Role of Dipicolinic Acid in Heat Resistance of Spores of Clostridium botulinum and Clostridium sporogenes PA3679 by Thermal and Pressure-assisted Thermal Processing

Abstract

Dipicolinic acid (DPA) is a major constituent of spores and reportedly provides protection against inactivation by various thermal processes; however, the relationship between DPA and resistance towards pressure-assisted thermal processing is not well understood. Thermal and pressure-assisted thermal inactivation studies of Clostridium botulinum nonproteolytic strains QC-B and 610-F, proteolytic strain Giorgio-A, and thermal surrogate Clostridium sporogenes PA3679 spores suspended in ACES buffer (0.05 M, pH 7.0) were performed to determine if a relationship exists between DPA release and log reduction of spores. Thermal inactivation at 80, 83, and 87 °C for nonproteolytic strains and 101, 105, and 108 °C for the proteolytic strain and thermal surrogate were conducted. Pressure-assisted thermal inactivation for nonproteolytic strains at 83 °C/600 MPa and for the proteolytic strain and thermal surrogate at 105 °C/600 MPa were performed. Surviving spores were enumerated by 5-tube MPN method for log reductions and analyzed for released DPA by liquid chromatography-tandem mass spectrometry. The correlation between MPN log reductions, released DPA, and D-values were calculated. A positive correlation between released DPA and log reduction of spores was observed for QC-B and 610-F at 80 and 83 °C (r = 0.6073 - 0.7755; P < 0.01). At 87 °C, a positive correlation was detected for 610-F (r = 0.4242, P < 0.05) and no correlation was observed for QC-B (r = 0.1641; P > 0.05). A strong, positive correlation (r = 0.8359 - 0.9284; P < 0.05) between released DPA and log reduction of spores was observed for Giorgio-A at 101, 105, and 108 °C, and a strong, positive correlation (r = 0.8402; P < 0.05) was observed for PA3679 at 101 °C. A positive correlation (r = 0.5646 - 0.6724; P < 0.01) was observed for QC-B, 610-F, and Giorgio-A after pressure-assisted thermal treatment. No correlation (r = 02494; P > 0.05) was found for PA3679 after pressure-assisted thermal treatment. These results suggest a correlation exists between DPA release and heat resistance; however, the level of correlation varied between strains and temperatures. The findings from this research may aid in the development of spore inactivation strategies targeting the thermal resistance profiles of various strains of C. botulinum spores.

Keywords: Clostridium botulinum; Dipicolinic acid; Pressure-assisted thermal processing; Spore inactivation; Spore resistance; Thermal processing.