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. 2016 Jul 14;11(7):e0159006.
doi: 10.1371/journal.pone.0159006. eCollection 2016.

Antimicrobial Activity and Possible Mechanism of Action of Citral against Cronobacter sakazakii

Chao Shi  1 , Kaikuo Song  1 , Xiaorong Zhang  1 , Yi Sun  1 , Yue Sui  1 , Yifei Chen  1 , Zhenyu Jia  1 , Huihui Sun  1 , Zheng Sun  1 , Xiaodong Xia  1   2
Affiliations

Antimicrobial Activity and Possible Mechanism of Action of Citral against Cronobacter sakazakii

Chao Shi et al. PLoS One. .

Abstract

Citral is a flavor component that is commonly used in food, beverage and fragrance industries. Cronobacter sakazakii is a food-borne pathogen associated with severe illness and high mortality in neonates and infants. The objective of the present study was to evaluate antimicrobial effect of citral against C. sakazakii strains. The minimum inhibitory concentration (MIC) of citral against C. sakazakii was determined via agar dilution method, then Gompertz models were used to quantitate the effect of citral on microbial growth kinetics. Changes in intracellular pH (pHin), membrane potential, intracellular ATP concentration, and membrane integrity were measured to elucidate the possible antimicrobial mechanism. Cell morphology changes were also examined using a field emission scanning electron microscope. The MICs of citral against C. sakazakii strains ranged from 0.27 to 0.54 mg/mL, and citral resulted in a longer lag phase and lower growth rate of C. sakazakii compared to the control. Citral affected the cell membrane of C. sakazakii, as evidenced by decreased intracellular ATP concentration, reduced pHin, and cell membrane hyperpolarization. Scanning electron microscopy analysis further confirmed that C. sakazakii cell membranes were damaged by citral. These findings suggest that citral exhibits antimicrobial effect against C. sakazakii strains and could be potentially used to control C. sakazakii in foods. However, how it works in food systems where many other components may interfere with its efficacy should be tested in future research before its real application.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Growth curves of C. sakazakii ATCC 29544 cultured in TSB with various concentrations of citral.
The lines represent the fit of the experimental data to the modified Gompertz model. Bars represent the standard deviation (n = 3).
Fig 2
Fig 2. Effects of citral on intracellular ATP production by C. sakazakii ATCC 29544.
Values represent the means of triplicate measurements. Bars represent the standard deviation (n = 3). **P ≤ 0.01.
Fig 3
Fig 3. Effects of citral on the intracellular pH of C. sakazakii ATCC 29544.
Values represent the means of triplicate measurements. Bars represent the standard deviation (n = 3). **P ≤ 0.01
Fig 4
Fig 4. Effects of citral on the membrane potentials of C. sakazakii ATCC 29544.
Negative values indicate hyperpolarization. Values represent the means of triplicate measurements. Bars represent the standard deviation (n = 3). **P ≤ 0.01.
Fig 5
Fig 5. Scanning electron micrographs of C. sakazakii ATCC 29544 untreated (A), treated with citral at MIC for 2 h (B), and treated with citral at 2MIC for 2 h (C).
See this image and copyright information in PMC

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