Anti-Bacillus cereus activity of three aromatic plants cultivated in the Region of Murcia (Spain)

Abstract

Bacillus cereus is a Gram-positive, endospore-forming, opportunistic human pathogen that causes two types of food-poisoning syndrome: emesis and diarrhea. The trend towards Refrigerated Processed Foods of Extended Durability (REPFEDs) and the increasing percentage of elderly and immunocompromised people will raise the importance of B. cereus as an aetiological agent of food-borne illness. The aim of this work was to evaluate the chemical variability of the essential oils of three Lamiaceae taxa: Origanum vulgare L., Salvia officinalis subsp. lavandulifolia (Vahl) Gams (syn. Salvia lavandulifolia Vahl), and Thymus hyemalis Lange in relation to their antibacterial activity against B. cereus strain CECT 131. A total of 30 individual plants (10 from each species) were used in the assay. The essential oils were extracted by hydrodistillation and the qualitative and quantitative compositions were analyzed using a gas chromatograph coupled to a mass spectrometric detector (GC-MS). The minimal inhibitory concentrations (MIC) were determined by a microdilution technique, using 96-well microplates. The minimal bactericidal concentrations (MBC) were established by pour-plating. To determine if the vegetative form or the endospore was the cause of the bacterial growth, Würtz staining was used: malachite green binds to the endospores and is used as a contrast dye, while safranin stains the vegetative cells. The tested concentrations of the essential oils ranged from 40 to 10000 ppm for O. vulgare and T. hyemalis, and from 1875 to 60000 ppm for
S. lavandulifolia. The herein tested essential-oil chemotypes of O. vulgare were: (A) 86% of carvacrol, (B) 77% of carvacrol and 6% of thymol, and (C) 65% of carvacrol and 18% of thymol; the chemotypes of S. lavandulifolia were: (D) 37% of camphor, 11% of α-pinene and 8% of eucalyptol, (E) 22% of camphor and 25% of eucalyptol, and (F) 55% of eucalyptol and 13% of camphor; and for
T. hyemalis, the chemotypes were: (G) 52% of carvacrol and 25% p-cymene, (H) 50% of thymol, 25% of p-cymene and 10% of γ-terpinene, and (I) 44% of
p-cymene and 41% of thymol. From the results it can be concluded: that the highest antimicrobial power was exhibited by chemotypes A of O. vulgare and
G of T. hyemalis (MIC = 156-312 ppm), for the exposure times of 24 and 48 h. In the case of S. lavandulifolia, the lowest MIC was that of chemotype D in the concertation range 5000-60000 ppm. The MBC for the endospores, for all the chemotypes tested, was higher than the concentrations evaluated.