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Cited by 20Probiotics and Fermented FoodsAnimal Nutrition and PhysiologyAquaculture disease management and microbiota

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Assessment of the Safety and Probiotic Properties of Enterococcus faecium B13 Isolated from Fermented Chili

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Xiao, Jingmin, Chen, Cai, Fu, Zhuxian, Wang, Shumin, Luo, Fan·2024·Microorganisms
DOI 10.3390/microorganisms12050994PMC PMC11123876
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Abstract

Enterococcus faecium B13, selected from fermentation chili, has been proven to promote animal growth by previous studies, but it belongs to opportunistic pathogens, so a comprehensive evaluation of its probiotic properties and safety is necessary. In this study, the probiotic properties and safety of B13 were evaluated at the genetic and phenotype levels in vitro and then confirmed in vivo. The genome of B13 contains one chromosome and two plasmids. The average nucleotide identity indicated that B13 was most closely related to the fermentation-plant-derived strain. The strain does not carry the major virulence genes of the clinical E. faecium strains but contains aac(6′)-Ii, ant (6)-Ia, msrC genes. The strain had a higher tolerance to acid at pH 3.0, 4.0, and 0.3% bile salt and a 32.83% free radical DPPH clearance rate. It can adhere to Caco-2 cells and reduce the adhesion of E. coli to Caco-2 cells. The safety assessment revealed that the strain showed no hemolysis and did not exhibit gelatinase, ornithine decarboxylase, lysine decarboxylase, or tryptophanase activity. It was sensitive to twelve antibiotics but was resistant to erythromycin, rifampicin, tetracycline, doxycycline, and minocycline. Experiments in vivo have shown that B13 can be located in the ileum and colon and has no adverse effects on experiment animals. After 28 days of feeding, B13 did not remarkable change the α-diversity of the gut flora or increase the virulence genes. Our study demonstrated that E. faecium B13 may be used as a probiotic candidate.

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Crossref · 10.3390/microorganisms12050994

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Figures

Figure 1 The complete genome of E. faecium B13. (A) Chromosomal genome map of E. faecium B13. (B) Plasmid 1 genome map of E. faecium B13. (C) Plasmid 2 genome map of E. faecium B13. The outermost circle of the circle map is the genome-sized logo; each scale is 0.1 Mb (for Figure 1A), 10 kb (for Figure 1B), and 1 kb (for Figure 1C). The second and third circles are CDS on the forward and reverse chains, and the assorted colors indicate different COG classifications of the CDS. The fourth circle is rRNA, or tRNA. The fifth circle is the GC content, and the outward orange part indicates that the GC content in the region is higher than the whole-genome average GC content. The inward green portion indicates that the GC content in the region is low; the higher peak value indicates a greater difference from the average GC content.
Figure 1 The complete genome of E. faecium B13. (A) Chromosomal genome map of E. faecium B13. (B) Plasmid 1 genome map of E. faecium B13. (C) Plasmid 2 genome map of E. faecium B13. The outermost circle of the circle map is the genome-sized logo; each scale is 0.1 Mb (for Figure 1A), 10 kb (for Figure 1B), and 1 kb (for Figure 1C). The second and third circles are CDS on the forward and reverse chains, and the assorted colors indicate different COG classifications of the CDS. The fourth circle is rRNA, or tRNA. The fifth circle is the GC content, and the outward orange part indicates that the GC content in the region is higher than the whole-genome average GC content. The inward green portion indicates that the GC content in the region is low; the higher peak value indicates a greater difference from the average GC content.CC BY · PubMed Central
Figure 2 Probiotic properties and safety assessment in vitro. (A) Auto-aggregation abilities of E. faecium B13. (B) Inhibition of Escherichia coli adhesion to Caco-2 cells by different modes. (C) Gelatinase activity of E. faecium B13 compared to S. aureus ATCC25923 (positive control) and sterile water (negative control). (D) Decarboxylase activity. (E) Hemolytic activities. * Indicates p < 0.05; ns indicates no significance.
Figure 2 Probiotic properties and safety assessment in vitro. (A) Auto-aggregation abilities of E. faecium B13. (B) Inhibition of Escherichia coli adhesion to Caco-2 cells by different modes. (C) Gelatinase activity of E. faecium B13 compared to S. aureus ATCC25923 (positive control) and sterile water (negative control). (D) Decarboxylase activity. (E) Hemolytic activities. * Indicates p < 0.05; ns indicates no significance.CC BY · PubMed Central
Figure 3 Fluorescence changes and histological sections of E. faecium B13 in the intestine. (A) Fluorescence values in intestines during 24 h. (B) Fluorescence values in the intestines during the whole experiment. (C) Paraffin sections were used to observe the distribution of E. faecium B13 in the colon and ileum. (D) Hematoxylin–eosin staining of the colon. scale bar: 50 μm.
Figure 3 Fluorescence changes and histological sections of E. faecium B13 in the intestine. (A) Fluorescence values in intestines during 24 h. (B) Fluorescence values in the intestines during the whole experiment. (C) Paraffin sections were used to observe the distribution of E. faecium B13 in the colon and ileum. (D) Hematoxylin–eosin staining of the colon. scale bar: 50 μm.CC BY · PubMed Central
Figure 4 Microbial community composition in the intestine. (A) Classification of microflora at the phylum level. (B) Classification of microflora at the gene level. (C) The relative abundance of Heminiphilus faecis between B13 and control groups. (D) PCoA analysis between the control and B13 groups.
Figure 4 Microbial community composition in the intestine. (A) Classification of microflora at the phylum level. (B) Classification of microflora at the gene level. (C) The relative abundance of Heminiphilus faecis between B13 and control groups. (D) PCoA analysis between the control and B13 groups.CC BY · PubMed Central

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Citations

Cites

Show all 90 references

Research works printed but not linked: 4

  1. Ding, 2017, Effects of Enterococcus faecium B13 Yielding Bacteriocins on Growth Performance, Nutrient Digestibility, Serum Immune Indexes and Fecal Microbiota of Weaned Piglets, Acta Vet. Zootech. Sin., 48, 1902
  2. Kara, 2020, Investigation of Antibiotic Resistance and Virulence Factors of Enterococcus faecium and Enterococcus faecalis Strains Isolated from Clinical Samples, Mikrobiyol. Bul., 54, 26
  3. Zhu, 2017, Novel lnu(G) gene conferring resistance to lincomycin by nucleotidylation, located on Tn6260 from Enterococcus faecalis E531, J. Antimicrob. Chemother., 72, 993
  4. Wei, 2020, Optimization of Fermentation Conditions for Bacteriocin Production from Enterococcus faecium B13, Food Res. Dev., 41, 194

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Metadata

Authors
Xiao, J., Chen, C., Fu, Z., Wang, S., & Luo, F.
Affiliations
  • Southwest Minzu University (CN)
  • Institute of Qinghai-Tibetan Plateau, Southwest Minzu University, Chengdu 610041, China
Corresponding author
Luo, Fan
ORCID
Venue
Microorganisms
Publisher
MDPI AG
Published
2024-05-15
Volume
12
Issue
5
Pages
994
ISSN
2076-2607 (ISSN-L)
DOI
10.3390/microorganisms12050994
PMC
PMC11123876
OpenAlex
W4396965634
Cited by
17
References
90
Citations by year
2026: 7 · 2025: 10
FWCI
3.65
Type
journal article
Language
en
Open access
Licence
Locations
  • Microorganisms (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • Natural Science Foundation of Sichuan Province (2022NSFSC0114, ZYN2024083)
  • the Central Universities Project of Southwest Minzu University (2022NSFSC0114, ZYN2024083)
Topics
  • Life Sciences › Agricultural and Biological Sciences › Food Science › Probiotics and Fermented Foods
  • Life Sciences › Agricultural and Biological Sciences › Animal Science and Zoology › Animal Nutrition and Physiology
  • Life Sciences › Immunology and Microbiology › Immunology › Aquaculture disease management and microbiota
Keywords
Enterococcus faecium, probiotic candidate, virulence genes, antibiotic resistance, acid and bile tolerance, antioxidant activity, cell adhesion, gut microbiota, plasmids, animal growth promotion

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