volatile amines which were isolated from Clostridium cultures by vacuum distillation and concentrated as hydrochloride salts . Headspace sampling after alkalinization of the salts under vacuum was the most suitable for subsequent gas chromatographic analysis . With ammonia-loaded helium as carrier gas, methylamines were separated on 4.8% PEG 2OM + 0.3% potassium hydroxide on Carbopack B, and other volatile amines on 28% Pennwalt 223 + 4% potassium hydroxide on Gas-Chrom R . Bacterial volatile amines (dimethylamine, trimethylamine, isobutylamine, 3-methylbutylamine, etc.) were detected with a flame-ionization detector and identified by gas chromatography--mass spectrometry in electron-impact and chemical ionization modes.

Lancet, 1985 Feb 2, 1(8423), 237 - 41
Continuous microbiological and pathological study of 70 sudden and unexpected infant deaths: toxigenic intestinal clostridium botulinum infection in 9 cases of sudden infant death syndrome; Sonnabend OA et al.; As part of a programme to exclude infection as the cause of death in infants who died suddenly and unexpectedly necropsies were carried out on 70 such infants . In 11 cases (15%), a pathological diagnosis could be made at necropsy; in 9 of these, causative bacteria or viruses were found . The 59 cases in which the cause of death could not be found had histological features characteristic of sudden infant death syndrome (SIDS) . Botulinum toxin was found in 9 SIDS cases (15%) . 8 of these infants had botulinum toxin and organisms of different types (A, B, C, F, G) in the contents of the ileojejunum or colon . 4 of them also had toxin in the serum . No botulinum toxin or organisms were found in the 11 infants who died of identified causes or 18 other infants who died in hospital of known causes.

Biochimie, 1985 Feb, 67(2), 241 - 8
Differential levels of ferredoxin and rubredoxin in Clostridium acetobutylicum; Marczak R et al.; Ferredoxin and rubredoxin levels have been determined in DEAE-cellulose treated extracts of Clostridium acetobutylicum using specific enzymatic assays . In contrast to ferredoxin, the content of rubredoxin is affected by various culture conditions; it fluctuates in the proportions of 1 to 3 according to the growth phase, 1 to 8 according to the medium composition, and 1 to 40 according to the pH . Highest rubredoxin level is obtained at the end of the acid phase when the cells grow in chemically defined medium, the pH of which is not controlled . Such variations suggest that rubredoxin as well as rubredoxin-reductase take part in an electron transport chain system inducible by the culture conditions.

Arch Microbiol, 1985 Feb, 141(1), 85 - 90
Characterization of citrate lyase from Clostridium sporosphaeroides; Quentmeier A et al.; Cells of Clostridium sporosphaeroides which were grown on citrate contained citrate lyase and citrate lyase acetylating enzyme, but no detectable citrate synthase and citrate lyase deacetylase activities . Citrate lyase from C . sporosphaeroides was purified to homogeneity as judged by polyacrylamide gel electrophoresis and high performance liquid chromatography . In contrast to the enzyme from Clostridium sphenoides, the addition of L-glutamate was not necessary for activity and stabilization of the enzyme . The purified enzyme had a specific activity of 34 U/mg protein and was comparable to other citrate lyases with respect to its molecular weight and subunit composition . Electron microscopic investigations showed that similar to the lyase from C . sphenoides and in contrast to all other citrate lyases examined so far, the majority of the enzyme molecules was present in "star" form.

Zh Mikrobiol Epidemiol Immunobiol, 1985 Feb, (2), 24 - 7
{Ultrastructural characteristics of the natural heteromorphic growth of Clostridium septicum}; Vaisman ISh; Periodic cultures of C . septicum strain No . 59, growing in Pope's broth, have been studied by means of transmission electron microscopy on ultrathin sections . During the lagphase of growth the inoculated bacilliary cells are sequentially converted into giant filamentous multinucleate forms . These changes, reflecting the reaction of phenotypical adaptation to new environmental conditions, are followed by the restoration of the initial phenotype via the fragmentation of the giant cells . At the same time heteromorphic growth becomes, also spontaneously, atypical in some of the multinucleate cells . This atypical heteromorphic growth results in pronounced degenerative changes leading to bacteriolysis and the death of microbial cells due to disturbances in the coordination of the formation of structural and functional complexes in these cells at the period of their phenotypical adaptation . Such pathology of microbial cells leads to elimination of individual cells with developmental defects, which is finally conducive to the sanitation of the population.

Antimicrob Agents Chemother, 1985 Feb, 27(2), 162 - 6
Comparative in vitro activity of cefbuperazone against anaerobic bacteria isolated from community hospitals; Goldstein EJ et al.; The activity of cefbuperazone against 266 strains of anaerobic bacteria was determined by the agar dilution method and compared with cefoxitin, moxalactam, piperacillin, and clindamycin . All strains were recent clinical isolates from community hospitals . All agents tested showed good activity against Bacteroides fragilis, Fusobacterium spp., Propionibacterium spp., Clostridium septicum, Clostridium perfringens, and the anaerobic, gram-positive cocci and gram-negative cocci . Cefbuperazone, cefoxitin, and moxalactam had poor activity against Bacteroides thetaiotaomicron, Bacteroides ovatus, and Bacteroides distasonis . The susceptibility of other Clostridium spp., Lactobacillus spp., and Eubacterium lentum was variable . Our community hospital isolates showed a difference in susceptibility patterns from those reported from university and research centers . This supports the recommendation that clinical microbiology laboratories, including those in community hospitals, need to perform susceptibility testing on representative clinical isolates.

Isr J Med Sci, 1985 Feb, 21(2), 150 - 3
Botulism intoxication after surgery in the gut; Isacsohn M et al.; Botulism intoxication is described in a 45-year-old woman who was hospitalized with symptoms suggesting an intestinal obstruction . At intervention a 20-cm length of necrotic small intestine was excised and prophylactic antibiotics were given . Five days after surgery--during which time the patient was fed only parenterally--dryness of the mouth, difficulty in swallowing, general weakness, and difficulty with speech and in keeping her eyes open were noted . The patient was alert and well oriented . The clinical symptoms and the electrophysiological studies suggested the diagnosis of botulism . A serum sample caused death in mice upon inoculation and two samples of feces were positive for the presence of Clostridium botulinum organisms . Supportive treatment with botulinum antitoxin was given and the patient was discharged in good condition.

Virology, 1985 Feb, 141(1), 144 - 7
Neuraminic acid is involved in the binding of influenza C virus to erythrocytes; Herrler G et al.; Neuraminidases of both viral and bacterial origin have been reported to be unable to destroy the cellular receptor for influenza C virus on chicken erythrocytes, in contrast to the receptors for influenza A and B virus . However, under appropriate conditions neuraminidases from both Vibrio cholerae and Clostridium perfringens were able (i) to make chicken red blood cells resistant against agglutination by influenza C virus and (ii) to reduce the hemagglutination-inhibiting activity of rat serum . Both effects were abolished in the presence of the neuraminidase inhibitor 2,3-dehydro-2-deoxyneuraminic acid (DDN) . These results indicate that contrary to previous assumptions sialic acid may very well be an essential component of the receptor for influenza C virus.

J Clin Microbiol, 1985 Feb, 21(2), 269 - 72
Evaluation of broth microdilution susceptibility results for anaerobic organisms by use of a rapid direct colony inoculum; Mangels JI et al.; A direct colony inoculum suspension procedure was compared with the overnight suspension procedure recommended for the broth microdilution anaerobic commercial system (Micro-Media Systems, Inc., Potomac, Md.) . Six National Committee for Clinical Laboratory Standards-recommended quality control organisms, Bacteroides fragilis ATCC 25285, Clostridium perfringens ATCC 13124, Bacteroides thetaiotaomicron ATCC 29741, Bacteroides vulgatus ATCC 29327, Peptococcus magnus ATCC 29328, Peptococcus asaccharolyticus ATCC 29743, and 50 anaerobic clinical isolates were tested against seven commonly tested antimicrobial agents . The minimum inhibitory concentration results from each suspension method (using the quality control organisms) were identical in 18 (78%) instances, and within +/- 1 log2 dilution in 96% of the comparisons . Results with the fresh clinical isolates also compared satisfactorily with the overnight procedure (97% were identical or within one dilution) . The Wilkins-Chalgren test medium failed to support the growth of most anaerobic gram-positive cocci and Bacteroides melaninogenicus strains.

J Clin Microbiol, 1985 Feb, 21(2), 251 - 4
Two bacteriophages of Clostridium difficile; Mahony DE et al.; Two temperate bacteriophages of differing morphology and host range were isolated by screening 94 isolates of Clostridium difficile . Phage 41 had a 300-nm flexible tail, whereas phage 56 had a shorter tail with a contractile sheath . Electron microscopy of phage 56 lysates exposed to elevated magnesium concentrations showed small virus-like particles which were 21 nm in diameter . The addition of MgCl2 to semisolid agar overlays enhanced both the titer and plaque size of phage 56 . Phage 56 was more temperature labile than phage 41 and demonstrated unusual lability in buffer at pH 7.0 . One-step growth and adsorption experiments revealed that both phages had latent periods of about 60 min, but phage 56 adsorbed to its indicator strain more efficiently . Phage 56, which was obtained from a toxigenic strain of C . difficile, was used to lysogenize its nontoxigenic indicator strain, but no conversion to toxigenicity was observed in this strain.

J Infect Dis, 1985 Feb, 151(2), 355 - 61
Population dynamics of ingested Clostridium difficile in the gastrointestinal tract of the Syrian hamster; Wilson KH et al.; The population dynamics of Clostridium difficile in the hamster gastrointestinal tract were studied after intragastric inoculation with organisms and a 51Cr tracer . Seventy-eight percent of spores germinated within the small intestine within 1 hr . Germinated spores and vegetative cells both showed two phases of elimination from the hamster cecum--an initial phase of rapid death that was not affected by antibiotic treatment followed by a phase of complete inhibition of multiplication . The latter phase of inhibition was not seen in antibiotic-treated animals and was thus attributable to the indigenous flora . The 51Cr tracer mixed well with cecal contents and was eliminated exponentially with a dilution rate constant ranging from -0.46/hr to -0.31/hr in normal hamsters . The hamster cecum was therefore dynamically analogous to a continuous flow system, a finding supporting the concept that anaerobic continuous flow cultures are useful in vitro models of the cecal ecosystem.

Br J Ophthalmol, 1985 Feb, 69(2), 143 - 8
Gas gangrene infection of the eyes and orbits; Crock GW et al.; The literature on Clostridium perfringens infections is reviewed up to 1983 . An additional case is reported with bilateral clostridial infections of the eye and orbit . One eye followed the classical course of relentless panophthalmitis, amaurosis, and orbital cellulitis ending in enucleation . The second eye contained intracameral mud and gas bubbles that were removed by vitrectomy instrumentation . Subsequent removal of the toxic cataract resulted in a final aided visual acuity of 6/18, N8 . This is the third report of a retained globe, and we believe the only known case where the patient was left with useful vision.

Infect Immun, 1985 Feb, 47(2), 349 - 52
Effects of Clostridium difficile toxins given intragastrically to animals; Lyerly DM et al.; We examined the activities of Clostridium difficile toxin preparations given intragastrically to hamsters, mice, and rats . The culture filtrate from a highly toxigenic strain of C . difficile caused hemorrhage and accumulation of fluid in the small intestine and cecum, diarrhea, and death in hamsters and mice . In rats, the culture filtrate caused only a small amount of fluid accumulation and slight hemorrhage along the small intestine . When toxin A was removed from the culture filtrate, the filtrate lost its activity . Preparations of homogeneous toxin A caused a response similar to that observed after the administration of culture filtrate . Hamsters were more sensitive to toxin A than mice or rats were . When hamsters were given multiple low doses of toxin A 1 week apart at a concentration which singly caused no response, they became ill and died, indicating that the toxin may have long-term effects . High amounts of toxin B did not cause any significant response when given intragastrically, unless initially mixed with low amounts of toxin A or given to hamsters with bruised ceca . These results suggest that toxins A and B act synergistically and that the action of toxin B may occur via the tissue damage caused by toxin A.

Immun Infekt, 1985 Feb, 13(1), 3 - 8
{Significance of Clostridium septicum as a cause of gas edema}; Schallehn G et al.; Gas gangrene caused by Clostridium septicum is relatively rare . In this paper 15 cases are described observed within the last 5 years . In 3 cases the gas gangrene was acquired exogenously . The other cases were endogenous infections . 9 of these cases were associated with colon carcinoma . C . septicum gas gangrene has a high lethality rate: 11 of the 15 patients died from the infection.

J Clin Microbiol, 1985 Feb, 21(2), 231 - 3
Selective isolation and rapid identification of Clostridium botulinum types A and B by toxin detection; Dezfulian M et al.; A simple procedure for rapid identification of Clostridium botulinum type A and B colonies from cultures and stool samples from infants with botulism was devised . The stool samples were directly streaked on C . botulinum isolation medium containing selective inhibitory agents . Typical lipase-positive colonies that appeared within 24 to 48 h were examined for the presence of botulinal toxin by the enzyme-linked immunosorbent assay and conventional mouse toxicity test . The amount of toxin associated with 48-h colonies of stock strains was comparable to that of 96-h broth culture . The quantity of toxin present in a single colony or combination of two was shown to be sufficient for toxin detection by the enzyme-linked immunosorbent assay . Of 42 additional stock strains tested in this manner, 41 (97.5%) were identified as toxigenic C . botulinum type A or B . The remaining one strain also proved to be toxigenic when it was tested as a concentrated cell suspension . This procedure should prove useful for large-scale serological screening of food and clinical specimens.

J Antimicrob Chemother, 1985 Feb, 15(2), 181 - 5
Antibiotic susceptibility of clinical isolates of clostridia; Brazier JS et al.; Over a 2 1/2 year period 361 clinical isolates of clostridia, representing 28 species, were tested for susceptibility to commonly used antimicrobial agents . Penicillin resistant strains were tested for their ability to produce beta-lactamase: of the commonly isolated species only Clostridium beijerinckii/butyricum produced the enzyme . Cl . perfringens exhibited a low incidence of resistance to all of the agents tested . Cl . difficile showed a high degree of resistance to penicillin and clindamycin, whilst all strains were sensitive to vancomycin . Chloramphenicol and amoxycillin/clavulanic acid were found to be highly active against most clostridia; metronidazole was the only agent to which no resistance was demonstrable.

Arch Biochem Biophys, 1985 Feb 1, 236(2), 593 - 602
Galactose-rich glycoproteins are on the cell surface of herpes virus-infected cells . 1 . Surface labeling and serial lectin binding studies of Asn-linked oligosaccharides of glycoprotein gC; Kumarasamy R et al.; Cell-surface glycoproteins of mock-infected and herpes simplex virus type 1 (HSV-1)-infected BHK-21 and HEp-2 cells were radiolabeled by incubation with galactose oxidase followed by reduction with NaB3H4 . The incorporation of radiolabel into glycoconjugates in both BHK-21 and HEp-2 cells was increased several fold following infection with HSV, showing an increase in surface-exposed Gal residues in the infected cells . This was further confirmed by an increase in binding of cell-surface-labeled glycoproteins gC and gB from HSV-infected BHK-21 cells to Ricinus communis agglutinin I, which is specific for beta-D-Gal residues . Prior treatment of cells with Clostridium perfringens neuraminidase enhanced the surface radiolabeling by the galactose oxidase/NaB3H4 method: HEp-2 cells exhibited over sixfold enhancement in labeling, while BHK-21 cells showed only a slight increase . HSV glycoprotein gC was the predominant cell-surface glycoprotein radiolabeled by the galactose oxidase/NaB3H4 method in virus-infected BHK-21 cells . The glycoprotein gC was purified by immunoaffinity column chromatography on monoclonal anti-gC-antibody-Sepharose . The radiolabel in the glycopeptides of gC was resistant to beta elimination, showing that it was associated only with Asn-linked oligosaccharides . A serial lectin affinity chromatography of glycopeptides on columns of concanavalin A-Sepharose, lentil (Lens culinaris) lectin-Sepharose, and Ricin I-agarose allowed the assignment of minimal oligosaccharide structures bearing terminal Gal residues in gC.

J Biochem (Tokyo), 1985 Feb, 97(2), 449 - 61
Studies on the interaction of Clostridium perfringens sialidase with sialic acid linked to the internal galactose in monosialogangliotetraosyl ceramide; Corfield AP et al.; Investigation of the action of highly purified Clostridium perfringens sialidase on ganglioside II3Neu5Ac-Gg4Cer and its oligosaccharide II3Neu5Ac-Gg4, in the presence and absence of sodium cholate, extend earlier results obtained with impure enzyme fractions . Sialidase labeled with 125I was found to bind to various ganglioside substrate micelles, including II3Neu5Ac-Gg4Cer, and to mixed ganglioside-sodium cholate micelles . No binding occurred between the enzyme and the ganglioside-derived oligosaccharide II3Neu5Ac-Gg4, even when radioactive II3Neu5Ac-Gg4-{3H}ol was used . The binding of sialidase to micellar substrate is a condition for enzymic hydrolysis . Correspondingly, II3Neu5Ac-Gg4Cer and II3Neu5Ac-Gg4Cer-sodium cholate micelles were hydrolyzed by the enzyme but II3Neu5Ac-Gg4 was not . Ganglioside oligosaccharide analogues containing an amino function at the reducing terminus or between two oligosaccharide chains, II3Neu5Ac-Gg4-NH2 and (II3Neu5Ac-Gg4)2NH, were hydrolyzed in the absence of cholate . A synthetic analogue of II3Neu5Ac-Gg4Cer containing only the fatty acid moiety and not the sphingosine residue (I1-deoxy-I1-stearamido-II3-monosialo-gangliotetraitol ) behaved as the ganglioside in the presence and absence of sodium cholate.

Food Chem Toxicol, 1985 Feb, 23(2), 275 - 85
Relevance of in vitro tests to in vivo acute skin inflammation: potential in vitro applications of skin keratome slices, neutrophils, fibroblasts, mast cells and macrophages; Parish WE; In vitro tests on cells or keratome slices of skin may reproduce, or indirectly reflect, the first event in acute inflammation, the cytotoxic action of an irritant on epithelial/epidermal cells . Keratome slices of human or animal skin release enzymes, show histochemical changes and demonstrate increased or decreased utilization of isotope-labelled amino acids when exposed to chemicals, including surfactants, or bacterial toxins (Clostridium perfringens) . The correlation with in vivo change is good for weak irritants and moderate to poor for strong irritants . Corrosive substances destroy the ability of the tissue to respond . Similar results have been obtained in tests on fibroblast cultures without or with an agar-keratin barrier . Neutrophils, or their separated granules, and mast cells have limited application in the prediction of chemical irritancy . The relevance and limitations of in vitro toxicological predictive tests are assessed in terms of the in vivo feature reflected by the in vitro test, the range of chemicals active in vitro compared to the in vivo responses, the ability to discriminate between intensities of reactions and the need for standards to compare results in different laboratories.

Acta Pathol Microbiol Immunol Scand {B}, 1985 Feb, 93(1), 41 - 8
The effect of Ca++ and Mg++ on the action of Clostridium perfringens enterotoxin on Vero cells; Granum PE; Clostridium perfringens enterotoxin binds to receptors on Vero cells . This process does not depend on the presence of divalent cations (Ca++, Mg++) . Binding of enterotoxin causes inhibition of 14C-leucine incorporation into proteins, probably because of depression of amino acid transport . The presence of Mg++ speeds up this effect of the enterotoxin . The enterotoxin produces membrane leakage only in the presence of Ca++, but additional Mg++ increases the rate of this process . These results indicate that the dissociation constant of the enterotoxin receptor interaction is reduced in the presence of Mg++ . A model for the mode of action of the enterotoxin is proposed.

J Hyg (Lond), 1985 Feb, 94(1), 69 - 79
The assessment and application of a bacteriocin typing scheme for Clostridium perfringens; Watson GN; A collection of 50 bacteriocins was assembled and used to type 802 isolates of Clostridium perfringens from food poisoning outbreaks and a variety of other sources . It was found that strains of the same serotype within an outbreak showed similar patterns of susceptibility to bacteriocins, and the use of "one difference' rule is proposed for interpretation of the typing patterns of epidemiologically related strains . Isolates of different serotype or of the same serotype isolated from different sources produced many variations in bacteriocin susceptibility patterns . Two computer programs were developed to assist in the interpretation of bacteriocin typing patterns . Their use showed that related and unrelated strains formed different clusters and enabled a range of the 20 most discriminatory bacteriocins to be selected . Isolates of C . perfringens from a wide range of sources were screened for their ability to produce bacteriocins . A much greater proportion of the strains from food poisoning outbreaks was bacteriocinogenic than were isolates from human and animal infections, various foods and the environment . The relevance of these findings to the occurrence of C . perfringens food poisoning is discussed.

J Bacteriol, 1985 Feb, 161(2), 636 - 40
Molecular analysis of transferable tetracycline resistance plasmids from Clostridium perfringens; Abraham LJ et al.; Conjugative tetracycline resistance plasmids from 15 Clostridium perfringens isolates from piggeries were analyzed by restriction endonuclease digestion and agarose gel electrophoresis . Seven isolates from one farm were found to carry a 47-kilobase pair (kb) plasmid, pJIR5, which had EcoRI, XbaI, and ClaI profiles that were identical to those of a previously characterized plasmid, pCW3 . An isolate from a second farm was found to carry a plasmid, pJIR6, which also was indistinguishable from pCW3 . Five additional isolates from a third farm carried a 67-kb plasmid, pJIR2, which had at least 29 kb of DNA in common with pCW3 . Finally, two isolates from a fourth farm were found to carry a 50-kb plasmid pJIR4, which appeared to consist of an entire pCW3 molecule with a 3-kb insertion . Comparative restriction maps of pCW3, pJIR2, and pJIR4 that identified the regions of homology among these plasmids were constructed . We suggest that many conjugative tetracycline resistance plasmids in C . perfringens may contain a pCW3-like core.

Proc Natl Acad Sci U S A, 1985 Feb, 82(4), 1160 - 4
Construction of the mycoplasma evolutionary tree from 5S rRNA sequence data; Rogers MJ et al.; The 5S rRNA sequences of eubacteria and mycoplasmas have been analyzed and a phylogenetic tree constructed . We determined the sequences of 5S rRNA from Clostridium innocuum, Acholeplasma laidlawii, Acholeplasma modicum, Anaeroplasma bactoclasticum, Anaeroplasma abactoclasticum, Ureaplasma urealyticum, Mycoplasma mycoides mycoides, Mycoplasma pneumoniae, and Mycoplasma gallisepticum . Analysis of these and published sequences shows that mycoplasmas form a coherent phylogenetic group that, with C . innocuum, arose as a branch of the low G+C Gram-positive tree, near the lactobacilli and streptococci . The initial event in mycoplasma phylogeny was formation of the Acholeplasma branch; hence, loss of cell wall probably occurred at the time of genome reduction to approximately to 1000 MDa . A subsequent branch produced the Spiroplasma . This branch appears to have been the origin of sterol-requiring mycoplasmas . During development of the Spiroplasma branch there were several independent genome reductions, each to approximately 500 MDa, resulting in Mycoplasma and Ureaplasma species . Mycoplasmas, particularly species with the smallest genomes, have high mutation rates, suggesting that they are in a state of rapid evolution.

J Mol Biol, 1985 Jan 5, 181(1), 147 - 9
Crystallization of an NAD+-dependent glutamate dehydrogenase from Clostridium symbiosum; Rice DW et al.; Crystals of a bacterial NAD+-dependent glutamate dehydrogenase (GDHase) have been grown over a wide range of pH values by using the hanging drop method of vapour diffusion with ammonium sulphate as the precipitant . Sodium dodecyl sulphate/polyacrylamide gel electrophoresis of this enzyme together with high pressure liquid chromatography/gel filtration, shows that this GDHase is hexameric like the GDHases of vertebrates . X-ray photographs of the crystals show that they diffract to at least 2.0 A, and an analysis of the diffraction pattern demonstrates that the hexamer is arranged in at least pseudo 32 symmetry.

Toxicon, 1985, 23(6), 903 - 12
Effect of Clostridium perfringens alpha toxin on the cardiovascular system of rats; Sakurai J et al.; Clostridium perfringens alpha toxin decreased heart rate, then elevated blood pressure, and finally caused some changes of electrocardiogram readings . The toxin decreased peripheral blood flow before blood pressure started to increase and the blood flow continued to decrease, without any affect on electrocardiogram readings, until the maximal pressure rise caused by the toxin . The toxin caused a rise in blood pressure in a dose-dependent manner . On the other hand, anti-alpha toxin antiserum inhibited both phospholipase C activity and pressor activity . When the toxin was pretreated with cysteine, calcium disodium ethylenediamine tetraacetate and calcium trisodium ethylenetriamine pentaacetate, pressor activity decreased, as well as phospholipase C activity . The results indicate that alpha toxin possesses pressor activity as well as phospholipase C activity.

Eur J Nucl Med, 1985, 11(6-7), 275 - 8
Synthesis of pyruvate-1-11C as a radiopharmaceutical for tumor imaging; Hara T et al.; Pyruvate-1-11C was prepared enzymatically by the exchange reaction of 11CO2 with the carboxyl group of pyruvic acid using pyruvate-ferredoxin oxidoreductase from Clostridium butyricum . 11C-Labeled pyruvate was purified by sublimation in specially made glassware . The radiochemical yield of pure pyruvate-1-11C was 80% 35 min after the end of bombardment . The distribution of 11C in tumor-bearing rabbits after an i.v . injection of pyruvate-1-11C was observed using a gamma camera . In contrast to normal organs, the tumor was positively visualized . We also conducted a number of successful clinical studies . A case of brain tumor which exhibited a positive image on positron-emission tomography (PET) using pyruvate-1-11C is presented.

Scand J Infect Dis, 1985, 17(3), 291 - 4
Characteristic manifestations of clostridium induced spontaneous gangrenous myositis; Narula A et al.; Spontaneous clostridial myonecrosis occurred in 30- and 69-year-old patients with pancytopenia (after treatment of acute myelogenous leukemia) and diabetes with neutropenia respectively . They presented with fever and sudden onset of pain plus tenderness in involved muscles . They rapidly deteriorated and died within hours after admission . A review of the literature for previous reports of this condition disclosed 31 additional cases . Mean age of patients was 50 years, male to female ratio was 2.2:1, and an underlying condition was present in all of them . Presenting manifestations were spontaneously occurring excruciating pain in the involved muscle (67%), generalized sepsis and shock (24%), nonpainful swelling in the involved muscles (6%) and pain, swelling and shock (3%) . Mortality rate was 91% (30/33) . The clostridial strains associated with this condition were identified in 31 cases, with Clostridium perfringens and C . septicum causing 28 of them . Bacteremia was described in 10 cases . Awareness of this rare catastrophe may aid in early recognition and surgical intervention which are essential for patient survival.

Infection, 1985, 13 Suppl 1, S46 - 9
A review of the use of cefotaxime in the treatment of skin and skin structure infections, with special reference to gram-positive pathogens; Karakusis PH et al.; Data compiled from computer-generated summaries of patient records submitted to Hoechst-Roussel Pharmaceuticals were reviewed regarding the efficacy and toxicity of cefotaxime in the therapy of skin and skin structure infections associated with gram-positive pathogens . In addition, published open and comparative trials employing cefotaxime in gram-positive and gram-negative skin infections were evaluated with respect to the pathogens isolated and the nature, severity and bacteriological and clinical outcome of the treated infections . Within the limitations of the data reviewed, cefotaxime appeared to be a safe and effective therapy in greater than 90% of infections including cellulitis, abscesses and necrotizing ulcers of the skin and subcutaneous tissues when associated with the isolation of susceptible gram-negative bacilli, methicillin-susceptible Staphylococcus aureus, or aerobic or anaerobic gram-positive pathogens susceptible to aqueous penicillin G . The data would indicate that cefotaxime is a suitable therapy for patients with presumed polymicrobial, non-crepitant infections of the skin or skin structures pending microbiological studies . However, cefotaxime cannot be recommended for similar infections due to organisms such as methicillin-resistant S . aureus or Pseudomonas aeruginosa that are commonly resistant to cefotaxime in vitro . Data regarding skin and skin structure infections associated with Clostridium spp . and enterococcal group D streptococci are either lacking or inconclusive with respect to the utility of cefotaxime.

Drugs, 1985, 29 Suppl 5, 57 - 63
Studies with temocillin in the hamster model of antibiotic-associated colitis; Boon RJ et al.; The studies reported here were designed to ascertain whether or not the new beta-lactam antibiotic, temocillin, would produce antibiotic-associated colitis in the hamster . The experiments were controlled with clindamycin and cefoxitin, which are known to induce antibiotic-associated colitis experimentally and clinically . All three antibiotics were administered to groups of animals both parenterally and orally . Clindamycin, at 1 mg/hamster, caused a slow onset of antibiotic-associated colitis by both routes, with death occurring at between 4 and 8 days . 80 to 100% of the animals had diarrhoea and showed signs of haemorrhage and caecal distension, with the caecal contents being Clostridium difficile toxin-positive . The onset of antibiotic-associated colitis after administration of cefoxitin was less marked at the 1 mg parenteral dose, with only 40% of the hamsters showing signs of colitis . At the higher doses of cefoxitin, colitis was more severe and the animals exhibited dramatic weight loss, with death occurring at between 3 and 5 days . The majority of animals had diarrhoea and were C . difficile toxin-positive; 60 to 80% also showed signs of haemorrhage and caecal distension . In contrast, the hamsters receiving temocillin remained healthy with no signs of diarrhoea, and showed consistent weight gain . No pathological abnormalities were observed and the caecal contents were toxin-negative . These results suggest that temocillin therapy in humans is unlikely to cause significant disturbance of the gastrointestinal flora.

Digestion, 1985, 32(1), 25 - 9
Effect of filtrate containing Clostridium difficile toxin on rectal mucosa maintained in organ culture; Allan A et al.; Rectal biopsies were maintained in organ culture over a 24-hour culture period, with good preservation of histological architecture . A filtrate containing Clostridium difficile toxin significantly inhibited the rise in epithelial alkaline phosphatase activity normally seen during culture . This effect was abolished by pre-incubation of the filtrate with Clostridium sordellii antitoxin, or heat inactivation . This effect is most probably due to a toxin of C . difficile . The method provides a new quantitative approach to the study of luminal toxins as possible pathogenic agents in idiopathic inflammatory diseases of the colon.

Microbiol Immunol, 1985, 29(2), 113 - 8
Germinability and heat resistance of spores of Clostridium difficile strains; Nakamura S et al.; Out of 111 Clostridium difficile strains, 108 produced spores in numbers of more than 10(5)/ml and the remaining three did not produce any spores in brain heart infusion medium . The germination frequency in the medium without lysozyme varied widely from strain to strain, ranging from less than 10(-8) to 10(0), and in 77 of the 108 strains the germination frequency was 10(-5) or less . The spores, when treated with sodium thioglycollate and then inoculated into the medium containing lysozyme, germinated in all of the 108 strains at a frequency of 10(-0.5) or more . The spores of two strains germinated at a frequency of more than 10(-0.5) in all methods . Spores of C . difficile strains were fairly highly heat-resistant; D100C values ranged from 2.5 to 33.5 min.

Scand J Infect Dis, 1985, 17(1), 77 - 82
Ceftriaxone: pharmacokinetics and effect on the intestinal microflora in patients with acute bacterial infections; Nilsson-Ehle I et al.; 12 patients with acute bacterial infections were treated with ceftriaxone, 1.5 g intravenously twice daily for 7-13 days . Pharmacokinetic variables were studied in 11 patients . In older subjects, serum half-lives were longer and serum clearances lower than in younger individuals . After the last dose, a larger increase in AUC compared to the first dose was observed in older patients and a biphasic elimination curve appeared in all patients but 2, with a terminal half-life of 15.6 h and 11.4 in old and young subjects, respectively . Estimated biliary clearances showed large individual variation, with a range of 0-16 ml/min X 1.73 m2 . Changes in the colonic microflora were pronounced . Almost total disappearance of staphylococci, streptococci and enterobacteria was found, and there was a marked tendency to overgrowth of yeasts and enterococci . One patient with the highest estimated biliary clearance of ceftriaxone developed diarrhoea after 7 days of therapy . A toxin-producing Clostridium difficile was isolated from the stool.

J Antimicrob Chemother, 1985 Jan, 15(1), 31 - 7
Interaction between penicillin, clindamycin or metronidazole and gentamicin against species of clostridia and anaerobic and facultatively anaerobic gram-positive cocci; Brook I et al.; Seven anaerobic and facultative Gram-positive cocci and 12 clostridial species were tested for in-vitro and in-vivo susceptibilities to penicillin, clindamycin, and metronidazole, used singly or in combination with gentamicin . The in-vitro tests consisted of determination of minimal inhibitory concentration (MIC), done without or with constant amounts of gentamicin . When used alone or in combination with penicillin or metronidazole, gentamicin had negligible effects on the bacteria . When used with clindamycin, gentamicin significantly reduced the MIC for one strain each of Peptococcus magnus and Clostridium difficile . The in-vivo tests were carried out in mice and consisted of studying the bacterial contents of abscesses induced by subcutaneous injection of bacterial suspensions . Synergy between gentamicin and penicillin, clindamycin or metronidazole was shown respectively in five, three and one strain . Consistency between in-vitro and in-vivo findings was present in the above mentioned strains only between gentamicin and clindamycin . The synergy between penicillin, clindamycin or metronidazole and gentamicin in Gram-positive anaerobic and facultative organisms may have clinical implications.

J Clin Pathol, 1985 Jan, 38(1), 82 - 5
Gas chromatographic identification of Clostridium difficile and detection of cytotoxin from a modified selective medium; Levett PN et al.; A modification of an existing selective medium for Clostridium difficile is described . Inclusion in the medium of DL nor-leucine and p-hydroxyphenylacetic acid enables identification of C difficile to be made directly from primary isolation plates by gas chromatographic detection of caproic acid and p-cresol . Plugs of agar withdrawn from the selective medium also allow the detection of cytotoxin production in vitro.

J Clin Microbiol, 1985 Jan, 21(1), 12 - 4
Monoclonal and specific polyclonal antibodies for immunoassay of Clostridium difficile toxin A; Lyerly DM et al.; Monoclonal antibody, affinity-purified antibody, and monospecific antiserum against toxin A were produced . The monoclonal antibody was an immunoglobulin G2a kappa chain isotype that immunoprecipitated toxin A, as shown by crossed immunoelectrophoresis . These antibodies were compared by counterimmunoelectrophoresis, latex agglutination, and indirect enzyme-linked immunosorbent assay for their sensitivity in detecting toxin A . Our findings indicate that these antibodies may be useful as immunodiagnostic reagents for Clostridium difficile disease.

Am J Obstet Gynecol, 1985 Jan 1, 151(1), 87 - 9
Pseudomembranous colitis following prophylactic antibiotic use in primary cesarean section; Arsura EL et al.; A report of a hospital outbreak of pseudomembranous colitis in three patients given prophylactic antibiotic therapy before and after primary cesarean section is presented . All patients shared the same ward and labor and delivery room, and the colitis occurred within an 8-day period . The diagnosis of pseudomembranous colitis was suspected clinically and confirmed by limited colonoscopy and biopsy followed by stool culture and toxin assay for Clostridium difficile . The high carrier rate of Clostridium difficile in the female urogenital tract and altered colonic motility during pregnancy, in addition to antibiotic use, may have contributed to the establishment of this disease . When diarrhea develops postoperatively in patients who have undergone cesarean section, pseudomembranous colitis as a potential serious complication must be kept in mind and necessary precautions taken to impede cross-contamination and development of secondary cases.

Am J Med, 1985 Jan, 78(1), 45 - 8
Clinical and endoscopic findings in patients early in the course of clostridium difficile-associated pseudomembranous colitis; Gebhard RL et al.; Endoscopic and clinical features are reported for 39 patients detected early in the course of pseudomembranous colitis . Disease was detected early by virtue of careful surveillance in patients in whom diarrhea developed . Early proctosigmoidoscopic findings in pseudomembranous colitis are illustrated . Clinical presentation includes development of fever, leukocytosis, abdominal pain, and even an ileus picture on radiography in addition to diarrhea.

J Mol Evol, 1985, 22(1), 20 - 31
New perspectives on bacterial ferredoxin evolution; George DG et al.; Recent evidence indicates that a gene transposition event occurred during the evolution of the bacterial ferredoxins subsequent to the ancestral intrasequence gene duplication . In light of this new information, the relationships among the bacterial ferredoxins were reexamined and an evolutionary tree consistent with this new understanding was derived . The bacterial ferredoxins can be divided into several groups based on their sequence properties; these include the clostridial-type ferredoxins, the Azotobacter-type ferredoxins, and a group containing the ferredoxins from the anaerobic, green, and purple sulfur bacteria . Based on sequence comparison, it was concluded that the amino-terminal domain of the Azotobacter-type ferredoxins, which contains the novel 3Fe:3S cluster binding site, is homologous with the carboxyl-terminal domain of the ferredoxins from the anaerobic photosynthetic bacteria . A number of ferredoxin sequences do not fit into any of the groups described above . Based on sequence properties, these sequences can be separated into three groups: a group containing Methanosarcina barkeri ferredoxin and Desulfovibrio desulfuricans ferredoxin II, a group containing Desulfovibrio gigas ferredoxin and Clostridium thermoaceticum ferredoxin, and a group containing Desulfovibrio africanus ferredoxin I and Bacillus stearothermophilus ferredoxin . The last two groups differ from all of the other bacterial ferredoxins in that they bind only one Fe:S cluster per polypeptide, whereas the others bind two . Sequence examination indicates that the second binding site has been either partially or completely lost from these ferredoxins . Methanosarcina barkeri ferredoxin and Desulfovibrio desulfuricans ferredoxin II are of interest because, of all the ferredoxins whose sequences are presently known, they show the strongest evidence of internal gene duplication.(ABSTRACT TRUNCATED AT 400 WORDS)

Proc Natl Acad Sci U S A, 1985 Jan, 82(2), 347 - 50
A selenium-containing nucleoside at the first position of the anticodon in seleno-tRNAGlu from Clostridium sticklandii; Ching WM et al.; In previous studies, the single selenonucleoside component of a selenium-containing tRNAGlu isolated from Clostridium sticklandii has been shown to be 5-methyl-aminomethyl-2-selenouridine . Here, we show that this selenonucleoside is most likely located at the "wobble" position of the anticodon of the clostridial seleno-tRNAGlu . Nuclease T1 digestion of this seleno-tRNAGlu generated one major selenium-containing oligonucleotide (25 bases long) . The selenium-containing residue within this oligonucleotide was located by sequence analysis of the oligonucleotide before and after removal of selenium by treatment with cyanogen bromide . The sequence of this oligonucleotide, A-A-C-C-G-C-C-C-U-U+-U-C-A+C-G-G-C-G-G-U-A-A-C-A-G, is homologous to that of the Escherichia coli tRNAGlu2 from residues 27 to 50, including the anticodon region and the variable loop, except that the E . coli tRNA has 5-methylaminomethyl-2-thiouridine instead of the selenonucleoside.

Drugs Exp Clin Res, 1985, 11(3), 201 - 5
Cefuroxime versus ceftriaxone prophylaxis in cardiovascular surgery; Geroulanos S et al.; Publication Types:

	Clinical Trial
	Randomized Controlled Trial

What Is Biotechnology?, What Is Listeria Monocytogenes?, What Is Amino Acid?, What Is Salmonella?, What Is Molecular Microbiology?, o, Microorganism, r, Microbe, o, Microbiology, o, Bacteria, a, Bacterium, i, Pseudomonas aeruginosa, r, Bacteria, r, Pseudomonas, e, Antimicrobials, o, Bacteriophage, a, Pseudomonas, n, Bacillus anthracis, i, Escherichia coli, e, Microorganisms, o, Staphylococcus, e, Neisseria, c, Campylobacter, o, Minimal inhibiting concentration, s, Enterobacteriacea, a, Staphylococcus, e, Lactobacillus, o, Lactobacillus, i, Haemophilus, o, Microorganisms, a, Escherichia coli, i, Antibiotics

Scientific Publications - Work Done by Microbiology Reader Bioscreen C
Agricultural Microbiology Anaerobic Microbiology Antimicrobial Susceptibility Artificial Atmosphere Bioassay of Antibiotics Biofilm Microbiology Bioreactor Technology Biotechnology Cell Biology Clinical Microbiology Environmental Microbiology Experiments with Yeast	Fermentation Food Microbiology Functional Genomics Gene Technology Growth Media Development Growth Rate and Lag Time Industrial Microbiology Medical/Pharmaceutical Field Microbiological Assay Microbiological Research Microbiology of Cosmetics go to a specific theme...	Military Microbiology Molecular Microbiology Mutagenicity and Genotoxicity Oral Microbiology Patents Postantibiotic Studies Soil Microbiology Spore Microbiology Veterinary Microbiology Waste/Wastewater Treatment Water Microbiology Wine Microbiology

� 2005 Transgalactic Ltd (manufacturer of Bioscreen C software) | Privacy Statement | P.O. Box 1393, 00101 Helsinki, Finland, phone: +358 9 85172920, fax: +358 9 8749481, e-mail: microbiology@bionewsonline.com

Last modified: May 25, 2005