Limits...
The analysis of para-cresol production and tolerance in Clostridium difficile 027 and 012 strains.

Dawson LF, Donahue EH, Cartman ST, Barton RH, Bundy J, McNerney R, Minton NP, Wren BW - BMC Microbiol. (2011)

Bottom Line: It has been proposed that the hpdBCA operon, rarely found in other gut microflora, encodes the enzymes responsible for the conversion of p-HPA to p-cresol.We show that the PCR-ribotype 027 strain R20291 quantitatively produced more p-cresol in-vitro and was significantly more tolerant to p-cresol than the sequenced strain 630 (PCR-ribotype 012).The mutants were equally able to tolerate p-cresol compared to the respective parent strains, suggesting that tolerance to p-cresol is not linked to its production.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Infectious & Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London, WC1E 7HT, UK.

ABSTRACT

Background: Clostridium difficile is the major cause of antibiotic associated diarrhoea and in recent years its increased prevalence has been linked to the emergence of hypervirulent clones such as the PCR-ribotype 027. Characteristically, C. difficile infection (CDI) occurs after treatment with broad-spectrum antibiotics, which disrupt the normal gut microflora and allow C. difficile to flourish. One of the relatively unique features of C. difficile is its ability to ferment tyrosine to para-cresol via the intermediate para-hydroxyphenylacetate (p-HPA). P-cresol is a phenolic compound with bacteriostatic properties which C. difficile can tolerate and may provide the organism with a competitive advantage over other gut microflora, enabling it to proliferate and cause CDI. It has been proposed that the hpdBCA operon, rarely found in other gut microflora, encodes the enzymes responsible for the conversion of p-HPA to p-cresol.

Results: We show that the PCR-ribotype 027 strain R20291 quantitatively produced more p-cresol in-vitro and was significantly more tolerant to p-cresol than the sequenced strain 630 (PCR-ribotype 012). Tyrosine conversion to p-HPA was only observed under certain conditions. We constructed gene inactivation mutants in the hpdBCA operon in strains R20291 and 630Δerm which curtails their ability to produce p-cresol, confirming the role of these genes in p-cresol production. The mutants were equally able to tolerate p-cresol compared to the respective parent strains, suggesting that tolerance to p-cresol is not linked to its production.

Conclusions: C. difficile converts tyrosine to p-cresol, utilising the hpdBCA operon in C. difficile strains 630 and R20291. The hypervirulent strain R20291 exhibits increased production of and tolerance to p-cresol, which may be a contributory factor to the virulence of this strain and other hypervirulent PCR-ribotype 027 strains.

Show MeSH

Related in: MedlinePlus

Tolerance to p-cresol. Strains R20291 and 630 were tested for their in-vitro tolerance to 0.1% p-cresol. * indicates a significant difference p < 0.01 Student's T-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3102038&req=5

Figure 1: Tolerance to p-cresol. Strains R20291 and 630 were tested for their in-vitro tolerance to 0.1% p-cresol. * indicates a significant difference p < 0.01 Student's T-test.

Mentions: The tolerance of strains 630 and R20291 to p-cresol was assessed in BHI broth as CFU counts per ml, expressed as a proportion of the untreated control for a four hour incubation period with 0.1% p-cresol (Figure 1). Strain R20291 (PCR-ribotype 027) showed a significant increase in survival to 0.1% p-cresol compared to strain 630 (PCR-ribotype 012) p < 0.01 using a Student's t-test (Figure 1). There was no significant difference in tolerance to p-cresol between 630 and 630Δerm, an erythromycin sensitive spontaneous mutant (data not shown). The 630Δerm strain was essential to construct and select gene inactivation mutants for further investigations of p-cresol tolerance and production, therefore subsequent analysis was performed with the 630Δerm strain.


The analysis of para-cresol production and tolerance in Clostridium difficile 027 and 012 strains.

Dawson LF, Donahue EH, Cartman ST, Barton RH, Bundy J, McNerney R, Minton NP, Wren BW - BMC Microbiol. (2011)

Tolerance to p-cresol. Strains R20291 and 630 were tested for their in-vitro tolerance to 0.1% p-cresol. * indicates a significant difference p < 0.01 Student's T-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3102038&req=5

Figure 1: Tolerance to p-cresol. Strains R20291 and 630 were tested for their in-vitro tolerance to 0.1% p-cresol. * indicates a significant difference p < 0.01 Student's T-test.
Mentions: The tolerance of strains 630 and R20291 to p-cresol was assessed in BHI broth as CFU counts per ml, expressed as a proportion of the untreated control for a four hour incubation period with 0.1% p-cresol (Figure 1). Strain R20291 (PCR-ribotype 027) showed a significant increase in survival to 0.1% p-cresol compared to strain 630 (PCR-ribotype 012) p < 0.01 using a Student's t-test (Figure 1). There was no significant difference in tolerance to p-cresol between 630 and 630Δerm, an erythromycin sensitive spontaneous mutant (data not shown). The 630Δerm strain was essential to construct and select gene inactivation mutants for further investigations of p-cresol tolerance and production, therefore subsequent analysis was performed with the 630Δerm strain.

Bottom Line: It has been proposed that the hpdBCA operon, rarely found in other gut microflora, encodes the enzymes responsible for the conversion of p-HPA to p-cresol.We show that the PCR-ribotype 027 strain R20291 quantitatively produced more p-cresol in-vitro and was significantly more tolerant to p-cresol than the sequenced strain 630 (PCR-ribotype 012).The mutants were equally able to tolerate p-cresol compared to the respective parent strains, suggesting that tolerance to p-cresol is not linked to its production.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Infectious & Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London, WC1E 7HT, UK.

ABSTRACT

Background: Clostridium difficile is the major cause of antibiotic associated diarrhoea and in recent years its increased prevalence has been linked to the emergence of hypervirulent clones such as the PCR-ribotype 027. Characteristically, C. difficile infection (CDI) occurs after treatment with broad-spectrum antibiotics, which disrupt the normal gut microflora and allow C. difficile to flourish. One of the relatively unique features of C. difficile is its ability to ferment tyrosine to para-cresol via the intermediate para-hydroxyphenylacetate (p-HPA). P-cresol is a phenolic compound with bacteriostatic properties which C. difficile can tolerate and may provide the organism with a competitive advantage over other gut microflora, enabling it to proliferate and cause CDI. It has been proposed that the hpdBCA operon, rarely found in other gut microflora, encodes the enzymes responsible for the conversion of p-HPA to p-cresol.

Results: We show that the PCR-ribotype 027 strain R20291 quantitatively produced more p-cresol in-vitro and was significantly more tolerant to p-cresol than the sequenced strain 630 (PCR-ribotype 012). Tyrosine conversion to p-HPA was only observed under certain conditions. We constructed gene inactivation mutants in the hpdBCA operon in strains R20291 and 630Δerm which curtails their ability to produce p-cresol, confirming the role of these genes in p-cresol production. The mutants were equally able to tolerate p-cresol compared to the respective parent strains, suggesting that tolerance to p-cresol is not linked to its production.

Conclusions: C. difficile converts tyrosine to p-cresol, utilising the hpdBCA operon in C. difficile strains 630 and R20291. The hypervirulent strain R20291 exhibits increased production of and tolerance to p-cresol, which may be a contributory factor to the virulence of this strain and other hypervirulent PCR-ribotype 027 strains.

Show MeSH
Related in: MedlinePlus