Lab Anim Sci, 1987 Oct, 37(5), 652 - 6 An evaluation of ampicillin pharmacokinetics and toxicity in guinea pigs; Young JD et al.; Sodium ampicillin was administered subcutaneously to 350-550 g male Dunkin Hartley guinea pigs at doses of 6, 8 and 10 mg/kg tid for 5 days . Over a period of 12 days, the lowest ampicillin dose appeared to be tolerated well . However, significant body weight reduction and mortality occurred with the two higher dosage regimens . Cecal cultures of dead animals confirmed the presence of Clostridium difficile, an organism associated with antibiotic-induced enterotoxemia . Assay of serum collected from ampicillin-treated animals revealed ampicillin concentrations of approximately 10 micrograms/ml at 5 minutes post-dosing which fell precipitously to less than 0.2 micrograms/ml at 60 minutes . Determination of biliary ampicillin levels during the 60 minutes after administration of a single 10 mg/kg SQ dose revealed concentrations ranging from 18 micrograms/ml to 90 micrograms/ml . Estimates of total urinary ampicillin content after a single 10 mg/kg SQ dose were less than 500 micrograms/animal at 7.5 minutes, but increased to greater than 2000 micrograms/animal at 60 minutes after dosing . Results of this study indicated that due to its short serum half-life, sodium ampicillin probably has little systemic therapeutic efficacy in guinea pigs . Because high concentrations of ampicillin accumulated in the urine and bile, the antibiotic probably would have therapeutic efficacy for urinary and intestinal infections . However, its associated toxicity at large doses probably precludes its use . In view of the rapid clearance of ampicillin in guinea pigs in comparison to other species, the pharmacokinetics of other antibiotics, especially those reported to be less toxic for guinea pigs, should be considered. J Antibiot (Tokyo), 1987 Oct, 40(10), 1375 - 82 Coloradocin, an antibiotic from a new Actinoplanes . I . Taxonomy, fermentation and biological properties; Jackson M et al.; Coloradocin was discovered in a screen for anti-anaerobe activity . The producing organism was determined to be a new species of Actinoplanes, designated Actinoplanes coloradoensis sp . nov . Coloradocin inhibits Bacteroides, Clostridium and other anaerobes . It does not inhibit most aerobic bacteria but is effective against Neisseria gonorrhoeae and Haemophilus influenzae . Coloradocin has low acute toxicity. J Clin Microbiol, 1987 Oct, 25(10), 1999 - 2000 Detection of Clostridium difficile toxin in various tissue culture monolayers; Maniar AC et al.; Thirty stool filtrates known to contain Clostridium difficile toxin based on previous testing on McCoy cells were tested for toxicity on primary African green monkey kidney (AGMK), McCoy, MRC-5, primary rhesus monkey kidney (RMK), and Vero cells . All 30 filtrates showed cytotoxic effect at greater than or equal to 1:100 dilution on McCoy and Vero cells . A total of 22 filtrates were positive on MRC-5 monolayers, while only 16 and 10 filtrates showed positive cytotoxic effect on AGMK and RMK cells, respectively . Another 630 stool specimens were tested on McCoy and Vero cells only . Of these stool filtrates, 70 were positive and 560 were negative with both cell lines, which thus gave 100% agreement . Vero cells can be used interchangeably with McCoy cells for the detection of C . difficile toxin in stool filtrates. South Med J, 1987 Oct, 80(10), 1318 - 20 Splenic abscess due to Clostridium septicum in a patient with multiple myeloma; Kinnaird DW et al.; Solitary abscesses of the spleen may occur as a consequence of septicemia from various septic foci, particularly intra-abdominal sites . A variety of causative organisms, including the clostridia, have been isolated . We have described a patient with multiple myeloma who had a solitary splenic abscess due to Clostridium septicum and who failed to survive despite seemingly appropriate treatment . Clinicians should be alert to this complication, which demands immediate splenectomy and appropriate antibiotic therapy. Mayo Clin Proc, 1987 Oct, 62(10), 901 - 5 Vancomycin; Hermans PE et al.; Vancomycin is a narrow-spectrum bactericidal antibiotic used primarily for treatment of serious staphylococcal infections . It is the alternative therapy of choice when the penicillins and cephalosporins cannot be used . Vancomycin is also used in (1) methicillin-resistant Staphylococcus aureus infections; (2) streptococcal endocarditis in conjunction with an aminoglycoside in patients intolerant of penicillin or ampicillin; (3) infections, including those involving prosthetic devices, caused by gram-positive organisms with multiple antibiotic resistance; (4) antibiotic-induced enterocolitis caused by Clostridium difficile; and (5) prophylaxis for endocarditis in patients who are at risk and cannot tolerate a penicillin, cephalosporin, or erythromycin . The major toxic effect associated with the use of vancomycin is ototoxicity, which may develop when serum levels exceed 30 micrograms/ml. Clin Orthop, 1987 Oct, (223), 194 - 7 Delayed postbacteremic prosthetic joint infection; Maniloff G et al.; Deep infection of a prosthetic joint is a devastating complication . One proposed mechanism of late prosthetic joint infection involves hematogenous spread from an extraarticular focus of infection . Two cases clearly demonstrate hematogenously acquired prosthetic joint infections, one caused by Clostridium perfringens and the other by Streptococcus pneumoniae . These cases were unusual in that a long asymptomatic period intervened between the primary bacteremic illness and the subsequent prosthetic infection . Patients with prosthetic joints who develop bacteremic infection at extraarticular sites should be treated promptly and aggressively with appropriate antibiotics . Prophylactic antibiotics should be strongly considered in the patient with a prosthetic joint who undergoes procedures likely to be associated with a bacteremia . Transient arthralgias at the time of bacteremia may represent the onset of the joint infection and should not be overlooked or attributed a priori to the patient's underlying arthritic or medical condition. Am J Clin Oncol, 1987 Oct, 10(5), 451 - 2 Clostridium difficile colitis induced by cytarabine; Roda PI; Pseudomembranous enterocolitis (PMC) has become a widely recognized syndrome of nausea, abdominal distention, and severe (frequently bloody) diarrhea (1) . While this syndrome was first associated with the administration of clindamycin, almost all antimicrobial drugs can serve as predisposing agents (2) . We wish to report a patient with typical PMC induced by the administration of cytarabine. Cornell Vet, 1987 Oct, 77(4), 328 - 38 Successful management of malignant edema caused by Clostridium septicum in a horse; Perdrizet JA et al.; The clinical course and successful therapeutic management of a horse with malignant edema caused by Clostridium septicum is described . This is believed to be the first report of a horse surviving malignant edema caused by C . septicum . A discussion of this disease syndrome, including etiology, pathogenesis, clinical signs, therapy, and diagnostic methods is presented. Vet Microbiol, 1987 Oct, 15(1-2), 115 - 20 Production of iota toxin by Clostridium spiroforme: a requirement for divalent cations; Carman RJ et al.; The effects of divalent cations (Ca2+, Co2+ and Zn2+) on the production of iota toxin by Clostridium spiroforme were studied . Toxin production had an absolute requirement for one or more cations in the range 1-5 mM . Using bispecific antisera, we showed that production of both the components of the toxin (ia and ib) were enhanced by divalent cations added to brain-heart infusion supplemented with peptone and glucose. Eur J Clin Microbiol, 1987 Oct, 6(5), 542 - 6 Epidemiological aspects of Clostridium difficile in a pediatric hospital and its role in diarrheal disease; Camorlinga-Ponce M et al.; The influence of antibiotics on the frequency of colonization by Clostridium difficile and the presence of its cytotoxin in infants and older children was examined to determine its role in diarrheal disease . Cytotoxin was more closely associated with cases of diarrhea, both in infants and in children than the microorganism, although not significantly . The isolates were typed by means of sensitivity to bacteriophages and bacteriocins and their cytotoxigenic potential was also determined . Less than 30% of the colonized patients had toxigenic strains . A study of strain variability over a four-year period in the same hospital showed that two bacteriophage-bacteriocin types and non-toxigenic strains predominated . The common presence of non-toxigenic strains could explain in part the lack of correlation between isolation of Clostridium difficile and diarrhea . Most of the non-toxigenic strains showed moderate resistance to tetracycline, a property which might explain their ability to persist for long periods in the hospital. Microb Pathog, 1987 Oct, 3(4), 279 - 86 Response of tissue-cultured cynomolgus monkey kidney cells to botulinum C2 toxin; Miyake M et al.; C2 toxin (C2T) elaborated by Clostridium botulinum types C and D is composed of two nonlinked protein components, designated components I and II . The toxin, a mixture of untrypsinized component I and trypsinized component II, induced marked morphological changes of tissue-cultured cynomolgus monkey kidney cells; the characteristic response of the cells to the toxin was rounding, which increased proportionally to log dose of the toxin . The components alone and a combination of untrypsinized components I and II showed little activity . The rounding of the cultured cells was not accompanied by inhibition of protein and nucleic acid syntheses of the cells, although the rounded cells ultimately lost viability . Immunofluorescence studies showed that component II, either trypsinized or untrypsinized, bound to the cell surface, whereas component I bound to the cells only in the presence of trypsinized component II . The present results support the previously proposed idea concerning the mode of action of C2T, that components I and II of C2T act together as a molecule with dual functions; component II as the recognizer of the receptor site on the cell surface membranes and component I as the effector in the cytoplasm by preferential inactivation of cytoskeletal actin, which results in alteration of cell morphology, and subsequently in cellular damage. Epidemiol Infect, 1987 Oct, 99(2), 355 - 9 Comparison of alcohol shock enrichment and selective enrichment for the isolation of Clostridium difficile; Riley TV et al.; Two enrichment methods were compared for their ability to recover Clostridium difficile from stool samples . One method used selective enrichment in an antibiotic-containing broth followed by detection with a latex particle agglutination (LPA) reagent . The other used enrichment in a non-selective broth following treatment of the specimen with alcohol . With clinical specimens enrichment culture was significantly more successful at detecting C . difficile than direct plating . Alcohol shock enrichment was twice as effective as direct culture, while selective broth enrichment was three times more effective . The use of LPA for screening selective enrichment broths for C . difficile should prove a cost-effective measure as only positive broths (about 20%) require subculture for confirmation. Avian Dis, 1987 Oct-Dec, 31(4), 904 - 6 Experimentally induced necrotic enteritis in chickens; Cowen BS et al.; A 1.3 to 10% incidence of necrotic enteritis was experimentally produced in broiler-type chickens in three of five trials . The incidence range observed was considerably narrower and lower than the 5.6-37.3% range reported in the literature . Clostridium perfringens was inconsistently isolated from the liver and intestine of dosed chickens. Med Trop (Mars), 1987 Oct-Dec, 47(4), 385 - 7 {Pseudomembranous colitis after antibiotic therapy associated with the presence of Entamoeba histolytica histolytica . Apropos of 2 cases}; Peghini M et al.; The authors report on two cases of pseudomembranous colitis (P.M.C.) developed in two Senegalese women of 38 and 36 years, and discovered at the 4th and 5th day respectively of an antibiotherapy based on ampicillin . In these two observations, cysts of Entamoeba histolytica histolytica were found in both feces and biopsies . They recall the circumstances of the occurrence, diagnosis techniques and treatment . They underline the unfrequency of this disease in Africa south of Sahara and they discuss the correlation with amoebiasis colitis . One has to keep in mind the possibility of a P.M.C . during any antibiotherapy, and consequently to have a rectoscopy to perform . Such an exploration is enough to pose a diagnosis . In day to day practice it is not necessary to show clearly the specific germ Clostridium difficile or its entero-toxin . To stop any antibiotherapy is required and beneficial . Metronidazole or Vancomycin are the best drugs in this case. Z Lebensm Unters Forsch, 1987 Oct, 185(4), 281 - 7 {Anaerobic spore formers in commercial spices and ingredients for infant food}; Eisgruber H et al.; Anaerobic spore-formers can cause food intoxication . Clostridium (Cl.) perfringens type-A strains are mainly involved . In addition to the original contamination of food, e.g., meat or milk, ingredients such as spices or meat extracts are the main source and low numbers of clostridia are present (i.e., up to 10(2) cells/g) . At present, information about the quantitative ecology of clostridia is incomplete; therefore 115 commercial food products were investigated (spices: n = 70, meat extracts: n = 15, instant foods: n = 30) . For the detection of low numbers of clostridia (less than 10(3)/g), the most probable number (MPN) technique was used . For the detection of higher numbers (greater than 10(3)/g) a cultivation method in solid media was developed . The mesophilic sulphite-reducing clostridia spores were detected by the pour-plating method with overlayer . Twenty-one of the 70 commercial spices, 7 of the 30 specimen of instant food and 12 of the 15 meat extracts were clostridia-positive . The low counts ranged between 3 X 10(0) and 10(3) cells/g, in most cases (2/3) in the range 10(1)/10(2)/g . Only in 6 cases were more than 10(3) clostridia/g detected . Most of those isolated were identified as Cl . perfringens (46 of 73 isolates) . Only in meat extracts did clostridia species other than Cl . perfringens strains predominate . Three of the 23 Cl . perfringens strains proved to be heat-resistant . A hydrated meat extract originally contaminated with low numbers was tested in a pilot study for the enrichment of clostridia . This extract showed an increase of up to 10(7) cells/ml after a 24-h incubation period.(ABSTRACT TRUNCATED AT 250 WORDS) J Biol Chem, 1987 Sep 15, 262(26), 12488 - 95 Purification and properties of an extracellular collagenolytic protease produced by the human oral bacterium Bacillus cereus (strain Soc 67); Makinen KK et al.; The major collagenolytic proteinase present in the culture filtrate of Bacillus cereus (strain Soc 67, isolated from the human oral cavity) has been purified to homogeneity by a procedure that comprised concentration of ultrafiltered growth medium on a Millipore PTTK00005 membrane, precipitation with ammonium sulfate, gel permeation chromatography, chromatofocusing, fast protein liquid chromatography on an anion-exchange column, and finally fast protein liquid chromatography on a gel column . The enzyme hydrolyzed, with decreasing rates, phenylazobenzyloxy-carbonyl-L-Pro-L-Leu Gly-L-Pro-D-Arg (PZ-PLGPA), furylacrylolyl-L-Leu-Gly-L-Pro-L-Ala, and furylacryloyl-L-Phe-Gly-Gly, while furylacryloyl-Gly-L-Leu-NH2 was not hydrolyzed . The enzyme degraded soluble and insoluble collagens, Azocoll and gelatin . Bradykinin was hydrolyzed at a high rate at the Phe-Ser bond . The enzyme was sensitive to pyrophosphate, L-cysteine, and L-histidine and could be totally inactivated in the presence of metal chelators . The enzyme contains 1 mol of Zn/mol and the hydrolysis of PZ-PLGPA is slightly increased by Ca2+ . The enzyme is readily inhibited by heavy metal cations, but Cu2+ and Ni2+ affected the catalysis in opposite ways: increasing levels of Cu2+ decreased the affinity of the enzyme for PZ-PLGPA, whereas Ni2+ had no effect . The effect of Cu2+ also depended on the pH and type of buffer used . Detailed chemical modification experiments suggested that the active site of the enzyme contains at least 1 tyrosyl and 1 lysyl residue, and 1 carboxyl group . The enzyme was not sensitive to sulfhydryl reagents and thiols did not activate the enzyme . The modification studies were unable to reveal active histidyl residues . The ability of the enzyme to hydrolyze PZ-PLGPA, furylacryloyl-L-Leu-Gly-L-Pro-L-Ala, furylacryloyl-L-Phe-Gly-Gly, and various collagenous materials, its inactivity toward furylacryloyl-Gly-L-Leu-NH2, and the results from the chemical modification studies suggest that the B . cereus (Soc 67) collagenolytic enzyme can be regarded as a true collagenase which resembles the Clostridium histolyticum collagenase(s). Eur J Biochem, 1987 Sep 15, 167(3), 425 - 30 Role of the essential thiol group in the thiol-activated cytolysin from Clostridium perfringens; Iwamoto M et al.; A hemolysin, 0-toxin, produced by Clostridium perfringens has one cysteinyl residue in the free thiol form which is essential for its hemolytic activity . The cysteinyl residue was shown to be located at a position about 5 kDa from the C terminus of the molecule by the method of cysteine-specific chemical cleavage . Modification of the residue with a thiol-blocking agent, 5,5'-dithiobis(2-nitrobenzoic acid), reduced the binding affinity of the toxin to sheep erythrocytes to 1/100 that of intact toxin, resulting in a failure of binding at low cell concentrations (0.5%) . Thus the failure of hemolysis at low cell concentrations is primarily ascribed to a decreased affinity of the toxin for erythrocytes . Effects of the modification on the lytic processes were examined using high cell concentrations where considerable amounts of modified toxin bound to the cells . The modified toxin hemolyzes erythrocytes once it binds to them; however, the efficiency of hemolysis is reduced by the modification . These, and additional results indicating that modification alters the sensitivity of toxin molecules to protease digestion, show that thiol-modification inactivates the toxin by affecting both binding and the subsequent lytic processes, probably through a conformational change introduced in the toxin molecules. Biochemistry, 1987 Sep 8, 26(18), 5826 - 33 Deuterium nuclear magnetic resonance studies on the plasmalogens and the glycerol acetals of plasmalogens of Clostridium butyricum and Clostridium beijerinckii; Malthaner M et al.; Deuterium nuclear magnetic resonance was used to investigate the structure of different lipid fractions isolated from the anaerobic bacteria Clostridium butyricum and Clostridium beijerinckii . The fractions isolated from C . butyricum were (1) phosphatidylethanolamine/plasmenylethanolamine and (2) the glycerol acetal of plasmenylethanolamine, and from C . beijerinckii similar fractions containing principally (1) phosphatidyl-N-monomethylethanolamine, along with its plasmalogen, and (2) the glycerol acetal of this plasmalogen were isolated . The third fraction from both species consisted largely of the acidic lipids phosphatidylglycerol and cardiolipin along with plasmalogen forms of these lipids . Palmitic acid with deuterium labels at C-2, C-3, or C-4 or oleic acid with deuterium labels at C-2 and C-9,10 was added to the growth medium and incorporated to various extents in the lipid fractions . Biochemical analysis showed that palmitic acid and oleic acid were preferentially bound to the sn-2 and sn-1 positions, respectively, of the glycerol backbone when both fatty acids were added to the medium . From the 2H NMR spectra, the hydrocarbon chain ordering near the lipid-water interface could be determined and appeared to be similar for all three lipid fractions . The deuterium quadrupole splitting and order parameter were low at the C-2 segment and increased by almost a factor of 2 at positions C-3 and C-4 for cells fed with deuterated palmitic acid along with unlabeled oleic acid . These results agree with previous findings on pure diacyl lipids in which the sn-2 chain was found to adopt a bent conformation at the carbon segment C-2 . However, two unusual quadrupole splittings could be detected for the plasmalogens.(ABSTRACT TRUNCATED AT 250 WORDS) Rev Infect Dis, 1987 Sep-Oct, 9(5), 1038 - 43 Anaerobic bacteremia in a general hospital: retrospective five-year analysis; Vazquez F et al.; Anaerobic bacteremia (116 cases) represented 5.4% of the total cases of bacteremia in the Hospital Nuestra Senora de Covadonga of Oviedo, Spain, during a five-year period (1981-1985) . Microbiologic data for all 116 cases and clinical data for 63 patients were analyzed . A total of 129 isolates were identified as gram-negative bacilli (45.7%), gram-positive bacilli (38.0%), gram-positive cocci (14.0%), and gram-negative cocci (2.3%) . Bacteroides fragilis and Clostridium perfringens were the most frequently occurring species . Anaerobic polymicrobial infection was detected in 21 patients . The most relevant clinical features were fever (79%), metastatic abscesses (33%), anemia (27%), septic shock (25%), and disseminated intravascular coagulation (6%) . The overall mortality rate was 25.4%, and the factors associated with a poor prognosis were age over 60 years, lack of adequate surgical treatment, severe underlying disease, metastatic foci, and polymicrobial and/or nosocomial infection. J Antimicrob Chemother, 1987 Sep, 20(3), 379 - 82 The in-vitro activity of doxycycline and minocycline against anaerobic bacteria; Robbins M et al.; The likelihood of bacterial resistance now prevents the use of oxytetracycline in the empirical therapy of anaerobic infections . This study investigates the in-vitro activity of two semi-synthetic derivatives, doxycycline and minocycline, against a range of anaerobic bacteria . MICs for each antibiotic were determined by an agar incorporation technique . Doxycycline and minocycline were four to eight times more active against the majority of strains than oxytetracycline . With the exception of Bacteroides bivius, almost 90% of strains were inhibited by 4 mg/l of doxycycline or minocycline, but resistance to the same concentration of oxytetracycline was present in 60% of the B . fragilis group, 30% of Peptostreptococcus spp . and 24% of Clostridium perfringens . Doxycycline and minocycline represent an alternative therapy for anaerobic infections where bacterial sensitivities are known. Am J Forensic Med Pathol, 1987 Sep, 8(3), 259 - 62 Homicidal death following blunt trauma in a vulnerable host, with secondary infections including local tetanus; Murphy GK; An uncommon type of homicide resulted from complications of an ordinarily nonfatal injury after a 59-year-old obese, hypertensive, diabetic man was struck in the face with a two-by-four, sustaining a grossly contaminated laceration . It was cleaned and sutured primarily, and a tetanus booster was given . On the fourth hospital day there was evidence of anerobic wound cellulitis, including Clostridium tetani . The wound was surgically debrided, but 2 days later the patient developed local tetanus . Only then was it discovered that he had never been immunized against tetanus . He did not develop systemic tetanus, but 2 days later he died with bronchopneumonia and sepsis . The assailant was indicted for involuntary manslaughter, but after a contentious trial he pleaded "no contest" to a reduced charge . The decedent was a vulnerable host, his contaminated facial laceration initiating an unbroken course of events that led to his death. J Gen Microbiol, 1987 Sep, 133 ( Pt 9), 2647 - 57 Binding and cytotoxic effects of Clostridium botulinum type A, C1 and E toxins in primary neuron cultures from foetal mouse brains; Kurokawa Y et al.; Binding of purified Clostridium botulinum type A, C1 and E toxins to cultured cells was studied by an immunocytochemical method . Type A and C1 toxins bound strongly to neuron cultures prepared from brains of foetal mice, but binding of type E toxin was weak . None of the toxin types bound to the feeder layer, composed of non-neuronal cells . The heavy-chain component of the type C1 toxin bound to neurons, but the light chain component did not . Type C1 toxin also bound only to cell lines of neuronal origin . When type C1 toxin {final concentration 4 x 10(2) LD50 (10 ng) per well} was added to primary neuron cultures in 96-well plates, degeneration of neuronal processes and rounding of neuronal somas were observed, but type A and E toxins did not produce such changes . The binding and cytotoxic activities of type C1 toxin were blocked by heat treatment (80 degrees C for 30 min) or by preincubation of the toxin with polyclonal anti-C1 IgG and some of the monoclonal antibodies which neutralized the toxin activity in mice . In the neuronal processes treated with C1 toxin, many degenerated mitochondria, membranous dense bodies and vesicles were observed by electron microscopy; these ultrastructural changes were similar to those of Wallerian degeneration in vivo. J Appl Bacteriol, 1987 Sep, 63(3), 217 - 26 Evaluation of a monoclonal antibody-based immunoassay for detecting type A Clostridium botulinum toxin produced in pure culture and an inoculated model cured meat system; Gibson AM et al.; A monoclonal antibody-based amplified enzyme-linked immunosorbent assay (ELISA) method for detecting Clostridium botulinum type A toxin was evaluated for its ability to detect the toxin in the supernatant fluid of pure cultures and after growth from Cl . botulinum spores inoculated into pork slurries . Slurries containing NaCl (1.5-4.5% w/v) and polyphosphate (0.3% w/v) were either unheated or heated, 80 degrees C/5 min + 70 degrees C/2 h, before storage at 15 degrees, 20 degrees or 27 degrees C . The presence of specific toxin was confirmed by mouse bioassay and results compared with those of the amplified ELISA method . A total of 49 strains, 39 Cl . botulinum and 10 Cl . sporogenes (putrefactive anaerobes), and 95 slurry samples were tested . Fourteen of 15 strains of type A Cl . botulinum and 34 of 36 slurry samples containing type A toxin were positive by ELISA . No false positive reactions occurred with Cl . botulinum types B, C, D, E and F, or with the 10 strains of Cl . sporogenes . However, toxin produced by one strain of Cl . botulinum type A (NCTC 2012) was not detected by the amplified ELISA. J Clin Pathol, 1987 Sep, 40(9), 1070 - 87 Automation in clinical microbiology: a new approach to identifying micro-organisms by automated pattern matching of proteins labelled with 35S-methionine; Tabaqchali S et al.; A new rapid automated method for the identification and classification of microorganisms is described . It is based on the incorporation of 35S-methionine into cellular proteins and subsequent separation of the radiolabelled proteins by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) . The protein patterns produced were species specific and reproducible, permitting discrimination between the species . A large number of Gram negative and Gram positive aerobic and anaerobic organisms were successfully tested . Furthermore, there were sufficient differences within species between the protein profiles to permit subdivision of the species . New typing schemes for Clostridium difficile, coagulase negative staphylococci, and Staphylococcus aureus, including the methicillin resistant strains, could thus be introduced; this has provided the basis for useful epidemiological studies . To standardise and automate the procedure an automated electrophoresis system and a two dimensional scanner were developed to scan the dried gels directly . The scanner is operated by a computer which also stores and analyses the scan data . Specific histograms are produced for each bacterial species . Pattern recognition software is used to construct databases and to compare data obtained from different gels: in this way duplicate "unknowns" can be identified . Specific small areas showing differences between various histograms can also be isolated and expanded to maximise the differences, thus providing differentiation between closely related bacterial species and the identification of differences within the species to provide new typing schemes . This system should be widely applied in clinical microbiology laboratories in the near future. Microbiol Sci, 1987 Sep, 4(9), 277 - 80 The cloned cel (cellulose degradation) genes of Clostridium thermocellum and their products; Beguin P et al.; More than 10 cel genes of Clostridium thermocellum were cloned and expressed in Escherichia coli . Four of them, celA, celB, celC and celD, were studied in detail . The corresponding endoglucanases were purified from E . coli and characterized . The over-produced endoglucanase D, purified from cytoplasmic granules, was crystallized. Microb Pathog, 1987 Sep, 3(3), 195 - 206 Interferon pretreatment enhances the sensitivity of Vero cells to Clostridium perfringens type A enterotoxin; McClane BA et al.; Treatment of Vero (African green monkey kidney) cells with interferon (IFN) before the addition of Clostridium perfringens type A enterotoxin (CPE) significantly increased the sensitivity of these cells to CPE . IFN pretreatment caused a subsequent two- to four-fold increase in CPE-induced membrane permeability alterations and also decreased the time of CPE treatment required before the onset of permeability alterations and morphologic damage . Enhancement of CPE activity was dependent on the amount of IFN added during pretreatment and on the duration of IFN pretreatment incubations . Potentiation of CPE activity was observed following pretreatment of Vero cells with natural human IFN-alpha or IFN-gamma or Roferon recombinant human IFN-alpha . However, pretreatment with mouse IFN did not affect CPE activity . IFN pretreatment did not grossly enlarge the size of the functional hole produced in plasma membranes by CPE . IFN pretreatment of Vero cells slightly increased CPE specific binding, but this effect occurred kinetically after the enhancement of CPE toxicity . These results suggest that IFN pretreatment enhances the action of CPE on IFN pretreated Vero cells by increasing the sensitivity of these cells to the action of CPE rather than by increasing CPE specific binding or by directly activating the CPE molecule . Additional studies are required to further clarify the mechanism by which IFN sensitized Vero cells to CPE. Can J Microbiol, 1987 Sep, 33(9), 773 - 6 Changes in the hydrophobic characteristics of Clostridium perfringens spores and spore coats by heat; Craven SE et al.; The hydrophobic characteristics of Clostridium perfringens NCTC 8679 spores were demonstrated by adherence to toluene in a toluene-aqueous partition system . Spores and spore coat preparations were hydrophobic . Vegetative cells and spores extracted with a dithiothreitol-sodium dodecyl sulfate treatment known to remove spore coats were not hydrophobic . A heat activation treatment (75 degrees C for 20 min) which promotes more rapid spore germination increased the hydrophobicity of intact spores and decreased that of isolated spore coats . The hydrophobic changes were reversed by washing and stabilized by 0.5% glutaraldehyde . Heat-induced hydrophobic changes were observed in spore coats prepared from spores that were preheated and washed before rupturing in a buffer containing glutaraldehyde . These results suggest the occurrence of a heat-induced change in the spore coat (possibly in the conformation of a macromolecule) which was stable only within the architectural confines of the intact spore. Biull Eksp Biol Med, 1987 Sep, 104(9), 376 - 8 {Use of collagenase from the hepatopancreas of the Kamchatka crab for isolating and culturing endothelial cells of the large vessels in man}; Goncharov NV et al.; The application of hepatopancreas collagenase from crab Paralithodes camtschatica for the isolation and cultivation of the endothelial cells was studied in human umbilical vein endothelium . The comparison of the enzyme from crab hepatopancreas with collagenase from Clostridium histoliticum has shown that the number of viable cells isolated from human umbilical vein by the crab enzyme was lower than in the case of microbial collagenase . However, this difference was not significant for subsequent cultivation of cells . Harvesting of the endothelial cells from the substrate during cultivation was more effective in the case when collagenase from crab hepatopancreas was used . It was shown that crab collagenase, in contrast with microbiological collagenase, was not a metal-dependent enzyme. Lancet, 1987 Aug 29, 2(8557), 496 - 500 Enteritis necroticans among Khmer children at an evacuation site in Thailand; Johnson S et al.; A severe illness characterised by bloody diarrhoea and intestinal dysfunction was recognised at an evacuation site on the Thai-Kampuchean border . From June, 1985, to July, 1986, the illness occurred in 62 Khmer children aged 10 months to 10 years (mean 4 years); it was characterised by bloody diarrhoea (94%), fever (90%), and abdominal pain (78%) . The overall mortality rate was 58% . Among 16 children who died and underwent necropsy, small-intestinal necrosis of varying severity was found; in 5 of these children small-intestinal lesions with areas of full-thickness necrosis were seen that histologically resembled those in cases of enteritis necroticans (pigbel) in Papua New Guinea . Beta-toxin-producing Clostridium perfringens type C was isolated from 2 of 23 children from whom specimens for anaerobic cultures were collected, and antibodies to beta toxin were detected in 5 of 9 survivors but not in 10 healthy, age-matched control children . These cases show that enteritis necroticans can cause substantial morbidity and mortality outside Papua New Guinea. Eur J Biochem, 1987 Aug 17, 167(1), 175 - 80 A 50-kDa fragment from the NH2-terminus of the heavy subunit of Clostridium botulinum type A neurotoxin forms channels in lipid vesicles; Shone CC et al.; 1 . A 50-kDa fragment representing the NH2-terminus of the heavy subunit of botulinum type A neurotoxin was found, at low pH, to evoke the release of K+ from lipid vesicles loaded with potassium phosphate . Similar K+ release was also observed with the intact neurotoxin, its heavy chain and a fragment consisting of the light subunit linked the 50-kDa NH2-terminal heavy chain fragment . The light subunit alone, however, was inactive . 2 . In addition to K+, the channels formed in lipid bilayers by botulinum neurotoxin type A or the NH2-terminal heavy chain fragment were found to be large enough to permit the release of NAD (Mr 665) . 3 . The optimum pH for the release of K+ was found to be 4.5 . Above this value K+ release rapidly decreased and was undetectable above pH 6.0 . 4 . The binding of radiolabelled botulinum toxin to a variety of phospholipids was assessed . High levels of toxin binding were only observed to lipid vesicles with an overall negative charge; much weaker binding occurred to lipid vesicles composed of electrically neutral phospholipids . 5 . A positive correlation between the efficiency of toxin-binding and the efficiency of K+ release from lipid vesicles was not observed . Whereas lipid vesicles containing the lipids cardiolipin or dicetyl phosphate bound the highest levels of neurotoxin, the toxin-evoked release of K+ was low compared to vesicles containing either phosphatidyl glycerol, phosphatidyl serine or phosphatidyl inositol . 6 . The implications of these observations to the mechanism by which the toxin molecule is translocated into the nerve ending are discussed. Biochem J, 1987 Aug 15, 246(1), 193 - 7 Characterization of alpha-amylase and pullulanase activities of Clostridium thermohydrosulfuricum . Evidence for a novel thermostable amylase; Melasniemi H; Thermostable extracellular alpha-amylase and pullulanase activities of Clostridium thermohydrosulfuricum E 101-69 were characterized in a crude enzyme preparation . The activities responded similarly to temperature and pH, with optima at 85-90 degrees C and pH 5.6 . The activities were stable at 65 degrees C, but were inactivated gradually in an identical manner at higher temperatures in the absence of Ca2+ and substrate . Ca2+ stabilized both activities similarly at high temperatures . Ca2+ also stimulated both activities, whereas EDTA reversed this stimulation . The activities were similarly inactivated at pH extremes . The two activities distributed in the same way during isoelectric focusing . The results suggest that the two activities are properties of the same protein, representing a novel, thermostable, amylase. Arch Biochem Biophys, 1987 Aug 15, 257(1), 217 - 29 Toxin A from Clostridium difficile binds to rabbit erythrocyte glycolipids with terminal Gal alpha 1-3Gal beta 1-4GlcNAc sequences; Clark GF et al.; The binding of Toxin A isolated from Clostridium difficile to rabbit erythrocyte glycolipids has been studied . Total lipid extracts from rabbit erythrocytes were subjected to thin-layer chromatography and toxin-binding glycolipids detected by using 125I-labeled Toxin A in a direct binding overlay technique . Two major and several minor toxin-binding glycolipids were detected in rabbit erythrocytes by this method . The results of structural analyses of the major toxin-binding glycolipids were consistent with a pentasaccharide-ceramide (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer) and a branched decasaccharide-ceramide (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3{Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-6}Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer) previously identified as the two most abundant glycolipids in rabbit erythrocytes . 125I-Toxin A binding to these glycolipids could be inhibited by bovine thyroglobulin, monospecific antiserum to the toxin, or by treatment of the glycolipids with alpha-galactosidase . The absence of toxin interaction with isoglobotriaosylceramide (Gal alpha 1-3Gal beta 1-4Glc-Cer) isolated from canine intestine suggested that the GlcNAc residue present in the terminal Gal alpha 1-3Gal beta 1-4GLcNAc sequence common to all known toxin binding glycoconjugates is required for carbohydrate-specific recognition by Toxin A . These observations are consistent with the proposed carbohydrate binding specificity of Toxin A for the nonreducing terminal sequence, Gal alpha 1-3Gal beta 1-4GlcNAc. Biochemistry, 1987 Aug 11, 26(16), 5042 - 8 Flavodoxin-cytochrome c interactions: circular dichroism and nuclear magnetic resonance studies; Tollin G et al.; Circular dichroism and 1H and 31P nuclear magnetic resonance spectroscopy have been used to investigate complex formation between cytochrome c and the flavodoxins from Azotobacter vinelandii and Clostridium pasteurianum . Such complexes are known to be involved in the mechanism of electron transfer between these two redox proteins . A large increase in ellipticity in the Soret band of the cytochrome heme was observed upon formation of the Clostridium flavodoxin complex, whereas much smaller changes were found for the complexes with either Azotobacter flavodoxin or an 8 alpha-imidazolyl-FMN-substituted Clostridium flavodoxin analogue . Similarly, the magnitudes of the perturbations of the contact-shifted heme proton resonances obtained upon complexation of cytochrome c by Azotobacter flavodoxin were much smaller than those previously shown for Clostridium flavodoxin {Hazzard, J . T., & Tollin, G . (1985) Biochem . Biophys . Res . Commun . 130, 1281-1286} . 31P nuclear magnetic resonance measurements were also consistent with differences in the interactions between the components in the complexes of the two flavodoxins with cytochrome c . It is suggested that these spectral changes are due to a loosening or opening of the heme crevice upon Clostridium flavodoxin binding, which allows closer contact between the heme and flavin prosthetic groups and results in a faster rate of electron transfer . The implications of these observations for biological oxidation-reduction processes are considered. Biochemistry, 1987 Aug 11, 26(16), 5036 - 42 Influence of 8 alpha-imidazole substitution of the FMN cofactor on the rate of electron transfer from the neutral semiquinones of two flavodoxins to cytochrome c; De Francesco R et al.; The effects of substituting an imidazole ring onto the 8 alpha-position of the FMN cofactor on the kinetics of electron transfer from the neutral semiquinone forms of Azotobacter and Clostridium flavodoxins to oxidized horse heart cytochrome c have been investigated by stopped-flow methods . Although 8 alpha-substitution does not alter the mechanistic pathway of the reaction, the rate constants are decreased by factors of 10-30, without significant changes in the equilibrium association constants of the intermediate electron-transfer complexes . Protonation of the imidazole ring further decreases the observed second-order rate constants for the electron-transfer reaction by factors of 20-50 . The pKa values for the 8 alpha-imidazole ring in both flavodoxin semiquinones were determined to be approximately 7 . In contrast, the reactions of the native flavodoxins with cytochrome c are pH independent . The results are consistent with a structural model of the intermediate complex {Simondsen, R . P., Weber, P . C., Salemme, F . R., & Tollin, G . (1982) Biochemistry 21, 6366-6375}, which postulates a close fit between the exposed dimethylbenzene ring of the FMN and the heme edge within a nonpolar interface region . The results further indicate that electron transfer is uncoupled from proton transfer, that it is the rate-limiting step, and that it occurs prior to proton transfer at all pH values . Finally, the results do not provide support for a direct role of the imidazole ring in the facilitation of one-electron transfer in those enzymes containing 8 alpha-N-histidylflavin coenzymes. J Biol Chem, 1987 Aug 5, 262(22), 10489 - 96 Clostridial pyruvate oxidoreductase and the pyruvate-oxidizing enzyme specific to nitrogen fixation in Klebsiella pneumoniae are similar enzymes; Wahl RC et al.; The chemical characterization, EPR properties, and mechanism of pyruvate:flavodoxin (ferredoxin) oxidoreductase from Klebsiella pneumoniae and Clostridium thermoaceticum have been investigated . A simple, specific, and sensitive assay and an efficient purification (based on the high affinity of these enzymes for a dye attached to agarose) are reported . The observed iron content of 8 atoms/subunit is twice that reported by others, whereas the contents of lipoate and flavin are less than 0.1 mol/subunit, in agreement with previous reports . Spectroscopic evidence suggests that the iron is present in Fe4S4(2+,1+) clusters . Reduction of the enzyme requires the presence of CoA as well as 1.1 pyruvate/subunit, which is very nearly the theoretical amount required the reduce two Fe4S(2+,1+) clusters . In the absence of CoA, stoichiometric amounts of pyruvate are decarboxylated, but the Fe/S centers are not reduced . We conclude that the K . pneumoniae and C . thermoaceticum enzymes are adapted to rapid reduction of low potential 1-e- carriers, similar to the pyruvate oxidoreductase of Halobacterium (Kerscher, L., and Oesterhelt, D . (1977) FEBS Lett . 83, 197-201), but different in that an Fe/S center-radical pair is used in the latter enzyme in place of the pair of Fe4S4 centers we find . The K . pneumoniae and C . thermoaceticum oxidoreductases appear to be mechanistically closely related to the Clostridium acidiurici enzyme (Uyeda, K., and Rabinowitz, J . C . (1971) J . Biol . Chem . 246, 3111-3119), differing as a class from the lipoate-containing, pyridine nucleotide-reducing enzyme present in aerobes (Reed, L . J . (1974) Accts . Chem . Res . 2, 740-746) . The function of the Klebsiella enzyme is to supply electrons to nitrogenase . This is accomplished in vitro with purified components via a nif-specific flavodoxin or other low potential 1-e- carriers such as viologen dyes or ferredoxins . The in vivo molar ratio of nitrogenase to the physiological reduction system, estimated from activity measurements of individual components in crude extracts, was 0.4:0.03:2:1 pyruvate oxidoreductase:flavodoxin:nitrogenase component II:nitrogenase component I. J Trop Med Hyg, 1987 Aug, 90(4), 189 - 92 Isolation of Clostridium difficile from diarrhoea patients in Bangladesh; Akhtar SQ; An attempt was made to detect Clostridium difficile and its toxin from the stools of 20 patients with antibiotic-associated diarrhoea (AAD), 35 with colitis, six with chronic diarrhoea and 300 with watery diarrhoea . Two toxigenic and three non-toxigenic strains were isolated from patients associated with antimicrobial therapy . All 300 stools from watery diarrhoea patients, not associated with antibiotics, were negative for Cl . difficile and its toxin . We conclude that Cl . difficile might be a cause of AAD in Bangladesh. Am J Clin Nutr, 1987 Aug, 46(2), 273 - 6 Microbial flora in the bypassed jejunum of patients with biliopancreatic bypass for obesity; Prakash G et al.; The microbial flora in the bypassed biliopancreatic intestinal segment was studied after obesity surgery . This procedure causes less diarrhea than jejunoileal bypass and appears to avoid extraintestinal complications . This report concerns type and quantity of bacteria colonizing the biliopancreatic segment and changes occurring after oral metronidazole treatment . Twelve specimens were aspirated in 10 patients via catheter inserted percutaneously during surgery . The specimens were plated immediately on selective and nonselective media under aerobic and anaerobic conditions . Essentially equal numbers of aerobes and anaerobes were recovered from the biliopancreatic segment with average counts of 10(4) cfu/mL and median counts of 10(5) cfu/mL . Four patients had counts of 10(7) cfu/mL . The most common aerobes were E . coli, Klebsiella, Gram-positive cocci, and Candida; among anaerobes, Clostridium and the Bacteroides fragilis group were most common . In three patients treated with metronidazole because of diarrhea, anaerobes were eliminated and diarrhea cleared. J Med Microbiol, 1987 Aug, 24(1), 41 - 52 The effects of Clostridium difficile crude toxins and toxin A on ileal and colonic loops in immune and non-immune rabbits; Ketley JM et al.; Rabbits were solidly immunised by parenteral injection of purified Clostridium difficile toxin A such that they resisted an intravenous challenge with a normally lethal dose of toxin A . Ileal and colonic loops constructed in non-immune and immune animals received challenge injections of crude culture filtrate or purified toxin A of C . difficile . Protection of ileum was manifest after sufficient initial mucosal damage resulted in release of high levels of antitoxin A into the loop lumen of immune animals . There was less fluid accumulation in ligated ileal loops of immune than of non-immune rabbits . Less protection was observed when loops were challenged with crude culture filtrate containing toxins A and B than when challenged with purified toxin A . In-vitro studies with Ussing chambers yielded no evidence for tissue-localised immunity as judged by electrical responses and histology of toxin-treated tissue from non-immune and immune animals . No differences were found in the degree of epithelial damage, or volume or composition of fluid accumulating in colonic loops of non-immune and immune rabbits challenged with toxin A or crude culture filtrate . However, in colonic loops of immune rabbits there was no overt tissue-localised haemorrhage, whereas in those of non-immune rabbits tissue-localised haemorrhage was marked . In contrast to our findings with ileal loops, fluid accumulating in colonic loops was watery and contained substantially less total protein and (in immune animals) antitoxin A. Gastroenterology, 1987 Aug, 93(2), 273 - 9 Differential effects of Clostridium difficile toxins A and B on rabbit ileum; Triadafilopoulos G et al.; The pathogenesis of Clostridium difficile enterocolitis appears to involve colonization of the bowel followed by release of toxin A, an enterotoxin, and toxin B, a cytotoxin . The purpose of this study was to determine the effect of purified toxins A and B on intestinal secretion, epithelial permeability, and morphology in perfused rabbit ileal loops . Intestinal permeability after toxin exposure was assessed by blood-to-lumen clearance of {3H}mannitol . Toxin A at doses of 5-100 micrograms/10 cm ileal loop caused a threefold to fivefold increase in {3H}mannitol permeability (p less than 0.001) vs . equal concentrations of toxin B or buffer control . In addition, perfusate from toxin A-exposed loops contained significantly more neutrophils (p less than 0.001) than toxin B or control loops . Toxin A caused severe epithelial cell necrosis with destruction of villi and polymorphonuclear infiltration . Electron microscopy of mucosa subjected to a low dose of toxin revealed widespread nonspecific dilatation of endoplasmic reticulum and mitochondrial swelling . In contrast to these effects of toxin A in ileal loops, in vitro experiments with ileal explants in short-term organ culture revealed that toxin A had no effect on epithelial cell permeability, protein synthesis, release of alkaline phosphatase, or morphology . Our results show that purified toxin A but not toxin B causes severe inflammatory enteritis in rabbit ileal loops, but has no discernable effect on rabbit ileum in vitro . We speculate that toxin A may contribute significantly to intestinal damage in C . difficile-associated colitis and diarrhea. Can J Microbiol, 1987 Aug, 33(8), 663 - 9 Oxidation of primary bile acids by a 7 alpha-hydroxysteroid dehydrogenase elaborating Clostridium bifermentans soil isolate; Sutherland JD et al.; A gram-positive, rod-shaped anaerobe (strain F-6) was isolated from soil . This organism was identified by cellular morphology as well as fermentative and biochemical data as Clostridium bifermentans . Strain F-6 formed 7-ketolithocholic acid from chenodeoxycholic acid and 7-ketodeoxycholic acid from cholic acid in whole cell cultures, but did not transform deoxycholic acid, ursodeoxycholic acid, or ursocholic acid . This reaction is reversible . The structures of 7-ketolithocholic acid and 7-ketodeoxycholic acid were verified by mass spectroscopy and by thin-layer chromatography using Komarowsky's spray reagent . When incubated with the strain F-6 glycine and taurine conjugates of the primary bile acids were partially hydrolyzed and transformed to 7-keto products . Optimal yields of 7-ketolithocholic acid and 7-ketodeoxycholic acid were obtained after 78 h of incubation . Culture pH changed with time and was characterized by an initial drop (1.1 pH units) and a gradual increase back to the starting pH (7.3) . Corroborating these observations, an inducible, NADP-dependent, 7 alpha-hydroxysteroid dehydrogenase was demonstrated in cell extracts of strain F-6 . A trace of NAD-dependent 7 alpha-hydroxysteroid dehydrogenase was also found . A substantial increase in the specific activity of the NADP-dependent 7 alpha-hydroxysteroid dehydrogenase was observed when either 7-ketolithocholic acid, chenodeoxycholic acid, or deoxycholic acid was included in the growth medium . Optimal induction of the NADP-dependent 7 alpha-hydroxysteroid dehydrogenase was achieved with 0.3-0.4 mM 7-ketolithocholic acid . Production of the enzyme(s) was optimal at 6-8 h of growth and the 7 alpha-hydroxysteroid dehydrogenases had a pH optimum of approximately 11.(ABSTRACT TRUNCATED AT 250 WORDS) Infect Immun, 1987 Aug, 55(8), 1779 - 83 Production, characterization, and protective effect of monoclonal antibodies to Clostridium chauvoei flagella; Tanaka M et al.; Monoclonal antibodies to flagella of Clostridium chauvoei were obtained by the fusion of murine myeloma cells (P3-X63-Ag8-U1) and spleen cells from BALB/c mice immunized with partially purified flagella of strain Okinawa . Enzyme-linked immunosorbent assay and Western blot analysis with partially purified flagella, flagellated cells, and nonflagellated mutants were used to show that five monoclonal antibodies are specific for the flagella . In the Western blot analysis, all five antiflagellar antibodies reacted strongly with the 56,000-molecular-weight protein, which corresponds to the flagellin . By using the ELISA-derived reactivity of monoclonal antibodies to the various clostridia and the competitive binding assay, we showed that the flagella of C . chauvoei had at least three epitopes . The three antiflagellar monoclonal antibodies (one immunoglobulin G and two immunoglobulin M) demonstrated passive protective effects in mice . These results strongly suggest that the flagella of C . chauvoei are important for protective immunity in mice. Epidemiol Infect, 1987 Aug, 99(1), 167 - 72 A food-poisoning incident caused by Clostridium botulinum toxin A in Japan; Otofuji T et al.; Food poisoning caused by Clostridium botulinum toxin A occurred in Japan . Eleven (31%) of 36 patients from 14 different areas died of botulism . Most of the patients had eaten commercial fried lotus-rhizome solid mustard without heating . The food, which implicated one of the special local products used for gifts in Kumamoto, was found to have been produced by a manufacturer in Kumamoto prefecture . In Fukuoka prefecture, two of three patients died on days 4 and 8 after eating the food; they had typical symptoms of botulism . A total of 42 packages of the food bought as gifts was collected from different districts in Fukuoka prefecture for examination for both organism and toxin . Thirteen of these (31%) were contaminated with the organism, and in 11 (26%) a small amount of toxin A had been produced. Appl Environ Microbiol, 1987 Aug, 53(8), 1902 - 6 Carbon monoxide-dependent chemolithotrophic growth of Clostridium thermoautotrophicum; Savage MD et al.; The acetogen Clostridium thermoautotrophicum was cultivated under CO-dependent chemolithotrophic conditions . CO-dependent growth profiles and energetics indicated that supplemental CO2 was fundamental to efficient growth at the expense of CO . Overall product stoichiometry approximated 6.5CO----CH3CO2H + 3.5CO2 + 0.6 cell C + 0.5 unrecovered C . Initial CO/CO2 ratios of 2 to 4 yielded optimal doubling times and cell yields . Maximal YCO values approximated 2.5 g of cell dry weight per mol of CO consumed; YH2 was considerably lower than YCO . Cross-transfer growth experiments and protein profiles indicated differential expression of genes between CO and methanol cultures. Appl Environ Microbiol, 1987 Aug, 53(8), 1880 - 4 Isolation of Clostridium botulinum type G from Swiss soil specimens by using sequential steps in an identification scheme; Sonnabend WF et al.; After Clostridium botulinum type G organisms and toxin were identified in necropsy specimens in cases of unexplained death in adults and infants (O . Sonnabend, W . Sonnabend, R . Heinzle, T . Sigrist, R Dirnhofer, and U . Krech, J . Infect . Dis . 143:22-27, 1981), extensive research to detect C . botulinum type G in soil samples from Switzerland was done . A total of 41 specimens from virgin soil and from cultivated land were examined for the presence of C . botulinum type G and other toxin types . Because of the lack of the lipase marker in type G, the detection of C . botulinum type G was based on the demonstration of type G organisms in enrichment cultures by a type G-specific enzyme-linked immunosorbent assay to detect both the type G toxin and antigen; enrichment cultures in which type G toxin or antigen was identified by enzyme-linked immunosorbent assay were then tested by a type G-specific gel immunodiffusion agar procedure . This method not only isolated strains of type G but also strains of Clostridium subterminale, a nontoxigenic variant of C . botulinum type G . As a consequence of the observed cross-reactions caused by strains of C . subterminale within this test system, all isolates of type G had to be definitively confirmed by mouse bioassay . The sequential steps of these methods seem to be very useful for detecting C . botulinum type G organisms . C . botulinum type G strains were isolated in five soil samples from different locations in close association with cultivated land.(ABSTRACT TRUNCATED AT 250 WORDS) Infect Immun, 1987 Aug, 55(8), 1873 - 7 Purification of Clostridium difficile toxin A by affinity chromatography on immobilized thyroglobulin; Krivan HC et al.; An efficient, single-step method for isolating highly purified toxin A from Clostridium difficile culture filtrates is described . The purification procedure was based on the affinity binding and release of toxin A to bovine thyroglobulin conjugated to agarose beads . The toxin strongly bound at 4 degrees C to the carbohydrate binding determinant Gal alpha 1-3Gal beta 1-4GlcNAc, a carbohydrate sequence which occurs on bovine thyroglobulin . Toxin bound to thyroglobulin at 4 degrees C, allowing its separation from the culture filtrate and contaminating proteins during the purification scheme . The toxin was eluted by increasing the temperature to 37 degrees C . The toxin-binding capacity was related to the amount of thyroglobulin immobilized on the gel: an affinity column containing 15 mg of bovine thyroglobulin per ml of gel bound 0.53 mg of toxin A per ml of gel . The percent recovery of purified toxin ranged from 56 to 80% and was inversely related to the amount of thyroglobulin coupled to the gel . The affinity-purified toxin was homogeneous as judged by crossed immunoelectrophoresis and gradient polyacrylamide gel electrophoresis and was immunologically identical to toxin A purified by conventional methods as determined by immunodiffusion analysis . The biochemical, hemagglutinating, and toxic properties of the toxin were preserved after affinity chromatography and were comparable with those of toxin A purified by conventional methods. J Steroid Biochem, 1987 Aug, 28(2), 189 - 92 Factors influencing production of 5(E)-19-nor-10-keto-vitamin D3 by rumen bacteria; Gardner RM et al.; Mixed populations of rumen bacteria or Clostridium hastiforme (a rumen isolate) catalyzed the oxidation of vitamin D3 to 5(E)-19-nor-10-keto-vitamin D3 . The reaction depended upon small amounts of O2 (less than 0.1% dissolved O2); when O2 was available, supernatant obtained from heat-killed mixed cultures also produced 5(E)-19-nor-10-keto-vitamin D3 . Results obtained by ultrafiltration indicated that at least two heat-stable factors of bacterial origin were involved . Lower rates of the same oxidation were observed when O2 was introduced to solutions containing vitamins D3 and L-cysteine . Oxygen radicals are known to be produced in such solutions and the involvement of such radicals in the D3 oxidation is probable since production in cysteine solutions was inhibited by superoxide dismutase and catalase. Zh Mikrobiol Epidemiol Immunobiol, 1987 Aug, (8), 62 - 4 {Use of the peroxidase method of immunoenzyme analysis for detecting Clostridium perfringens cells}; Kaplunova OP et al.; The possibility of using the peroxidase method of the enzyme immunoassay for the detection of C . perfringens cells in the culture fluid of strains, as well as in affected tissues in cases of anaerobic gas gangrene infection, has been shown . This method is highly specific, convenient and takes but 1.5 hrs. Poult Sci, 1987 Aug, 66(8), 1326 - 30 Influence of a wheat diet on mortality of broiler chickens associated with necrotic enteritis; Branton SL et al.; In an experiment to determine methods of incorporating soft winter wheat into broiler diets, a significant increase in mortality was observed in broilers fed wheat in crumbled diets . This increase in mortality was associated with necrotic enteritis with Clostridium perfringens indicated as the causative pathogen and complicated by a coccidiosis outbreak . When yellow corn was used in the diet, mortality was 2.9% . Use of all wheat, ground with a hammer mill, increased mortality to 28.9% . However, roller mill-ground wheat diet resulted in a mortality of 18.1% . When the grain component was approximately 50% wheat and 50% corn, mortality was 12.6% for broilers fed hammer mill-ground wheat and 3.4% for roller mill-ground wheat . Grain and feed were tested for several mycotoxins . Low levels of deoxynivalenol were found in both corn and wheat diets, but no differences between the corn and wheat-based diets were found that would explain the incidence of enteritis. J Lipid Res, 1987 Aug, 28(8), 949 - 54 Synthesis of a thiophosphate analog of dioctanoylphosphatidylcholine: a phospholipase C substrate; Snyder WR; Dioctanoylthiophosphatidylcholine, a racemic thiophosphate analog of L-alpha-dioctanoylphosphatidylcholine, has been synthesized and isolated by flash chromatography . In contrast with the didecanoylthiophosphatidylcholine synthesized previously, the analog is easily dispersed on sonication in aqueous media and is rapidly hydrolyzed to produce a free thiol group in the presence of the extracellular phospholipase C from either Bacillus cereus or Clostridium perfringens . When 5,5'-dithiobis (2-nitro-benzoic acid) was included as a thiol reactive chromogenic agent, the resultant measurement of product release, as an increase in absorbance at 412 nm, showed a linear relationship with added enzyme. J Clin Microbiol, 1987 Aug, 25(8), 1579 - 80 Bilateral abscessed orchiepididymitis associated with sepsis caused by Veillonella parvula and Clostridium perfringens: case report and review of the literature; Arrosagaray PM et al.; Veillonella species is a gram-negative coccus which is part of the anaerobic normal flora in the oral cavity, small intestine, upper respiratory tract, vagina, and urinary tract . The role that this organism plays in infection is not well known, and it is generally associated with other bacteria . We present a case of bilateral abscessed orchiepididymitis associated with septicemia due to Veillonella parvula and, later, to Clostridium perfringens, with the development of severe renal insufficiency and septic shock, which resolved favorably with antibiotic therapy, treatment of shock, and hyperbaric oxygen therapy . In reviewing the literature, we have not found any other case of sepsis due to Veillonella sp . associated with urological disorders. J Infect Dis, 1987 Aug, 156(2), 324 - 33 Effects of alpha and theta toxins from Clostridium perfringens on human polymorphonuclear leukocytes; Stevens DL et al.; Two toxins, alpha (phospholipase C) and theta (oxygen-labile hemolysin), were purified from Clostridium perfringens type A and assayed for toxic effects on human polymorphonuclear leukocytes (PMNLs) . Crude preparations containing both toxins totally inhibited chemotaxis and chemiluminescence responses of PMNLs and reduced PMNL viability . Purified alpha toxin did not alter PMNL viability, chemotactic responsiveness, or morphology but did enhance opsonized zymosan-induced PMNL chemiluminescence over a wide range of toxin concentrations . theta Toxin, at 12.5 hemolytic units (HU) per 10(5) PMNLs, reduced cell viability and induced marked PMNL morphological changes . Concentrations of theta toxin between 4 and 32 HU per 10(5) PMNLs inhibited PMNL chemiluminescence in a dose-dependent manner, whereas a lower concentration enhanced the PMNL chemiluminescent response to opsonized zymosan . Effects on chemotaxis were also dose dependent . Increased PMNL random migration was observed at a concentration of theta toxin of 0.06 HU per 2.5 X 10(5) PMNLs (P less than .05), whereas concentrations of greater than 0.08 HU per 2.5 X 10(5) PMNLs reduced both directed and random migration (P less than .05). J Biol Chem, 1987 Jul 25, 262(21), 10355 - 61 Transphosphatidylation activity in Clostridium butyricum . Evidence for a secondary pathway by which membrane phospholipids may be synthesized and modified; Walton PA et al.; Membrane particles from Clostridium butyricum, incubated with Triton X-100 and 32P-labeled phosphatidylethanolamine, phosphatidylglycerol, or phosphatidylserine, resulted in the labeling of three phospholipids . These unknown phospholipids incorporated label from the phosphate and acyl chains of the substrate, but not from the head group . Two-dimensional TLC of the intact lipids and their deacylation products showed that these lipids were phosphatidic acid, cardiolipin, and the previously unreported phosphatidyltriton . The reaction involved the transfer of the phosphatidyl moiety of the substrate molecule in a phospholipase D-like manner . The reaction displayed sigmoidal kinetics, did not require divalent cations, possessed an acidic pH optimum, and was sensitive to thermal inactivation . Differences in thermal sensitivity and pH optimum indicated that a distinct enzyme activity may be involved in the formation of cardiolipin . A primary alcohol group was required on the acceptor molecule, which could be either amphipathic or water-soluble . Addition of exogenous unlabeled phosphatidylglycerol resulted in the increased formation of cardiolipin, with a concomitant decrease in the level of phosphatidyltriton formed . Labeled phosphatidylethanolamine, phosphatidylglycerol, or phosphatidylserine could be formed upon addition of their corresponding alcoholic head group to incubations containing a 32P-labeled phosphatidyl donor and Triton X-100 . These results indicate that, in C . butyricum, enzymic steps exist that would allow remodeling of the membrane phospholipids, without requiring de novo biosynthesis. J Biol Chem, 1987 Jul 25, 262(21), 9945 - 7 An investigation of hydrogenase I and hydrogenase II from Clostridium pasteurianum by resonance Raman spectroscopy . Evidence for a {2Fe-2S} cluster in hydrogenase I; Macor KA et al.; Resonance Raman spectra are reported for hydrogenase I and II from Clostridium pasteurianum . These spectra show overlapping bands with contributions from {4Fe-4S} clusters, known to be present in these enzymes, and from novel FeS centers of hitherto undefined structure . For hydrogenase I there are strong bands at 288 and 394 cm-1, which are seen in {2Fe-2S} proteins and in no other FeS species so far examined . In contrast these bands do not appear for hydrogenase II, whose resonance Raman spectrum is dominated by {4Fe-4S} cluster modes . These results provide the first structural information on the hydrogenase I FeS center involved in H2 activation and demonstrate structural differences between hydrogenase I and hydrogenase II. J Biol Chem, 1987 Jul 25, 262(21), 10086 - 92 A new endo-beta-galactosidase releasing Gal alpha 1----3Gal from carbohydrate moieties of glycoproteins and from a glycolipid; Fushuku N et al.; Culture fluid of Clostridium perfringens contained an endoglycosidase releasing a galactosyl disaccharide from glycans derived from glycoproteins of teratocarcinoma OTT6050 . The endoglycosidase was purified by ammonium sulfate precipitation, Sephadex G-200 column chromatography, and DEAE-Sephadex A-25 column chromatography . The structure of the disaccharide product was determined to be Gal alpha 1----3Gal . The enzyme also released the disaccharide from bovine thyroglobulin glycopeptide and from a pentaglycosyl ceramide (Gal alpha 1----3Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4Glc-Cer) . Therefore, we concluded that the enzyme was an endo-beta-galactosidase cleaving Gal beta 1----4GlcNAc linkage . Keratan sulfate and the antigenic determinant of blood group type B were resistant to the enzyme: the specificity of the enzyme is different from other endo-beta-galactosidases so far described . The enzyme could act on intact thyroglobulin and defucosylated ovarian cyst mucin. J Biol Chem, 1987 Jul 5, 262(19), 8994 - 9003 Enzymatic reactions in the degradation of 5-aminovalerate by Clostridium aminovalericum; Barker HA et al.; The anaerobic degradation of 5-aminovalerate to valerate, acetate, propionate, and ammonia by Clostridium aminovalericum was shown to involve the following intermediates: glutaric semialdehyde, 5-hydroxyvalerate, 5-hydroxyvaleryl-CoA, 4-pentenoyl-CoA, 2,4-pentadienoyl-CoA, trans-2-pentenoyl-CoA, L-3-hydroxyvaleryl-CoA, 3-ketovaleryl-CoA, acetyl- and propionyl-CoA and the corresponding acylphosphates, valeryl-CoA, and possibly 3-pentenoyl-CoA . With exception of the enzyme presumably reducing 2,4-pentadienoyl-CoA to 3-pentenoyl-CoA, enzymes catalyzing the formation and utilization of the above intermediates were demonstrated in extracts . Trans-2-pentenoyl-CoA was shown to be the immediate precursor of valeryl-CoA . The reduction of 2-pentenoyl-CoA was found to be coupled to the oxidation of 4-pentenoyl-CoA to 2,4-pentadienoyl-CoA . Several enzymes catalyzing the above reactions were partially purified and some of their properties determined . A high pressure liquid chromatography method of identifying and estimating most of the above mentioned CoA thiolesters was developed. J Biol Chem, 1987 Jul 5, 262(19), 9130 - 5 Purification and characterization of pyruvate:NADP+ oxidoreductase in Euglena gracilis; Inui H et al.; Pyruvate:NADP+ oxidoreductase was homogeneously purified from crude extract of Euglena gracilis . The Mr of the enzyme was estimated to be 309,000 by gel filtration . The enzyme migrated as a single protein band with Mr of 166,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, suggesting that the enzyme consists of two identical polypeptides . The absorption spectrum of the native enzyme exhibited maxima at 278, 380, and 430 nm, and a broad shoulder was observed around 480 nm; the maximum at 430 nm was eliminated by reduction of the enzyme with dithionite . Reduction of the enzyme with pyruvate and CoA and reoxidation with NADP+ were proved from changes of absorption spectra . The enzyme contained 2 molecules of FAD and 8 molecules of iron . It was also indicated that the enzyme was thiamine pyrophosphate-dependent . The enzyme was oxygen-sensitive, and the reaction was affected by the presence of oxygen . Pyruvate was the most active substrate, but the enzyme was slightly active for 2-oxobutyrate, 3-hydroxypyruvate, and oxalacetate, but not for glyoxylate and 2-oxoglutarate . The native electron acceptor was NADP+, whereas NAD+ was completely inactive . Methyl viologen, benzyl viologen, FAD, and FMN were utilized as artificial electron acceptors, whereas spinach and Clostridium ferredoxins were inactive . Pyruvate synthesis by reductive carboxylation of acetyl-CoA with NADPH as the electron donor occurred by the reverse reaction of the enzyme . The enzyme also catalyzed a pyruvate-CO2 exchange reaction and electron-transfer reaction from NADPH to other electron acceptors like methyl viologen . These results indicate that pyruvate:NADP+ oxidoreductase in E . gracilis is clearly distinct from either the pyruvate dehydrogenase multienzyme complex or pyruvate:ferredoxin oxidoreductase. Zentralbl Bakteriol Mikrobiol Hyg {A}, 1987 Jul, 265(3-4), 340 - 52 Identification of anaerobic bacteria using high-performance liquid chromatography; Krausse R et al.; 20 species of the genera Clostridium, Bacteroides, Fusobacterium and the type strains of Peptostreptococcus anaerobius, Peptococcus asaccharolyticus, Veillonella parvula and Propionibacterium acnes were examined for the production of volatile (VFA) and nonvolatile (NVFA) short-chain fatty acids using high-performance liquid chromatography (HPLC) with a column for organic acids (Aminex HPX-87H) . 10 min are needed for sample preparation and the VFA and NVFA were detected simultaneously in a single chromatographic run . The total time required to run each chromatogram was approximately 60 min . With regard to the production of short-chain fatty acids in culture media it was possible to identify the species of the genera rapidly and clearly . The results illustrate the role of HPLC in determinating short-chain fatty acid products as an additional means for rapid differentiation between closely related anaerobic bacterial species . The aim of the method developed is to establish a complete automatization for the identification of anaerobic bacteria using an automatic sampling system and a microprocessor-controlled chromatography unit. J Wildl Dis, 1987 Jul, 23(3), 376 - 85 Epizootic necrotic enteritis in wild geese; Wobeser G et al.; Outbreaks of a disease characterized by severe necrotic enteritis occurred among Canada geese (Branta canadensis), lesser snow geese (Anser caerulescens), Ross' geese (A . rossi), and white-fronted geese (A . albifrons) on lakes in Saskatchewan and Manitoba during the autumn of 1983, 1984 and 1985 . Ducks using the lakes were apparently not affected . Lesions in the geese closely resembled those described in enteritides in other species associated with the proliferation of Clostridium perfringens in the small intestine . Clostridium perfringens was present in large numbers in the affected areas of the intestine of the geese; other pathogens were not identified . It is hypothesized that an abrupt change in diet as geese begin to feed on grain disrupts the intestinal microflora, allowing C . perfringens to proliferate in the upper small intestine . Toxins produced by the bacteria then cause mucosal necrosis . Protease-inhibitory substances in some grains might also have a role in the disease. J Clin Microbiol, 1987 Jul, 25(7), 1244 - 7 Commercial latex agglutination test for detection of Clostridium difficile-associated diarrhea; Kelly MT et al.; A commercially available latex agglutination test for Clostridium difficile was compared with a cell culture cytotoxin assay and bacteriological culture for the laboratory diagnosis of C . difficile-associated diarrhea and colitis (CAD) . Stool specimens from 626 patients were tested by the three methods, and specimens from 118 patients (19%) were positive by at least one of the methods . The results of the three tests agreed in 88% of the specimens tested, overall, but they agreed in only 34% of the 118 positive specimens . Ninety-three patients were evaluated to assess the significance of positive and negative results for each assay . Of 40 patients found to have CAD, 70% were positive by the cytotoxin assay, 78% were positive by the latex agglutination test, and 90% were culture positive . Of 53 patients who did not have CAD, 2% were positive by the cytotoxin assay, 8% were positive by the latex test, and 4% were culture positive . The detection of CAD was improved by using the tests in combination, and 97% of specimens positive by two or three methods were from patients who had CAD . Testing of multiple specimens from individual patients also increased the sensitivity of detection of CAD . The results suggest that the latex agglutination test may be useful for rapid diagnosis of CAD, especially in laboratories that lack cell culture facilities . However, the accuracy of CAD detection is improved when the latex test is used in combination with culture or the cytotoxin assay. J Am Vet Med Assoc, 1987 Jul 1, 191(1), 73 - 4 Botulism as a sequel to open castration in a horse; Bernard W et al.; Clostridium botulinum and type-B C botulinum toxin were isolated from a necrotic wound that developed subsequent to castration in a 2-year-old Thoroughbred gelding . The horse had clinical signs of botulism and was successfully treated with wound debridement, C botulinum type-B antitoxin, potassium penicillin, and supportive care. J Cell Physiol, 1987 Jul, 132(1), 168 - 72 Calcium and calmodulin in cellular intoxication with Clostridium difficile toxin B; Caspar M et al.; In cultured human lung fibroblasts treated with Clostridium difficile toxin B, the development of the cytopathogenic effect was inhibited by the proton ionophore monensin but was not affected by some other ionophores . The calcium channel blockers verapamil and LaCl3 protected the cells against intoxication, as did the calmodulin antagonists trifluoperazine, amitriptyline, R 24571, and dansylcadaverine . Since these agents could not prevent intoxication when added after the toxin internalization was completed, we suggest that calmodulin and uptake of extracellular calcium are needed for the internalization but not for the cytosolic action of the toxin. Infect Immun, 1987 Jul, 55(7), 1686 - 91 Role of volatile fatty acids in colonization resistance to Clostridium difficile in gnotobiotic mice; Su WJ et al.; Clostridium difficile is an agent involved in the development of antibiotic-associated pseudomembranous colitis . The purpose of this work was to investigate the role of volatile fatty acids (VFAs) in resistance to colonization by C . difficile by using a gnotobiotic animal model . Accordingly, germfree mice were associated with different hamster flora, and the VFAs in their cecal contents were measured by gas chromatography . The results showed that VFAs were produced mainly by the intestinal flora, especially by the strictly anaerobic bacteria . In these associated mice, the concentrations of acetic, propionic, and butyric acids were higher than those of other acids, but at pH 6.8 the MICs of these three acids in vitro for C . difficile were more than 200 mu eq/ml . In gnotobiotic mice monoassociated with C . difficile and in the isolated ceca of these mice, VFAs did not inhibit the growth of C . difficile . In gnotobiotic mice which were diassociated with C . difficile and C . butyricum and given drinking water with a lactose concentration of 20%, the cecal contents included about the same amount of butyric acid as did those of the monoassociated mice, although the population of C . difficile remained the same . Therefore, it is suggested that VFAs alone cannot inhibit intestinal colonization by C . difficile and that, consequently, other inhibitory mechanisms are also present. Infect Immun, 1987 Jul, 55(7), 1610 - 5 Biochemical studies on the effect of Clostridium difficile toxin B on actin in vivo and in vitro; Mitchell MJ et al.; We describe a simplified procedure for purification of Clostridium difficile toxin B . In this procedure, cytotoxicity is associated with a single protein band with a molecular mass of 230 kilodaltons . We used direct fluorescent staining of actin filaments to study the effect of this toxin on cultured cells . Morphologic changes were preceded by a decrease in the number and length of stress fibers followed by their disappearance with condensation of cellular actin around the nucleus . We then showed that cells treated with either cytochalasin B or toxin B had a significant increase in the monomeric actin pool as quantitated by DNase I inhibition . In contrast to the cytochalasins, toxin B had no direct effect on the rate or extent of actin polymerization or network formation in vitro . Cytoplasmic extracts of toxin B-treated cells had a significantly lower level of modulating activity on actin assembly and interactions in vitro compared with extracts of untreated cells . These results suggest that the action of toxin B on cells is due to direct or indirect effects on cellular proteins involved in controlling the state of actin assembly in the cells. Infect Immun, 1987 Jul, 55(7), 1541 - 6 Factors influencing the phagocytosis of Clostridium difficile by human polymorphonuclear leukocytes; Dailey DC et al.; Phagocytosis of Clostridium difficile by human polymorphonuclear leukocytes (PMNs) and the possible role of the clostridial toxins in this process were investigated . Phagocytosis of C . difficile was independent of aerobiosis and clearly depended on opsonization . Either complement or antibodies to C . difficile could serve as opsonins . Toxigenic strains of C . difficile were more resistant to phagocytosis than were nontoxigenic strains . Pretreatment of PMNs with as much as 10,000 units of toxins from culture filtrates of C . difficile for 2 h had no effect on either the phagocytic activity of PMNs or their viability as determined by trypan blue exclusion . In contrast, treatment of human embryonic intestinal cells with the same amount of toxin under identical conditions resulted in cell death. Antimicrob Agents Chemother, 1987 Jul, 31(7), 1111 - 6 In vitro and in vivo antibacterial activities of T-2588, a new oral cephalosporin, compared with those of other oral beta-lactam antibiotics; Okamoto S et al.; T-2588, the pivaloyloxymethyl ester of T-2525, {6R, 7R}-7-{(z)-2-(2-aminothiazol-4-yl)-2-methoxyiminoacetoamido} -3- {(5-methyl-2H-tetrazol-2-yl)methyl}-3-cephem-4-carboxylic acid, is a new oral cephalosporin . T-2525 had a widely expanded antibacterial spectrum against gram-negative and gram-positive bacteria . T-2525 was more active in vitro than cefaclor, cephalexin, and amoxicillin against members of the family Enterobacteriaceae and Branhamella catarrhalis . Moreover, it exhibited superior in vitro activity against Streptococcus pyogenes, Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria gonorrhoeae . T-2525 was highly stable to various beta-lactamases, which were classified as Richmond and Sykes types Ia, Ib, Ic, III, IV, and Vc . It had high affinities for the lethal (essential) penicillin-binding proteins of Escherichia coli, Clostridium perfringens, and Bacteroides fragilis . T-2588 had excellent therapeutic effect on systemic infections in mice with various species of gram-negative bacteria, including beta-lactamase-producing bacteria. J Steroid Biochem, 1987 Jul, 28(1), 49 - 54 Cofactor requirements of steroid-17-20-desmolase and 20 alpha-hydroxysteroid dehydrogenase activities in cell extracts of Clostridium scindens; Krafft AE et al.; Two neutral steroid-transforming activities were demonstrated in cell extracts of Clostridium scindens . Steroid-17-20-desmolase and 20 alpha-hydroxysteroid dehydrogenase were found to be inducible in cells cultured in the presence of cortisol . Both activities required manganese ions and NAD+ or NADH for activity . Cortisol, cortisone and 11-desoxycortisol were substrates as well as inducers of steroid-17-20-desmolase and 20 alpha-hydroxysteroid dehydrogenase activities . 17 alpha-Hydroxyprogesterone was an effective inducer but did not serve as a substrate for either enzyme activity . C . scindens is the first bacterial species of the normal human intestinal flora reported to elaborate inducible steroid-17-20-desmolase and 20 alpha-hydroxysteroid dehydrogenase activities . The results of cofactor, substrate specificity and induction studies suggest that these two activities may reside in the same enzyme complex. Clin Pharm, 1987 Jul, 6(7), 570 - 4 Management of tetanus; Olsen KM et al.; Two cases of tetanus are presented, and the diagnosis, clinical features, and management of tetanus are reviewed . The first patient, an 86-year-old woman, had marked muscle rigidity but was able to breathe spontaneously . A dark eschar with purulent drainage was noted on her left foot, but Clostridium tetani was not isolated . She was placed in a semidark room and was treated with penicillin G; tetanus immune globulin (TIG) 5000 units i.m.; tetanus toxoid 0.5 mL i.m.; diazepam, chlorpromazine, and morphine for sedation, muscle relaxation, and analgesia; ranitidine for stress ulcer prophylaxis; heparin for prevention of deep-vein thrombosis; and peripheral-vein nutrition . Her condition improved gradually, and she was discharged to a rehabilitation institute after 32 days . The second patient, a 46-year-old woman, experienced progressive descending paralysis and required ventilatory support . She had a periodontal abscess, but cultures of the drainage were negative . She was placed in a semidark room and treated with erythromycin, TIG, tetanus toxoid, diazepam, pancuronium bromide, morphine, ranitidine, and heparin . Autonomic instability occurred during the second and third weeks, but cardiac output was maintained without treatment . The patient was extubated after five weeks, and was transferred out of the intensive-care in the following week . The diagnosis of tetanus is based primarily on characteristic findings of muscle rigidity and reflex spasms; cultures for C . tetani are of limited value . A history of trauma or injury is common . Pulmonary infections and cardiovascular instability are the most common complications . Therapy consists of ventilatory support; control of neuromuscular symptoms with benzodiazepines, narcotics, and neuromuscular blockers; antibiotic therapy.(ABSTRACT TRUNCATED AT 250 WORDS) Res Vet Sci, 1987 Jul, 43(1), 134 - 5 Oral dosing of mice with Clostridium botulinum type C cultures and culture filtrates; Smith GR et al.; In two of 14 tests in which adult mice were dosed per os, whole broth culture of Clostridium botulinum type C was more lethal than culture sterilised by membrane filtration . The results indicated that occasionally--though not usually--significant bacterial multiplication and toxigenesis occurred in the gut. J Clin Microbiol, 1987 Jul, 25(7), 1336 - 7 Kinetics study of immunological response to Clostridium botulinum toxin; Dezfulian M et al.; A double-antibody enzyme-linked immunosorbent assay (ELISA) for detection of humoral antibody to type A botulinal toxin was developed . This assay was used to study the kinetics of antibody response of a volunteer to botulinal toxoid . The circulating type A antitoxin was first detected by the ELISA 2 weeks after the first booster injection of the toxoid . The antibody titer stayed level until the second booster at 12 weeks . The titer then continued to rise throughout the remaining study period . The neutralizing antibody to type A toxin was detected by mouse assay 15 weeks after detection of antitoxin by the ELISA. J Clin Microbiol, 1987 Jul, 25(7), 1225 - 7 Isolation of Clostridium difficile and detection of cytotoxin in the feces of diarrheic foals in the absence of antimicrobial treatment; Jones RL et al.; Clostridium difficile was isolated from the feces of 27 of 43 diarrheic foals (63%), and cytotoxin was detected in feces from 28 diarrheic foals (65%) . The foals had not received any antimicrobial treatment before the onset of diarrhea . C . difficile was not isolated from feces of 18 normal foals without diarrhea and 62 adult horses (P less than 0.005) . This finding of C . difficile and its toxins in association with diarrhea in foals adds another possible cause to the list of infectious agents which may cause diarrhea in foals. J Immunol, 1987 Jul 1, 139(1), 262 - 70 Cell surface sialic acid influences tumor cell recognition in the mixed lymphocyte reaction; Powell LD et al.; The Ia+ B cell lymphoma, AKTB-1b, fails to stimulate thymic lymphocytes in a one-way mixed lymphocyte reaction unless pretreated with sialidase or inhibitors of N-linked oligosaccharide processing . A comparison of different sialidases and sialyltransferases suggests that the removal of only a subset of total surface sialic acid, rather than net desialylation of the cell surface, is required . Three sialidases were compared, including Vibrio cholerae (VC) and Clostridium perfringens (CP), which will cleave alpha 2-3, alpha 2-6, and alpha 2-8, sialic acid linkages, and Newcastle Disease virus (NDV), which will remove only alpha 2-3 and alpha 2-8 linked sialic acid . When treated with equivalent units of sialidase, CP-, VC-, and NDV-treated cells were 24-fold, sixfold, and threefold better stimulators than untreated cells . In contrast, VC released 1.3-fold and 2.5-fold more sialic acid per cell than did CP or NDV, respectively . Furthermore, VC was superior in reducing the levels of binding of the sialic acid-specific lectin, Limulus polyphemus agglutinin, in exposing Gal beta 1-3GalNAc and Gal beta 1-4GlcNAc residues, and in desialylating gangliosides . Two-dimensional gel analysis indicated that VC and CP were both equal and superior to NDV in the desialylation of iodinatable cell-surface proteins, including H-2Kk, I-A beta k, and a highly sialylated 65,000 dalton protein of unknown identity . Maximal resialylation of CP-treated cells with exogenously added CMP-NANA and either the alpha 2-3(Gal beta 1-3GalNAc) or alpha 2-6(Gal beta 1-4GlcNAc) sialyltransferase did not reduce the stimulatory capacity of these cells . However, resialylation of VC-treated cells with just CMP-NANA alone resulted in 49% reversal of their stimulatory capacity, and no additional reversal could be achieved with either of the sialyltransferases . Although the alpha 2-6(Gal beta 1-4GlcNAc) sialyltransferase was capable of adding back approximately 10% of the sialic acid removed, the endogenous activity added back approximately 0.1% of the total sialic acid removed . SDS-PAGE gels of the sialylated cells indicated that the exogenously added sialyltransferase labeled many different proteins, whereas the endogenous activity labeled far fewer proteins, predominantly in 46,000 and 25,000 m.w . range . Both the desialylation and resialylation data suggest that the sialidase-dependent stimulation is due to the desialylation of specific membrane structures . Together with previous studies, these data suggest that the sialic acids involved are probably alpha 2-6 linked to N-linked glycosyl moieties. Prikl Biokhim Mikrobiol, 1987 Jul-Aug, 23(4), 547 - 51 {Microbial inhibitors of phospholipase C}; Rozhanskaia TI et al.; Inhibitors of the Clostridium perfringens phospholipase C were prepared from the filtrates of the culture liquids of Streptomyces saracetidus and Streptomyces species using soluble and cross-linked polyelectrolytes . The technological scheme of isolation involves ultrafiltration . The inhibitors produced by the two strains had different chemical nature . The preparation obtained from Str . saraceticus was proved to be a complex of inhibitors that were separated by gel-chromatography into a major polypeptide with a molecular weight of 5500-6500 and a low-molecular weight glycopeptide . The inhibitor obtained from Str . species was found to be a high-molecular weight protein. Arch Dis Child, 1987 Jul, 62(7), 743 - 4 T activation haemolysis and death after blood transfusion; Placzek MM et al.; A 30 week gestation infant developed necrotising enterocolitis associated with Clostridium perfringens septicaemia at 3 weeks of age . He responded to treatment with intravenous fluids, antibiotics, and blood, but his blood haemolysed . Because of anaemia further blood was given, but, within minutes he died . Examination of his red cells showed an increase in T activation. J Wildl Dis, 1987 Jul, 23(3), 488 - 91 Clostridium perfringens as the cause of death of a captive Atlantic bottlenosed dolphin (Tursiops truncatus); Buck JD et al.; A previously healthy captive female bottlenosed dolphin (Tursiops truncatus) died suddenly . At necropsy, Clostridium perfringens was isolated from dorsal muscle, blood, left heart ventricle, thoracic fluid, and abdominal fluid . An identical strain was recovered from pool water . A male dolphin in the same pool had inflicted several "rake" marks on the dorsal surface of the female . Water-borne bacteria probably entered these lesions which served as the focus for anaerobe penetration and spread. J Clin Microbiol, 1987 Jul, 25(7), 1333 - 5 Plasmid analysis as a means of strain differentiation in Clostridium perfringens; Mahony DE et al.; A total of 114 Clostridium perfringens isolates were serotyped and examined for plasmids . Fifty-two strains were from hospitalized patients with diarrhea or from hospital environments, and 62 epidemiologically unrelated isolates were obtained from food poisoning outbreaks . All strains were screened for bacteriocin production against a common indicator strain of C . perfringens . In the one significant hospital outbreak of C . perfringens diarrhea, three to five plasmid types were found in strains of the predominant serotype, but no similar correlation between serotype and plasmid type was found in random isolates from a variety of sources . All of the strains associated with the diarrhea outbreak produced bacteriocins, whereas 63% of the strains from various sources produced bacteriocins . The typing data suggest a promising differentiating capability for plasmid analysis in the epidemiological study of outbreaks of food poisoning, diarrhea, or infections caused by C . perfringens. J Biochem (Tokyo), 1987 Jul, 102(1), 163 - 70 Purification and properties of collagenase from a Streptomyces species; Endo A et al.; A collagenase active against native, insoluble collagen was isolated from the culture filtrate of a streptomycete which has been designated Streptomyces sp . C-51 . Collagenase was produced by growing this strain in media containing gelatin . Purification by ammonium sulfate fractionation and chromatographies on DEAE-Toyopearl and DEAE-Cellulofine columns produced active enzyme which was free of contaminating proteins, including nonspecific proteinases, but which contained two active subspecies (I and II) . Both subspecies were purified by preparative slab gel electrophoresis . The apparent molecular weights were 100,000 for the homogeneous fraction I and 90,000-110,000 for the microheterogeneous fraction II . These two subspecies were most active at pH 8-9, very similar in amino acid composition and immunologically identical . Some other properties of the Streptomyces C-51 collagenase were compared with those of Clostridium histolyticum collagenase . Substrate specificity, insensitivity to N-ethylmaleimide and diisopropyl fluorophosphate, and sensitivity to certain metal ion complexing agents were similar for the collagenases from both microorganisms. Arch Microbiol, 1987 Jul, 148(2), 107 - 14 Screening for plasmids in the genus Clostridium; Lee CK et al.; A plasmid screening was performed on 150 strains out of 75 clostridial species using a modification of the alkaline-lysis procedure . In 26 strains representing 21 species one or more plasmid bands were detected ranging in size from 3 to more than 100 kilobase pairs . Clostridium aceticum proved to contain a single small plasmid (pCA1) of 5.4 kbp as revealed by restriction analysis and electron microscopy . A physical map of pCA1 has been constructed . Spontaneous mutants of C . aceticum defective in autotrophic growth have been isolated . No direct correlation between plasmid content and autotrophy could be found. Antimicrob Agents Chemother, 1987 Jul, 31(7), 1135 - 6 Treatment of Clostridium difficile colitis in hamsters with a lipopeptide antibiotic, LY146032; Dong MY et al.; LY146032, an acidic lipopeptide antibiotic which inhibits the biosynthesis of cell wall peptidoglycan, was found to be effective in delaying death in a hamster model of pseudomembranous colitis . A dose of 0.05 mg/day was effective . The equivalent protection with vancomycin required a dose 100-fold higher, i.e., 5 mg/day. Antimicrob Agents Chemother, 1987 Jul, 31(7), 1039 - 45 Genetic characterization of a Clostridium difficile erythromycin-clindamycin resistance determinant that is transferable to Staphylococcus aureus; Hachler H et al.; The transferable macrolides-lincosamides-streptogramin B (MLS) resistance determinant of clinical isolates of Clostridium difficile, designated ermZ, has been shown to share homology with ermB, which is associated with Staphylococcus aureus transposon Tn551 . Homology within Tn551 was confined to less than or equal to 1.3 kilobases, whereas no homology could be demonstrated between Tn551 sequences external to ermB and MLS-resistant C . difficile . Transfer of ermZ from C . difficile to S . aureus was achieved by means of the filter mating technique, suggesting that (conjugative?) intergeneric exchange between clostridia and staphylococci may also occur in nature . S . aureus transcipients were shown to contain additional DNA from C . difficile besides ermZ . This additional DNA appeared to be present in MLS-susceptible C . difficile and might form part of an as yet undemonstrated insertion sequence element associated with ermZ of resistant strains. Can J Microbiol, 1987 Jul, 33(7), 589 - 92 Inhibition of macromolecular synthesis by caffeine in Clostridium perfringens; Labbe RG et al.; Caffeine (2 mg/mL) inhibited the incorporation of {14C}adenine into actively growing cells of Clostridium perfringens NCTC 8679 in a dose-dependent manner . Also reduced by caffeine was incorporation of {14C}thymidine and 14C-labeled amino acids . No effect on guanine, uracil, adenosine, guanosine, or uridine was detected . Actual incorporation of {14C}caffeine or {14C}thymine in control cultures did not occur. Biochemistry, 1987 Jun 30, 26(13), 3943 - 8 Substrate activity of synthetic formyl phosphate in the reaction catalyzed by formyltetrahydrofolate synthetase; Smithers GW et al.; Formyl phosphate, a putative enzyme-bound intermediate in the reaction catalyzed by formyltetrahydrofolate synthetase (EC 6.3.4.3), was synthesized from formyl fluoride and inorganic phosphate {Jaenicke, L . v., & Koch, J . (1963) Justus Liebigs Ann . Chem . 663, 50-58}, and the product was characterized by 31P, 1H, and 13C nuclear magnetic resonance (NMR) . Measurement of hydrolysis rates by 31P NMR indicates that formyl phosphate is particularly labile, with a half-life of 48 min in a buffered neutral solution at 20 degrees C . At pH 7, hydrolysis occurs with P-O bond cleavage, as demonstrated by 18O incorporation from H2(18)O into Pi, while at pH 1 and pH 13 hydrolysis occurs with C-O bond cleavage . The substrate activity of formyl phosphate was tested in the reaction catalyzed by formyltetrahydrofolate synthetase isolated from Clostridium cylindrosporum . Formyl phosphate supports the reaction in both the forward and reverse directions . Thus, N10-formyltetrahydrofolate is produced from tetrahydrofolate and formyl phosphate in a reaction mixture that contains enzyme, Mg(II), and ADP, and ATP is produced from formyl phosphate and ADP with enzyme, Mg(II), and tetrahydrofolate present . The requirements for ADP and for tetrahydrofolate as cofactors in these reactions are consistent with previous steady-state kinetic and isotope exchange studies, which demonstrated that all substrate subsites must be occupied prior to catalysis . The k cat values for both the forward and reverse directions, with formyl phosphate as the substrate, are much lower than those for the normal forward and reverse reactions . Kinetic analysis of the formyl phosphate supported reactions indicates that the low steady-state rates observed for the synthetic intermediate are most likely due to the sequential nature of the normal reaction. J Am Vet Med Assoc, 1987 Jun 15, 190(12), 1550 - 5 Isolation of Clostridium perfringens from neonatal calves with ruminal and abomasal tympany, abomasitis, and abomasal ulceration; Roeder BL et al.; Eight neonatal calves (2 to 21 days old) with suspected abomasal displacement or intestinal obstruction after acute onset of abdominal tympany, colic, depression, or death were referred to Kansas State University for clinical examination or for necropsy . Results of routine hematologic and serum chemical analyses did not reveal consistent changes . Necropsy revealed abomasal distention, with various degrees of abomasitis, hemorrhage, and ulceration, but did not reveal evidence of displaced abomasum or obstructed intestine . Specimens of ruminal contents collected via stomach tube or at necropsy and abomasal contents collected at necropsy were obtained for anaerobic bacteriologic culture . Clostridium perfringens was isolated from all specimens, and on the basis of toxin neutralization tests in mice, 7 were type A and one was type E . Copper concentrations in serum and tissues were within normal limits . It appeared that the acute abdominal syndrome in these neonatal calves was unrelated to copper deficiency, and that C perfringens, particularly type A, may have had an appreciable contributory role in its pathogenesis. Zentralbl Bakteriol Mikrobiol Hyg {A}, 1987 Jun, 265(1-2), 99 - 112 Methodological aspects of a serodiagnostic Clostridium tumour test--experience with spontaneous canine tumours; Fabricius EM et al.; The paper describes the development of appropriate antigen and method combinations for the microbiological cancer test using the non-oncolysing strain Clostridium butyricum CNRZ 528 in dogs with spontaneous tumours . The diagnostic rod antibodies could be determined quantitatively by the complement fixation test if a short-time warm fixation and a long-time cold fixation procedure were combined in separate runs and if two different antigens, a rod corpuscular antigen and a rod surface antigen, were used . Since complement-fixing antibodies were not always detected in cases of malignant tumours, we additionally used the passive haemagglutination method after pre-absorbing the sera with cross-reacting clostridial antigens . The efficiency of the microbiological cancer test could not be substantially increased, however, by the method combination the reliability of the evaluation of low seropositive titres was improved. Zentralbl Bakteriol Mikrobiol Hyg {A}, 1987 Jun, 265(1-2), 33 - 44 Oncolysis by Clostridium oncolyticum M55 and subsequent enzymatic determination of sialic acid in serum; Marth E et al.; Since the discovery of the "Clostridium tetani phenomenon", various apathogenic clostridia have been used for tumour lysis . Experiments have been conducted to achieve a tumour diagnosis using radiolabelled antibodies to clostridia . In addition, a method has been described that distinguishes, with variable success, between healthy and tumour-carrying animals by means of hemagglutination . The method outlined here uses the fact that malignant cells produce a multitude of sialic acid compounds which lie on the cell membrane and are also connected to the lipid layer of the tumour cell membrane . The apathogenic Clostridium oncolyticum M55 only germinates and multiplies in the malignant tumour tissue . Thus; bacterial hydrolases can enter the tumour tissue and lead to oncolysis . Subsequently the glycocompounds which can be detected by means of an enzymatic determination of the concentration of neuraminic acid (one of the sialic acids) in the serum are washed out into the peripheral blood . We observed these processes in mice in the Ehrlich ascites solid carcinoma and in the Lewis lung carcinoma . Using this method it was possible to detect tumour growth at an early stage with impressive accuracy . The Lewis lung carcinoma which secretes only small amounts of sialic acid glycocompounds cannot be distinguished from the control group by determination of sialic acid concentration . It was possible to detect a 52% increase in the amount of sialic acid after administration of spores of clostridia . This method makes it possible to increase the tumour marker sialic acid through manipulation of the tumour, using apathogenic clostridia, and to measure of sialic acid concentration as an indicator of the metabolic products of the tumour. Zh Mikrobiol Epidemiol Immunobiol, 1987 Jun, (6), 11 - 5 {Determination of the biological activity of Clostridium difficile toxins in in vivo and in vitro experiments}; Tret'iakov VA et al.; The biological activity of the filtrates of 29 C . difficile strains was studied in vivo (suckling white mice) and in vitro (cell cultures of different species and origin) . The action of the filtrates on the experimental models in vivo was evaluated from the cytotoxic effect index, while in vitro the intensity of the cytotoxic effect was evaluated from the percentage of dead cells in the monolayer . The results of the comparative determination of toxicity characteristics in vivo and in vitro demonstrated that cell cultures were more sensitive experimental models than suckling white mice . The use of cell cultures permitted the quantitative evaluation of the cytotoxic activity of the filtrates under study, as well as the detection of their cell-directed action at minimal concentrations. Eur J Clin Microbiol, 1987 Jun, 6(3), 352 - 6 Clostridium difficile-associated diarrhoea in uremic patients; Aronsson B et al.; An outbreak of 94 episodes of Clostridium difficile-associated diarrhoea in 62 patients in a nephrology ward over a two-year period was investigated . Quantitative stool cultures were performed on ten uremic patients not on antibiotics and without diarrhoea and on ten healthy controls . All diarrhoeal episodes were associated with Clostridium difficile, and no other bacterial pathogens were isolated . Thirty-two relapses occurred in 16 patients, fourteen of the relapses without preceding antibiotic exposure . Clostridium difficile could not be isolated from the environment of the patients . Uremic patients, who had a significantly increased number of Clostridium spp . in their stools, are predisposed to Clostridium difficile infections. Arch Surg, 1987 Jun, 122(6), 655 - 61 Clostridium difficile diarrhea in critically ill burned patients; Grube BJ et al.; We followed up 112 patients in the University of Washington Burn Center, Seattle, for the development of Clostridium difficile diarrhea . Diarrhea developed in 20 patients with a mean burn size of 42%, mean age of 38 years, and 49 mean total antibiotic days, for an incidence of 17% . Eleven patients had 16 episodes of nonspecific diarrhea . Nine patients had 11 episodes of C difficile-positive diarrhea and 15 episodes of nonspecific diarrhea for an incidence of 45% of all patients with diarrhea . There were no differences in patient age, burn size, or length of stay between the groups . When the 31 episodes of nonspecific diarrhea were compared with the 11 episodes of C difficile diarrhea, there were no differences between the groups in temperature, albumin levels, or total number of antibiotic days preceding the episodes of diarrhea . The only significant finding that differed between the two types of diarrhea was the white blood cell count on the day of diagnosis . The nonspecific diarrhea was self-limited, requiring antimotility agents in 45% of the episodes . The C difficile diarrhea responded promptly to vancomycin hydrochloride, with resolution of symptoms in an average of 3.3 days . There were two recurrences, and both responded to a second course of vancomycin. Infect Immun, 1987 Jun, 55(6), 1461 - 5 Activation of botulinum C2 toxin by trypsin; Ohishi I; C2 toxin (C2T) elaborated by Clostridium botulinum types C and D is composed of two dissimilar protein components, designated components I and II . The biological activity of C2T is enhanced by treating the toxin with trypsin . This activation of C2T is observed as a result of mixing untrypsinized component I and trypsinized component II but not as a result of mixing trypsinized component I and untrypsinized component II . The data presented here show that the maximum lethality of C2T, determined by mixing untrypsinized component I and trypsinized component II, was attained by treating component II with trypsin at a ratio of 10:1 on a protein basis for 30 min at 35 degrees C at pH 7.5 . The activation of component II was always accompanied by a change in the molecular weight of the component from 101,000 to 88,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) . However, the gel filtration of trypsinized component II resulted in the separation of two active components, with apparent molecular weights, estimated from the elution volume by gel filtration, of 365,000 and 74,000 . The high-molecular-weight component II had hemagglutination and hemolytic activities, whereas the low-molecular-weight component II has only hemagglutination activity . These two molecular species of active component II had approximately the same lethality, when mixed with component I, and gave a single band in SDS-PAGE, with a molecular weight of 88,000, the same as that of trypsin-activated component II under different reaction conditions . The results indicate that the activation of C2T by trypsin is due to the molecular conversion of component II from molecular weight 101,000 to 88,000 as determined by SDS-PAGE and that the trypsin-activated component II tends to form an oligomer of the active component II. Zentralbl Bakteriol Mikrobiol Hyg {A}, 1987 Jun, 265(1-2), 45 - 56 Characterization of proteins, insoluble at low temperature, produced by Clostridium botulinum type C and C . subterminale . Their antigenic relationship with a similar protein synthesized by C . botulinum type G; Gimenez JA et al.; The production of a protein insoluble at low temperature ("cryoprotein"), by cultures of Clostridium botulinum type G has been shown to be a metabolic characteristic also shared by C . botulinum type C and by C . subterminale . These new cryoproteins have been purified and some of their chemical and immunological properties studied . It was found that both proteins were chemically very similar among themselves and to the cryoprotein isolated from C . botulinum type G . All these proteins are formed by a single polypeptide chain of approximately Mr = 180,000, with closely related amino acid compositions, isoelectric points and do not contain either free cysteine or disulfide bridges . Homologous and heterologous radioimmunoassays established the existence of an antigenic similitude among the cryoproteins from C . botulinum type G and C . subterminale thus becoming the first purified antigens which relate both bacterial species . If the production of cryoproteins can be shown to be a generalized phenomenon within the genus Clostridium these substances would provide an important tool to examine immunological and genetical relatedness between strains in this bacterial group. J Appl Bacteriol, 1987 Jun, 62(6), 479 - 90 The effect of sodium chloride and temperature on the rate and extent of growth of Clostridium botulinum type A in pasteurized pork slurry; Gibson AM et al.; A selective medium was used to enumerate Clostridium botulinum growing in the presence of natural spoilage organisms in a model cured pork slurry . The growth responses of a mixed spore inoculum of six strains of Cl . botulinum type A were studied