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Antimicrobial activity of phenyllactic acid against Klebsiella pneumoniae and its effect on cell wall membrane and genomic DNA

  • Veterinary Microbiology - Research Paper
  • Published:
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Abstract

As Klebsiella pneumoniae (KP) has acquired high levels of resistance to multiple antibiotics, it is considered a worldwide pathogen of concern, and substitutes for traditional antibiotics are urgently needed. 3-Phenyllactic acid (PLA) has been reported to have antimicrobial activity against food-borne bacteria. However, there was no experiment evidence for the exact antibacterial effect and mechanism of PLA kills pathogenic KP. In this study, the Oxford cup method indicated that PLA is effective to KP with a minimum inhibitory concentration of 2.5 mg/mL. Furthermore, PLA inhibited the growth and biofilm formation of in a time- and concentration-dependent manner. In vivo, PLA could significantly increase the survival rate of infected mice and reduce the pathological tissue damage. The antibacterial mode of PLA against KP was further explored. Firstly, scanning electron microscopy illustrated the disruption of cellular ultrastructure caused by PLA. Secondly, measurement of leaked alkaline phosphatase demonstrated that PLA disrupted the cell wall integrity of KP and flow cytometry analysis with propidium iodide staining suggested that PLA damaged the cell membrane integrity. Finally, the results of fluorescence spectroscopy and agarose gel electrophoresis demonstrated that PLA bound to genomic DNA and initiated its degradation. The anti-KP mode of action of PLA was attributed to the destruction of the cell wall, membrane, and genomic DNA binding. These findings suggest that PLA has great potential applications as antibiotic substitutes in feed additives against KP infection in animals.

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Data availability

All data analyzed during this study are included in this published article. The raw data of this study are available from the author Gaowei Hu upon reasonable request.

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Funding

This study was supported by Zhejiang Provincial Natural Science Foundation of China (Grant No. LTGN23C180001), Science and Technology Plan Project of Taizhou (Grant No. 22nya08), and National College Students Innovation and Entrepreneurship Program (Grant No. 202210350030).

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Authors and Affiliations

  1. College of Life Sciences, Taizhou key Laboratory of Biomass Functional Materials Development and Application, Taizhou University, Taizhou, Zhejiang, 318000, China

    Jianyun Yu, Chunli Hong, Longfei Yin, Qingbo Ping & Gaowei Hu

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  1. Jianyun Yu
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  2. Chunli Hong
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Correspondence to Gaowei Hu.

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All study procedures were approved by the Animal Care and Use Committee of Taizhou University (Approval No. TZXY-2022-20221046) and were in accordance with the “Zhejiang province animal use nursing ethics guide” (Zhejiang, China).

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ESM 1

Fig. S1. Detection of antibacterial ability of PLA after treatment at different temperatures. 1×MIC PLA was added to 1 mL LB liquid medium and incubated at 4°C，37°Cand 100°Cfor 1 h. After that, 1×106 CFU/mL of KP was added to above LB liquid medium. The mixture PLA and KP was grown at 37°C in a shaking incubator at 180 rpm for 24 h, the bacterial fluid without the PLA was used as the negative control. The growth of KP was determined by the value of OD600. (PNG 90 kb)

High Resolution image (TIF 563 kb)

ESM 2

Fig. S2. Detection of the effect of PLA on bacterial protein expression by SDS-PAGE. KP cells at a density of 1×108 CFU/mL in LB was incubated with different concentrations of PLA (0, 0.5× and 1× MIC) at 37°C with shaking at 180 rpm for 6 h. Then, the cells were collected by centrifuging at 8000×g for 10 min, washed 3 times with NS, and lysed using a bacterial active protein extraction kit. Bacterial proteins were separated by 12% acrylamide SDS-PAGE and stained with Coomassie brilliant blue; M: Protein marker. (PNG 152 kb)

High Resolution image (TIF 597 kb)

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Yu, J., Hong, C., Yin, L. et al. Antimicrobial activity of phenyllactic acid against Klebsiella pneumoniae and its effect on cell wall membrane and genomic DNA. Braz J Microbiol 54, 3245–3255 (2023). https://doi.org/10.1007/s42770-023-01126-8

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