Effect of Various Sodium Taurocholate Preparations on the Recovery of <i>Clostridium difficile</i> Spores

Kamiya, Shigeru; Yamakawa, Kiyotaka; Ogura, Hisashi; Nakamura, Shinichi

doi:10.1111/j.1348-0421.1987.tb01344.x

Cited by 9 publications

(6 citation statements)

References 10 publications

(9 reference statements)

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“…dzflcile spores, the thioglycollate-lysozyme method was shown to be slightly less effective than the taurocholate method for untreated spores (Kamiya et al, 1987). However, in the present study, it was clearly shown that the heat-or alkalialtered C .…”

Section: Discussioncontrasting

confidence: 42%

“…The second method utilising sodium taurocholate was as described by Raibaud et al (1980) and Wilson et al (1982). Briefly, 0.1 ml of each 10-fold dilution of spore suspension was pipetted into 9-cm petri dishes and mixed with 20ml of GS-BHI agar medium containing sodium taurocholate 0.1% w/v (guaranteed reagent grade, Nakarai Chemicals Ltd, Kyoto, Japan) and was prepared before use as described by Kamiya et al (1987). The relative recovery rate (%) of spores was expressed as the ratio of the number of colonies recovered by either method after heat-or alkalitreatment to that of colonies recovered by the taurocholate method after heating at 70°C for 10 min, a technique which yields nearly 100% recovery (Kamiya et al, 1987).…”

Section: Recovery Of Heat-or Alkali-treated Sporesmentioning

confidence: 99%

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Recovery of spores of Clostridium difficile altered by heat or alkali

Kamiya

Yamakawa²,

Ogura³

et al. 1989

Journal of Medical Microbiology

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Summary. The effect of heating or alkali-treatment on spore recovery in ordinary growth medium was examined for four strains of Clostridium dificile. Heating spores at 80°C for 10 min produced 95-50-99-95% decreases in the recovery rates. Treatment with 0.1 N NaOH for 15 rnin produced 99-47 and 99.83% decreases in spore recovery rates for two of the four strains. The influence of either addition of lysozyme after treatment with sodium thioglycollate (thioglycollate-lysozyme method) or addition of sodium taurocholate (taurocholate method) on recovery of heat-or alkali-treated C. dificile spores was also examined. Viable spores of all strains altered by heating at 90°C or 100°C for 10 min could not be recovered at all by the taurocholate method. Nor did this method allow recovery of alkali-altered spores treated with >0-2 N NaOH for 15 min. On the other hand, 10-47% of altered spores heated at 90°C for 10 min were recovered by the thioglycollate-lysozyme method, and alkali-altered spores treated with 0.1-0.3 N NaOH for 15 min were as completely recovered by this method as untreated spores. These results indicate that the thioglycollate-lysozyme method is more effective than the taurocholate method for recovery of the heat-or alkali-altered C. dificile spores.

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Section: Discussioncontrasting

confidence: 42%

Section: Recovery Of Heat-or Alkali-treated Sporesmentioning

confidence: 99%

Recovery of spores of Clostridium difficile altered by heat or alkali

Kamiya

Yamakawa²,

Ogura³

et al. 1989

Journal of Medical Microbiology

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“…9 Briefly, an 0.1-ml aliquot of ten fold diluted bacterial culture was pipetted into 9-cm-diameter petri dishes and mixed with 20 ml of brain-heart infusion (Nissui Pharmaceutical, Tokyo, Japan) supplemented with 0.8% glucose, 1.0% soluble starch, 0.05% l-cysteine·HCl, 0.1% taurocholic acid sodium salt (Guaranteed reagent grade; Nacalai Tesque, Kyoto, Japan) and 1.3% agar. The inoculated agar plates were incubated anaerobically at 37°C for 3 days.…”

Section: Quantitative Determination Of Bacterial Cellsmentioning

confidence: 99%

Laboratory diagnosis of toxigenic Clostridium difficile by polymerase chain reaction: presence of toxin genes and their stable expression in toxigenic isolates from Japanese individuals

Karasawa

Nojiri

Hayashi

et al. 1999

Journal of Gastroenterology

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Clostridium difficile causes pseudomembranous colitis and antibiotic-associated diarrhea. The definitive diagnosis of C. difficile infection is finally accomplished by the isolation of toxigenic C. difficile. However, only a small number of Japanese clinical laboratories are able to reach a definitive diagnosis of C. difficile infection, probably because simple reliable assays for toxins in the isolates are not available. In this study, we examined the compatibility of a polymerase chain reaction (PCR) assay and tissue culture assay to identify toxigenic C. difficile, in toxigenic and nontoxigenic C. difficile isolates from Japanese patients and healthy carriers. The specificity of PCR primers was demonstrated by restriction endonuclease digestion and seminested PCR in C. difficile VPI 10463 strain. No PCR product was amplified in the eight other clostridial species used to check the specificity of the PCR assay. The detection limit was 10(3) cells. Both toxin A and toxin B genes (the genes encoding the major virulence factors of C. difficile) were detected in 58 toxigenic C. difficile isolates, which showed a wide range of cytotoxic activity in tissue culture assays. Neither of the toxin genes was carried by 40 nontoxigenic strains of C. difficile. The results of this study strongly suggest that a definitive diagnosis of C. difficile infection can be accomplished by PCR detection of the toxin genes rather than by tissue culture assay of isolates.

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“…The germination rate of C. difficile was evaluated by the methods described previously (17,31). To count the number of spores, a suspension of C. difficile cells was heated at 70°C for 10 min to kill all the vegetative cells.…”

Section: Methodsmentioning

confidence: 99%

Molecular and Microbiological Characterization of Clostridium difficile Isolates from Single, Relapse, and Reinfection Cases

et al. 2012

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In this study, we investigated the correlation between the microbiological characteristics of Clostridium difficile clinical isolates and the recurrence of C. difficile-associated disease (CDAD). Twenty C. difficile isolates recovered from 20 single infection cases and 53 isolates from 20 recurrent cases were analyzed by pulsed-field gel electrophoresis (PFGE) and PCR ribotyping, and the cytotoxicity, antimicrobial susceptibility, and sporulation/germination rates of the isolates were examined. Recurrent cases were divided into relapse or reinfection cases by the results of C. difficile DNA typing. Among the 20 recurrent cases, 16 cases (80%) were identified to be relapse cases caused by the initial strain and the remaining 4 cases (20%) were identified to be reinfection cases caused by different strains. All 73 isolates were susceptible to both vancomycin and metronidazole, but resistance against clindamycin, ceftriaxone, erythromycin, and ciprofloxacin was found in 87.7%, 93.2%, 87.7%, and 100% of the isolates, respectively. No correlations between DNA typing group, cytotoxicity, and sporulation rate of isolates and infection status, i.e., single, relapse, or reinfection, were observed. However, the isolates recovered from relapse cases showed a significantly higher germination rate when incubated in medium lacking the germination stimulant sodium taurocholate. These results indicate that the germination ability of C. difficile may be a potential risk factor for the recurrence of CDAD.

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Effect of Various Sodium Taurocholate Preparations on the Recovery of Clostridium difficile Spores

Cited by 9 publications

References 10 publications

Recovery of spores of Clostridium difficile altered by heat or alkali

Recovery of spores of Clostridium difficile altered by heat or alkali

Laboratory diagnosis of toxigenic Clostridium difficile by polymerase chain reaction: presence of toxin genes and their stable expression in toxigenic isolates from Japanese individuals

Molecular and Microbiological Characterization of Clostridium difficile Isolates from Single, Relapse, and Reinfection Cases

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