Abstract
BackgroundIt is generally reported that garlic (Allium sativum) harms pathogenic but not beneficial bacteria. Although numerous studies supporting the alleged garlic effects on pathogens are available, there are limited studies to prove this claim for beneficial bacteria. We have recently shown that garlic exhibits antibacterial activity against probiotic bifidobacteria. The aim of the current study was to elucidate the mechanism of action of garlic clove extract (GCE) on Bifidobacterium bifidum LMG 11041, B. longum LMG 13197 and B. lactis Bb12 using Fourier transform infrared (FT-IR) spectroscopy and flow cytometry.MethodsCultures (1 × 108 CFU ml-1) were individually incubated for 6 h at 37°C in garlic clove extract containing allicin at a corresponding predetermined minimum bactericidal concentration for each strain. For FTIR, an aliquot of each culture was deposited on CaF2 slide and vacuum dried. The slides were immediately viewed using a Bruker Vertex 70 V FT-IR spectrometer equipped with a Hyperion microscope and data analyzed using OPUS software (version 6, Bruker). Spectra were smoothed with a Savitsky-Goly function algorithim, base-line corrected and normalized. Samples for flow cytometry were stained using the Live/Dead BacLight bacterial viability kit L7012. Data compensation and analysis was performed using a BD FACSAria and FlowJo (version 7.6.1).ResultsFourier transform infrared spectroscopy showed changes in spectral features of lipids and fatty acids in cell membranes, proteins, polysaccharides and nucleic acids. Spectral data as per principle component analysis (PCA) revealed segregation of control and GCE-treated cells for all the tested bifidobacteria. Flow cytometry not only showed increase in numbers of membrane damaged and possibly lysed cells after GCE treatment, but also displayed diffuse light scatter patterns for GCE treated cells, which is evidence for changes to the size, granularity and molecular content of the cells.ConclusionGarlic has multiple target sites in bifidobacteria, penetrating the cell membrane and entering the cytoplasm, where it causes changes to carbohydrates, fatty acids, proteins and nucleic acids. These changes, for example, modification of membrane properties, may prevent exposed bifidobacteria from colonizing the intestinal mucosa. Loss of colonization potential would render them less efficient as probiotics.
Highlights
It is generally reported that garlic (Allium sativum) harms pathogenic but not beneficial bacteria
We have recently reported a change of bifidobacteria from rod to cocci shaped cells with cross- walls due to treatment with garlic clove extract (GCE) [4]
The mechanism of action of garlic towards bifidobacteria is similar to that which was reported for pathogenic bacteria
Summary
It is generally reported that garlic (Allium sativum) harms pathogenic but not beneficial bacteria. We have recently revealed sensitivity of bifidobacteria to antibacterial effects of garlic [4], as well as initiated the process of elucidating its mechanism of action on these bacteria. Electron micrographs showed that garlic induced unusual morphological changes in bifidobacteria [5]. It is not yet known whether garlic inhibition on probiotic cells is confined to the cell membrane or if there are other targets. We anticipate additional targets as allicin, the main active compound in garlic, has been reported to kill pathogens through total inhibition of RNA synthesis, partial inhibition of DNA and protein synthesis, and alteration of the electrochemical ability and induce apoptosis in cells [6,7,8,9]. It is known to affect microbial lipid biosynthesis, signal transduction, as well as react with thiol-containing proteins [10,11,12]
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