Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2020 Aug 11:8:e9543.
doi: 10.7717/peerj.9543. eCollection 2020.

Antibacterial activity and mechanism of sanguinarine against Providencia rettgeri in vitro

Qian Zhang  1   2   3   4 , Yansi Lyu  1 , Jingkai Huang  1 , Xiaodong Zhang  1 , Na Yu  1 , Ziping Wen  1 , Si Chen  3   5
Affiliations

Antibacterial activity and mechanism of sanguinarine against Providencia rettgeri in vitro

Qian Zhang et al. PeerJ. .

Abstract

Background: Sanguinarine (SAG), a benzophenanthridine alkaloid, occurs in Papaveraceas, Berberidaceae and Ranunculaceae families. Studies have found that SAG has antioxidant, anti-inflammatory, and antiproliferative activities in several malignancies and that it exhibits robust antibacterial activities. However, information reported on the action of SAG against Providencia rettgeri is limited in the literature. Therefore, the present study aimed to evaluate the antimicrobial and antibiofilm activities of SAG against P. rettgeri in vitro.

Methods: The agar dilution method was used to determine the minimum inhibitory concentration (MIC) of SAG against P. rettgeri. The intracellular ATP concentration, intracellular pH (pHin), and cell membrane integrity and potential were measured. Confocal laser scanning microscopy (CLSM), field emission scanning electron microscopy (FESEM), and crystal violet staining were used to measure the antibiofilm formation of SAG.

Results: The MIC of SAG against P. rettgeri was 7.8 μg/mL. SAG inhibited the growth of P. rettgeri and destroyed the integrity of P. rettgeri cell membrane, as reflected mainly through the decreases in the intracellular ATP concentration, pHin and cell membrane potential and significant changes in cellular morphology. The findings of CLSM, FESEM and crystal violet staining indicated that SAG exhibited strong inhibitory effects on the biofilm formation of P. rettgeri and led to the inactivity of biofilm-related P. rettgeri cells.

Keywords: Antibiofilm; Antimicrobial; Providencia rettgeri; Sanguinarine.

PubMed Disclaimer

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1
Figure 1. Effects of SAG on the growth kinetics of P. rettgeri cells.
Effect of SAG on the growth of P. rettgeri. Bacterial cells were incubated and grown in TBS with 0, 1/8, 1/4, 1 and 2 MIC of SAG at 37 °C. Error bars are SD of three replicates.
Figure 2
Figure 2. SAG treatment led to a decrease in the intracellular ATP concentrations, pH and membrane potential of P. rettgeri.
Effects of SAG on P. rettgeri: (A) intracellular and (B) extracellular ATP levels, (C) membrane potential, and (D) pHin. Data are expressed as mean ± SD. **P < 0.01 vs. 0 MIC.
Figure 3
Figure 3. SAG treatment increased the cell membrane permeability of P. rettgeri.
Effects of SAG on the cell membrane integrity of P. rettgeri cells through CLSM: (A) P. rettgeri cells exposed to 1% DMSO; (B) P. rettgeri cells exposed to SAG at 1 MIC, and (C) P. rettgeri cells exposed to SAG at 2 MIC.
Figure 4
Figure 4. Treatment with SAG led to changes in the cell morphology of P. rettgeri.
Effects of SAG on the cell structure of P. rettgeri through TEM. (A) P. rettgeri cells treated with 1% DMSO; (B) P. rettgeri cells treated with SAG at 1 MIC, and (C) P. rettgeri cells treated with SAG at 2 MIC.
Figure 5
Figure 5. Treatment with SAG led to changes in the cell morphology of P. rettgeri.
Effects of SAG on the cell morphology of P. rettgeri using FESEM: (A) P. rettgeri cells exposed to 1% DMSO; (B) P. rettgeri cells exposed to SAG at 1 MIC, and (C) P. rettgeri cells exposed to SAG at 2 MIC.
Figure 6
Figure 6. Inhibitory effects of SAG on the biofilm formation of P. rettgeri.
The biofilm formation index was tested by crystal violet staining with different concentrations of SAG in 96-well plates. Data are expressed as mean ± SD. *P < 0.05 vs. 0 MIC, **P < 0.01 vs. 0 MIC.
Figure 7
Figure 7. Effect of SAG on the biofilm formation of P. rettgeri.
Images of FESEM (A–D; magnification, 10,000′×g) and CLSM (E–H). (A and E) P. rettgeri cells exposed to 1% DMSO; (B and F) P. rettgeri cells exposed to SAG at 1/16 MIC; (C and G) P. rettgeri cells exposed to SAG at 1/8 MIC, and (D and H) P. rettgeri cells exposed to SAG at 1/4 MIC.
Figure 8
Figure 8. Inactivation effect of SAG on the biofilm of P.rettgeri cells.
Inactivation effect of SAG on P. rettgeri cells within biofilms. 1D (A–D) and 3D (E–H) images of CLSM. (A and E) P. rettgeri cells within biofilms unexposed to SAG. (B and F) P. rettgeri cells within biofilms exposed to SAG at 1 MIC. (C and G) P. rettgeri cells within biofilms exposed to SAG at 2 MIC. (D and H) P. rettgeri cells within biofilms exposed to SAG at 4 MIC.
Figure 9
Figure 9. SAG could change the biofilm matrix composition of P. rettgeri cells.
Effects of different concentrations of SAG on the levels of carbohydrates, extracellular proteins, and extracellular DNA inside P. rettgeri biofilms. (A–D) eDNA labeled red (PI). (E–H) Carbohydrates labeled green (WGA). (I–L) Proteins labeled red (SYPRO Ruby). Scale bar, 10 μm.
Figure 10
Figure 10. Effect of SAG on biofilms of diffusion.
(A) P. rettgeri cells treated with 0 MIC and gatifloxacin; (B) P. rettgeri cells treated with 1/16 MIC and gatifloxacin; (C) P. rettgeri cells treated with 1/8 MIC and gatifloxacin; (D) P. rettgeri cells treated with 1/4 MIC and gatifloxacin.
See this image and copyright information in PMC

References

    1. Armbruster CE, Smith SN, Yep A, Mobley HLT. Increased incidence of urolithiasis and bacteremia during Proteus mirabilis and Providencia stuartii coinfection due to synergistic induction of urease activity. Journal of Infectious Diseases. 2014;209(10):1524–1532. doi: 10.1093/infdis/jit663. - DOI - PMC - PubMed
    1. European Committee for Antimicrobial Susceptibility Testing (EUCAST) of the European Society of Clinical Microbiology and Infectious Dieases (ESCMID) EUCAST definitive document E.DEF 3.1, June 2000: determination of minimum inhibitory concentrations (MICs) of antibacterial agents by agar dilution. Clinical Microbiology and Infection: The Official Publication of the European Society of Clinical Microbiology and Infectious Diseases. 2000;6(9):509–515. doi: 10.1046/j.1469-0691.2000.00142.x. - DOI - PubMed
    1. Firatli E, Unal T, Onan U, Sandalli P. Antioxidative activities of some chemotherapeutics: a possible mechanism in reducing gingival inflammation. Journal of Clinical Periodontology. 1994;21(10):680–683. doi: 10.1111/j.1600-051X.1994.tb00786.x. - DOI - PubMed
    1. Giaouris E, Heir E, Hébraud M, Chorianopoulos N, Langsrud S, Møretrø T, Habimana O, Desvaux M, Renier S, Nychas G-J. Attachment and biofilm formation by foodborne bacteria in meat processing environments: causes, implications, role of bacterial interactions and control by alternative novel methods. Meat Science. 2014;97(3):298–309. doi: 10.1016/j.meatsci.2013.05.023. - DOI - PubMed
    1. Gonelimali FD, Lin J, Miao W, Xuan J, Charles F, Chen M, Hatab SR. Antimicrobial properties and mechanism of action of some plant extracts against food pathogens and spoilage microorganisms. Frontiers in Microbiology. 2018;9:1639. doi: 10.3389/fmicb.2018.01639. - DOI - PMC - PubMed

LinkOut - more resources