Analysis of the germination of individual Clostridium perfringens spores and its heterogeneity

• Published in: Journal of applied microbiology 2011-11, Vol.111 (5), p.1212-1223
• Main Authors: Wang, G, Zhang, P, Paredes‐Sabja, D, Green, C, Setlow, P, Sarker, M.R, Li, Y.‐Q
• Format: Article
• Language: English
• Subjects: Bacillus - physiology; Biological and medical sciences; Clostridium perfringens; Clostridium perfringens - physiology; cortex; dipicolinic acid; Fundamental and applied biological sciences. Psychology; germination; heat; Hot Temperature; Hydrolysis; Microbiology; microscopy; Microscopy, Interference; monitoring; Picolinic Acids - pharmacology; potassium chloride; Potassium Chloride - pharmacology; receptors; spore germination; spores; Spores, Bacterial - physiology
• ISSN: 1364-5072; 1365-2672
• DOI: 10.1111/j.1365-2672.2011.05135.x

Aims: To analyse the germination and its heterogeneity of individual spores of Clostridium perfringens. Methods and Results: Germination of individual wild‐type Cl. perfringens spores was followed by monitoring Ca‐dipicolinic acid (CaDPA) release and by differential interference contrast (DIC) microscopy. Following the addition of KCl that acts via germinant receptors (GRs), there was a long variable lag period (Tlag) with slow release of c. 25% of CaDPA, then rapid release of remaining CaDPA in c. 2 min (ΔTrelease) and a parallel decrease in DIC image intensity, and a final decrease of c. 25% in DIC image intensity during spore cortex hydrolysis. Spores lacking the essential cortex‐lytic enzyme (CLE) (sleC spores) exhibited the same features during GR‐dependent germination, but with longer average Tlag values, and no decrease in DIC image intensity because of cortex hydrolysis after full CaDPA release. The Tlag of wild‐type spores in KCl germination was increased significantly by lower germinant concentrations and suboptimal heat activation. Wild‐type and sleC spores had identical average Tlag and ΔTrelease values in dodecylamine germination that does not utilize GRs. Conclusions: Most of these results were essentially identical to those reported for the germination of individual spores of Bacillus species. However, individual sleC Cl. perfringens spores germinated inefficiently with either KCl or exogenous CaDPA, in contrast to CLE‐deficient Bacillus spores, indicating that germination of these species’ spores is not completely identical. Significance and Impact of the Study: This work provides information on the kinetic germination and its heterogeneity of individual spores of Cl. perfringens.