Strain-Dependent Adhesion Variations of Shouchella clausii Isolated from Healthy Human Volunteers: A Study on Cell Surface Properties and Potential Probiotic Benefits
Abstract
:1. Introduction
2. Materials and Methods
2.1. Bacterial Strains and Culture Conditions
2.2. Characterization of S. clausii Strains
2.3. Adhesion of S. clausii Strains to HT-29 and Caco-2 Cell Lines
2.3.1. Cell Adhesion Assay
2.3.2. Competitive Exclusion of Salmonella Typhimurium Adhesion from HT-29 Cell Line Using the S. clausii Strains
2.3.3. Adhesion to ECM Components
2.3.4. Determination of Cell Surface Characteristics
Cell Surface Hydrophobicity
Auto-Aggregation and Co-Aggregation Assay
2.4. Genome Sequencing and Annotation
2.5. In Silico Characterization of Cell Surface Protein
2.6. Molecular Docking Studies
2.7. Statistical Analysis
3. Results and Discussion
3.1. In Vitro Resistance of S. clausii Strains to Simulated Gastric and Intestinal Fluids
3.2. Safety Assessment of S. clausii Strains
3.3. Disease-Preventing Traits of S. clausii Strains
3.4. Health-Promoting Effects of S. clausii Strains
Properties | S. clausii Strains | |||
---|---|---|---|---|
B619/R | B603/Nb | B106 | B637/Nm | |
Identification | ||||
% 16S rRNA gene identity with S. clausii DSM 8716 | 99.85 | 99.85 | 99.85 | 99.87 |
Average Nucleotide Identity (ANI) (%) | 95.45 | 95.42 | 95.44 | 95.44 |
Digital DNA-DNA Hybridization (DDH) (%) | 62.5 | |||
Physiological properties a | ||||
Optimum Temperature (°C) | 30 (22–45) | 30 (22–45) | 30 (22–45) | 30 (22–45) |
Optimum pH | 10 (7–10) | 10 (7–10) | 8 (7–10) | 10 (7–10) |
Growth in presence of NaCl (%) | 4 (0–10%) | 2 (0–10%) | 4 (0–10%) | 4 (0–10%) |
Survival in Gastrointestinal fluids b | ||||
Survival (%) in SGF (120 min) | 88 ± 8 | 57 ± 3 | 94 ± 19 | 88 ± 4 |
Survival (%) in SIF (180 min) | 70 ± 1 | 36 ± 0 | 66 ± 2 | 71 ± 2 |
Antibiotic resistance profile c (*strains were resistant to the listed antibiotics) | Streptomycin, Neomycin, Clarithromycin, Metronidazole, Chloramphenicol | Novobiocin, Rifampicin, Clarithromycin, Metronidazole, Amoxicillin, Chloramphenicol | Chloramphenicol, Clarithromycin, Metronidazole | Tetracycline, Chloramphenicol Clarithromycin, Metronidazole, Amoxicillin |
Disease Preventing traits | ||||
Antimicrobial activity against pathogens d | P. aeruginosa (14.5 ± 0.7 mm) | P. aeruginosa (27 ± 1.4 mm), S. aureus subsp. aureus (20 ± 2.1 mm) | P. aeruginosa (11 ± 0.8 mm), E. faecalis (24.5 ± 0.7 mm), and S. aureus subsp. aureus (23 ± 2.6 mm) | P. aeruginosa (15.6 ± 1.1 mm), E. faecalis (10.6 ± 1.1 mm), and S. aureus subsp. aureus (18 ± 2.8 mm) |
Health Promoting traits | ||||
Protease activity e | + | + | + | + |
Lipase activity f | + | + | + | - |
β-galactosidase activity g | + | + | + | + |
Antioxidant activity (%) h | 80 ± 2 | 95 ± 1 | 73 ± 3 | 68 ± 1 |
Cholesterol-reducing ability (%) in SIF i | 46 ± 7 | 55 ± 1 | 52 ± 1 | 60 ± 2 |
Exo-polysaccharide production (mg/L) j | 467 ± 10 | 463 ± 9 | 625 ± 11 | 532 ± 5 |
3.5. Adhesion of S. clausii Strains to Human Colonic Epithelial Cells
3.6. Competitive Exclusion of S. Typhimurium Adhesion by S. clausii to HT-29 Cells
3.7. In Vitro Binding of S. clausii Strains to ECM Components
3.8. Cell Surface Properties of S. clausii Strains
3.9. Genomic Insights into the Adhesion Potential of S. clausii Strains
3.10. Comparative Analysis of Adhesion Proteins in S. clausii Strains
3.11. In Silico Characterization of the Cell Surface Protein of S. clausii Strains
3.11.1. Predicted 3D-Structure of the Cell Surface Protein
3.11.2. Docking of the Cell Wall Surface Anchor Family Protein (LPxTG Motif) with Gut Mucins
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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S. clausii Strain | Number of S. clausii Cells Adhering/100 HT-29 Cells | Number of S. clausii Cells Adhering/100 Caco-2 Cells |
---|---|---|
B619/R | 360 ± 14 | 182 ± 10 |
B603/Nb | 114 ± 6 | 106 ± 9 |
B106 | 211 ± 23 | 210 ± 30 |
B637/Nm | 11 ± 3 | 19 ± 1 |
S. clausii Strain | % Exclusion of S. Typhimurium from HT-29 Cell Line by S. clausii Strain |
---|---|
B619/R | 37 ± 3 |
B603/Nb | 19 ± 1 |
B106 | 7 ± 1 |
B637/Nm | 18 ± 1 |
S. clausii Strain | Binding to ECM Components (O.D. 600 nm) | ||
---|---|---|---|
Mucin | Fibrinogen | Collagen | |
B619/R | 0.250 ± 0.008 | 0.203 ± 0.008 | 0.175 ± 0.004 |
B603/Nb | 0.206 ± 0.039 | 0.105 ± 0.002 | 0.202 ± 0.003 |
B106 | 0.241 ± 0.032 | 0.154 ± 0.005 | 0.163 ± 0.003 |
B637/Nm | 0.247 ± 0.009 | 0.204 ± 0.002 | 0.048 ± 0.002 |
Strains of S. clausii | Hydrophobicity (%) | ||||
---|---|---|---|---|---|
Chloroform | Ethyl Acetate | Toluene | Xylene | n-Hexane | |
B619/R | 19.1 ± 7.3 | 19.3 ± 0.5 | 32.7 ± 1.8 | 12.1 ± 0.1 | 3.1 ± 1.2 |
B603/Nb | 17.6 ± 0.2 | 20.4 ± 0.3 | 44.8 ± 0.5 | 33.1 ± 1.3 | 19.7 ± 0.7 |
B106 | 11.0 ± 0 | 4.4 ± 1.5 | 11.2 ± 0.2 | 4.2 ± 0.1 | 4.6 ± 1.2 |
B637/Nm | 27.3 ± 3.6 | 57.4 ± 0.5 | 67.4 ± 1.1 | 61.7 ± 0.4 | 60.4 ± 0 |
Strains of S. clausii | Auto-aggregation (%) | % Co-Aggregation of S. clausii Strains with Pathogens after 4 h | |||||||
---|---|---|---|---|---|---|---|---|---|
E. coli | S. aureus subsp. aureus | E. faecalis | S. dysenteriae | K. pneumoniae | P. aeruginosa | E. aerogenes | S. Typhimurium | ||
B619/R | 43.1 ± 4.4 | 14.8 ± 1.7 | 38.1 ± 3.6 | 16.2 ± 1.0 | 42.1 ± 1.3 | 10.4 ± 1.1 | 12.4 ± 1.5 | 37.3 ± 4.4 | 24.3 ± 3.6 |
B603/Nb | 35.8 ± 2.4 | 45.5 ± 0.2 | 39 ±0.5 | 28 ± 0.01 | 28.5 ± 5.4 | 28.4 ± 0.8 | 41.2 ± 9.3 | 27.1 ± 2.1 | 32.1 ± 0.3 |
B106 | 45.7 ± 0.4 | 30 ± 1.2 | 42.9 ± 0.8 | 28 ± 3.4 | 26.3 ± 3.3 | 21.6 ± 2.1 | 42.5 ± 3.4 | 26.9 ± 3.1 | 24.5 ± 0.6 |
B637/Nm | 75.9 ± 0.9 | 33 ± 0 | 43.9 ± 0.1 | 34.1 ± 0.08 | 41.6 ± 2.2 | 30.3 ± 1.2 | 35.8 ± 0.08 | 40.0 ± 0.07 | 29.3 ± 1.4 |
Docking Complex | Energy (kcal/mol) |
---|---|
LPxTG with MUC1 | −635.4 |
LPxTG with MUC3A | −938.5 |
LPxTG with MUC4 | −1317.1 |
LPxTG with MUC12 | −1122.9 |
LPxTG with MUC13 | −952.1 |
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Dhakephalkar, T.; Pisu, V.; Margale, P.; Chandras, S.; Shetty, D.; Wagh, S.; Dagar, S.S.; Kapse, N.; Dhakephalkar, P.K. Strain-Dependent Adhesion Variations of Shouchella clausii Isolated from Healthy Human Volunteers: A Study on Cell Surface Properties and Potential Probiotic Benefits. Microorganisms 2024, 12, 1771. https://doi.org/10.3390/microorganisms12091771
Dhakephalkar T, Pisu V, Margale P, Chandras S, Shetty D, Wagh S, Dagar SS, Kapse N, Dhakephalkar PK. Strain-Dependent Adhesion Variations of Shouchella clausii Isolated from Healthy Human Volunteers: A Study on Cell Surface Properties and Potential Probiotic Benefits. Microorganisms. 2024; 12(9):1771. https://doi.org/10.3390/microorganisms12091771
Chicago/Turabian StyleDhakephalkar, Tanisha, Vaidehi Pisu, Prajakta Margale, Siddhi Chandras, Deepa Shetty, Shilpa Wagh, Sumit Singh Dagar, Neelam Kapse, and Prashant K. Dhakephalkar. 2024. "Strain-Dependent Adhesion Variations of Shouchella clausii Isolated from Healthy Human Volunteers: A Study on Cell Surface Properties and Potential Probiotic Benefits" Microorganisms 12, no. 9: 1771. https://doi.org/10.3390/microorganisms12091771