Effect of cumin (Cuminum cyminum) seed essential oil on biofilm formation and plasmid Integrity of Klebsiella pneumoniae

Abstract

Seeds of the cumin plant (Cuminum cyminum L.) have been used since many years in Iranian traditional medicine. We assessed the effect of cumin seed essential oil on the biofilm-forming ability of Klebsiella pneumoniae strains and on the integrity of a native resistance plasmid DNA from K. pneumoniae isolates, treated with essential oil. Antibacterial coaction between the essential oil and selected antibiotic disks were determined for inhibiting K. pneumoniae. The essential oil of the cumin seeds was obtained by hydrodistillation in a Clavenger system. A simple method for the formation of biofilms on semiglass lamellas was established. The biofilms formed were observed by scanning electron microscopy (SEM). The effect of essential oil on plasmid integrity was studied through the induction of R-plasmid DNA degradation. The plasmid was incubated with essential oil, and agarose gel electrophoresis was performed. Disk diffusion assay was employed to determine the coaction. The essential oil decreased biofilm formation and enhanced the activity of the ciprofloxacin disk. The incubation of the R-plasmid DNA with essential oil could not induce plasmid DNA degradation. The results of this study suggest the potential use of cumin seed essential oil against K. pneumoniae in vitro, may contribute to the in vivo efficacy of this essential oil.

Keywords: Biofilm; Cuminum cyminum; Klebsiella pneumoniae; essential oil; plasmid.

Figure 1

Biofilm formation on the surface of lamellas in broth media containing essential oil dilution by Klebsiella pneumoniae ATCC 13883. The bars indicate means ± standard errors of the means (error bars) from three experiments. (Reproduction size at column width)

Figure 2

SEM images of Klebsiella pneumoniae ATCC 13883 biofilms exposed to sub-MICs of essential oil of Cuminum cyminum seeds. Biofilms of K. pneumoniae formed by 24-hour incubation, in the presence of essential oil. Preparation and observation under the SEM were carried out as described in the text. The bar represents 20 μm. (Reproduction size at column width) a: Control. A biofilm formed in the medium without essential oil. b: Formed in the presence of 1/4 × MIC of essential oil (0.2-0.87 μg/ml). c: Formed in the presence of 1/2 × MIC of essential oil (0.4-1.75 μg/ml)

Figure 3

A percentage of 0.8% agarose gel electrophoresis of the plasmid (resistance plasmid) treated with essential oil. Aliquots of R-plasmid DNA (5 μl) were incubated with 5 μl (35 μg/ml) of the essential oil for 45 minutes at 37°C. After incubation, the reaction mixtures were submitted to agarose gel electrophoresis. Lane 1: Plasmid extracted from isolate 1 plus essential oil. Lane 2: Plasmid extracted from isolate 2 plus essential oil. I = supercoiled form (SC); II = open circle form (OC), and III = linear form (L). (Reproduction size at column width)