Genomic insight into Enterococcus faecium biofilm formation isolated from catheter associated urinary tract infection
-
Ayu Rahayu
,
Sayyidah Auliany Aminy
,
Dian Asti Umaroh
,
Niar Gusnaniar
,
Akhirta Atikana
,
Puspita Lisdiyanti
,
Estelle Perraud
,
Christine Imbert
,
Titik Nuryastuti
Abstract
Background and Objectives: Enterococcus faecium is a major nosocomial pathogen frequently associated with catheter-related infections and biofilm-mediated antimicrobial resistance. This study aimed to investigate the genomic determinants and structural characteristics underlying biofilm formation in a clinical E. faecium isolate recovered from a catheter-associated urinary tract infection (CAUTI).
Materials and Methods: A clinical isolate, E. faecium MFKCC-BF.UR33, was subjected to whole-genome sequencing and comprehensive genome annotation to identify biofilm-associated and antimicrobial resistance genes. Phylogenetic analysis was performed to determine its relatedness to reference clinical strains. Structural modeling of selected biofilm-associated proteins was conducted using AlphaFold2 to assess their three-dimensional conformations and potential as therapeutic targets.
Results: Whole-genome sequencing revealed a 3.32 Mb genome with a G+C content of 37.35%, comprising 2,321 predicted coding sequences and no detectable plasmids. Genome annotation identified several biofilm-associated genes, including sgrA, acm, fss3, scm, and efbA, as well as antimicrobial resistance genes (antA/B, tetM, tetL, efmA, and liaFSR). Phylogenetic analysis placed the isolate within the E. faecium clade, closely related to other clinical reference strains. Structural modeling demonstrated high-confidence 3D structures for Fss3 and EfbA proteins.
Conclusion: The integration of genomic and structural analyses provides insights into the genetic and structural basis of biofilm formation in E. faecium and establishes a foundation for future functional and structure-based investigations.
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11. Popović N, Dinić M, Tolinački M, Mihajlović S, Terzić-Vidojević A, Bojić S, et al. New Insight into Biofilm Formation Ability, the Presence of Virulence Genes and Probiotic Potential of Enterococcus sp. Dairy Isolates. Front Microbiol 2018; 9: 78.
12. Kraszewska Z, Skowron K, Kwiecińska-Piróg J, Grudlewska-Buda K, Przekwas J, Wiktorczyk-Kapischke N, et al. Antibiotic Resistance of Enterococcus spp. Isolated from the Urine of Patients Hospitalized in the University Hospital in North-Central Poland, 2016-2021. Antibiotics (Basel) 2022; 11: 1749.
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14. Cattoir V, Giard JC. Antibiotic resistance in Enterococcus faecium clinical isolates. Expert Rev Anti Infect Ther 2014; 12: 239-248.
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20. Kopit LM, Kim EB, Siezen RJ, Harris LJ, Marco ML. Safety of the surrogate microorganism Enterococcus faecium NRRL B-2354 for use in thermal process validation. Appl Environ Microbiol 2014; 80: 1899-1909.
21. Gan YQ, Zhang T, Gan YQ, Zhao Z, Zhu B. Complete genome sequences of two Enterococcus faecium strains and comparative genomic analysis. Exp Ther Med 2020; 19: 2019-2028.
22. Choi S, Park J, Kim EB. Draft genomes and antibiotic resistance profiles of four Enterococcus faecium strains isolated from Cheonggukjang, Korean fermented soybean paste. Korean J Microbiol 2025; 61: 185-187.
23. Hendrickx AP, van Luit-Asbroek M, Schapendonk CM, van Wamel WJ, Braat JC, Wijnands LM, et al. SgrA, a nidogen-binding LPXTG surface adhesin implicated in biofilm formation, and EcbA, a collagen binding MSCRAMM, are two novel adhesins of hospital-acquired Enterococcus faecium. Infect Immun 2009; 77: 5097-5106.
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25. Nuryastuti T, van der Mei HC, Busscher HJ, Iravati S, Aman AT, Krom BP. Effect of cinnamon oil on icaA expression and biofilm formation by Staphylococcus epidermidis. Appl Environ Microbiol 2009; 75: 6850-6855.
26. Choi DG, Baek JH, Han DM, Khan SA, Jeon CO. Comparative pangenome analysis of Enterococcus faecium and Enterococcus lactis provides new insights into the adaptive evolution by horizontal gene acquisitions. BMC Genomics 2024; 25: 28.
27. Zaghloul HAH, El Halfawy NM. Genomic insights into antibiotic-resistance and virulence genes of Enterococcus faecium strains from the gut of Apis mellifera. Microb Genom 2022; 8(11): mgen000896.
28. Khalil MA, Alorabi JA, Al-Otaibi LM, Ali SS, Elsilk SE. Antibiotic Resistance and Biofilm Formation in Enterococcus spp. Isolated from Urinary Tract Infections. Pathogens 2023; 12: 34.
29. Singh KV, La Rosa SL, Somarajan SR, Roh JH, Murray BE. The fibronectin-binding protein EfbA contributes to pathogenesis and protects against infective endocarditis caused by Enterococcus faecalis. Infect Immun 2015; 83: 4487-4494.
30. Qin X, Galloway-Peña JR, Sillanpaa J, Roh JH, Nallapareddy SR, Chowdhury S, et al. Complete genome sequence of Enterococcus faecium strain TX16 and comparative genomic analysis of Enterococcus faecium genomes. BMC Microbiol 2012; 12: 135.
31. Cui P, Feng L, Zhang L, He J, An T, Fu X, et al. Antimicrobial Resistance, Virulence Genes, and Biofilm Formation Capacity Among Enterococcus species From Yaks in Aba Tibetan Autonomous Prefecture, China. Front Microbiol 2020; 11: 1250.
32. Nowakiewicz A, Ziólkowska G, Troscianczyk A, Zieba P, Gnat S. Determination of resistance and virulence genes in Enterococcus faecalis and E. faecium strains isolated from poultry and their genotypic characterization by ADSRRS-fingerprinting. Poult Sci 2017; 96: 986-996.
33. Le Neindre K, Dejoies L, Reissier S, Guérin F, Felden B, Cattoir V. Small RNA-mediated regulation of the tet(M) resistance gene expression in Enterococcus faecium. Res Microbiol 2022; 173: 103941.
34. Tran TC, Panesso D, Zhao A, Khan A, Axell-House DB, Shamoo Y, et al. LiaX, a Member of the LiaFSR System, is Essential for Cell Envelope Adaptation System in Enterococcus faecium. Open Forum Infect Dis 2022; 9(Suppl 2): ofac492.207.
35. Tymoszewska A, Szylińska M, Aleksandrzak-Piekarczyk T. The LiaFSR-LiaX System Mediates Resistance of Enterococcus faecium to Peptide Antibiotics and to Aureocin A53- and Enterocin L50-Like Bacteriocins. Microbiol Spectr 2023; 11(3): e0034323.
36. Jumper J, Evans R, Pritzel A, Green T, Figurnov M, Ronneberger O, et al. Highly accurate protein structure prediction with AlphaFold. Nature 2021; 596: 583-589.
37. Zhang Y, Li P, Pan F, Liu H, Hong P, Liu X, et al. Applications of AlphaFold beyond Protein Structure Prediction [Internet]. 2021 [cited 2024 Jun 30]. Available from: http://biorxiv.org/lookup/doi/10.1101/2021.11.03.467194 doi:10.1101/2021.11.03.467194
38. Laskowski RA, MacArthur MW, Thornton JM. PROCHECK: validation of protein-structure coordinates. In: International Tables for Crystallography. Vol F 2012: 684-687.
39. Tsaban T, Varga JK, Avraham O, Ben-Aharon Z, Khramushin A, Schueler-Furman O. Harnessing protein folding neural networks for peptide-protein docking. Nat Commun 2022; 13: 176.
40. de Been M, Pinholt M, Top J, Bletz S, Mellmann A, van Schaik W, et al. Core Genome Multilocus Sequence Typing Scheme for High- Resolution Typing of Enterococcus faecium. J Clin Microbiol 2015; 53: 3788-3797.
2. Sinha B, Sengupta C, Sahoo PK, Nath T, Mukherjee O (2022). Genetic regulations of microbial biofilm. In: A Complete Guidebook on Biofilm Study. Ed, Roy DN. 1st ed. Elsevier. Amsterdam, pp. 35-58.
3. Schulze A, Mitterer F, Pombo JP, Schild S. Biofilms by bacterial human pathogens: Clinical relevance - development, composition and regulation - therapeutical strategies. Microb Cell 2021; 8: 28-56.
4. Roy R, Tiwari M, Donelli G, Tiwari V. Strategies for combating bacterial biofilms: A focus on anti-biofilm agents and their mechanisms of action. Virulence 2018; 9: 522-554.
5. Maehara Y, Shimono N, Uchida Y, Ogawa E, Otaguro S, Kanamoto Y, et al. Nosocomial transmission of vancomycin-resistant enterococci in our Hospital. Jpn J Environ Infect 2008; 23: 327–331.
6. Assefa M, Amare A. Biofilm-Associated Multi-Drug Resistance in Hospital-Acquired Infections: A Review. Infect Drug Resist 2022; 15: 5061-5068.
7. Gallardo-Moreno AM, González-Martín ML, Bruque JM, Pérez-Giraldo C, Gómez-García AC. The effects of urine and temperature on the physicochemical surface properties and adhesion behaviour of uropathogenic bacteria. J Adhes Sci Technol 2003; 17: 1223-1233.
8. Donelli G, Guaglianone E. Emerging role of Enterococcus spp in catheter-related infections: biofilm formation and novel mechanisms of antibiotic resistance. J Vasc Access 2004; 5: 3-9.
9. Hashem YA, Amin HM, Essam TM, Yassin AS, Aziz RK. Biofilm formation in enterococci: genotype-phenotype correlations and inhibition by vancomycin. Sci Rep 2017; 7: 5733.
10. Zheng JX, Bai B, Lin ZW, Pu ZY, Yao WM, Chen Z, et al. Characterization of biofilm formation by Enterococcus faecalis isolates derived from urinary tract infections in China. J Med Microbiol 2018; 67: 60-67.
11. Popović N, Dinić M, Tolinački M, Mihajlović S, Terzić-Vidojević A, Bojić S, et al. New Insight into Biofilm Formation Ability, the Presence of Virulence Genes and Probiotic Potential of Enterococcus sp. Dairy Isolates. Front Microbiol 2018; 9: 78.
12. Kraszewska Z, Skowron K, Kwiecińska-Piróg J, Grudlewska-Buda K, Przekwas J, Wiktorczyk-Kapischke N, et al. Antibiotic Resistance of Enterococcus spp. Isolated from the Urine of Patients Hospitalized in the University Hospital in North-Central Poland, 2016-2021. Antibiotics (Basel) 2022; 11: 1749.
13. Sattari-Maraji A, Jabalameli F, Node Farahani N, Beigverdi R, Emaneini M. Antimicrobial resistance pattern, virulence determinants and molecular analysis of Enterococcus faecium isolated from children infections in Iran. BMC Microbiol 2019; 19: 156.
14. Cattoir V, Giard JC. Antibiotic resistance in Enterococcus faecium clinical isolates. Expert Rev Anti Infect Ther 2014; 12: 239-248.
15. Peng Z, Yan L, Yang S, Yang D. Antimicrobial-Resistant Evolution and Global Spread of Enterococcus faecium Clonal Complex (CC) 17: Progressive Change from Gut Colonization to Hospital-Adapted Pathogen. China CDC Wkly 2022; 4: 17-21.
16. Govindarajan DK, Kandaswamy K. Virulence factors of uropathogens and their role in host pathogen interactions. Cell Surf 2022; 8: 100075.
17. Høiby N, Bjarnsholt T, Moser C, Bassi GL, Coenye T, Donelli G, et al. ESCMID guideline for the diagnosis and treatment of biofilm infections 2014. Clin Microbiol Infect 2015; 21 Suppl 1: S1-S25.
18. Werneburg GT. Catheter-Associated Urinary Tract Infections: Current Challenges and Future Prospects. Res Rep Urol 2022; 14: 109-133.
19. Hrvat F, Hasanić O, Aleta A, Spahić A, Džuho A, Hukić M (2021). Comparison of Biofilm Category Determination Using TCP Method Depending on Signal Molecule Adherence. In: CMBEBIH 2021. Ed, Badnjevic A, Gurbeta Pokvić L. Springer. Cham, pp. 575-581.
20. Kopit LM, Kim EB, Siezen RJ, Harris LJ, Marco ML. Safety of the surrogate microorganism Enterococcus faecium NRRL B-2354 for use in thermal process validation. Appl Environ Microbiol 2014; 80: 1899-1909.
21. Gan YQ, Zhang T, Gan YQ, Zhao Z, Zhu B. Complete genome sequences of two Enterococcus faecium strains and comparative genomic analysis. Exp Ther Med 2020; 19: 2019-2028.
22. Choi S, Park J, Kim EB. Draft genomes and antibiotic resistance profiles of four Enterococcus faecium strains isolated from Cheonggukjang, Korean fermented soybean paste. Korean J Microbiol 2025; 61: 185-187.
23. Hendrickx AP, van Luit-Asbroek M, Schapendonk CM, van Wamel WJ, Braat JC, Wijnands LM, et al. SgrA, a nidogen-binding LPXTG surface adhesin implicated in biofilm formation, and EcbA, a collagen binding MSCRAMM, are two novel adhesins of hospital-acquired Enterococcus faecium. Infect Immun 2009; 77: 5097-5106.
24. Sun H, Fang Y, Hu S, Lyu D, Dong X, Wang J. Molecular epidemiology, antimicrobial resistance, and virulence determinants of Enterococcus faecium bloodstream isolates: a 7-year study in Shandong, China. BMC Microbiol 2025; 25: 765.
25. Nuryastuti T, van der Mei HC, Busscher HJ, Iravati S, Aman AT, Krom BP. Effect of cinnamon oil on icaA expression and biofilm formation by Staphylococcus epidermidis. Appl Environ Microbiol 2009; 75: 6850-6855.
26. Choi DG, Baek JH, Han DM, Khan SA, Jeon CO. Comparative pangenome analysis of Enterococcus faecium and Enterococcus lactis provides new insights into the adaptive evolution by horizontal gene acquisitions. BMC Genomics 2024; 25: 28.
27. Zaghloul HAH, El Halfawy NM. Genomic insights into antibiotic-resistance and virulence genes of Enterococcus faecium strains from the gut of Apis mellifera. Microb Genom 2022; 8(11): mgen000896.
28. Khalil MA, Alorabi JA, Al-Otaibi LM, Ali SS, Elsilk SE. Antibiotic Resistance and Biofilm Formation in Enterococcus spp. Isolated from Urinary Tract Infections. Pathogens 2023; 12: 34.
29. Singh KV, La Rosa SL, Somarajan SR, Roh JH, Murray BE. The fibronectin-binding protein EfbA contributes to pathogenesis and protects against infective endocarditis caused by Enterococcus faecalis. Infect Immun 2015; 83: 4487-4494.
30. Qin X, Galloway-Peña JR, Sillanpaa J, Roh JH, Nallapareddy SR, Chowdhury S, et al. Complete genome sequence of Enterococcus faecium strain TX16 and comparative genomic analysis of Enterococcus faecium genomes. BMC Microbiol 2012; 12: 135.
31. Cui P, Feng L, Zhang L, He J, An T, Fu X, et al. Antimicrobial Resistance, Virulence Genes, and Biofilm Formation Capacity Among Enterococcus species From Yaks in Aba Tibetan Autonomous Prefecture, China. Front Microbiol 2020; 11: 1250.
32. Nowakiewicz A, Ziólkowska G, Troscianczyk A, Zieba P, Gnat S. Determination of resistance and virulence genes in Enterococcus faecalis and E. faecium strains isolated from poultry and their genotypic characterization by ADSRRS-fingerprinting. Poult Sci 2017; 96: 986-996.
33. Le Neindre K, Dejoies L, Reissier S, Guérin F, Felden B, Cattoir V. Small RNA-mediated regulation of the tet(M) resistance gene expression in Enterococcus faecium. Res Microbiol 2022; 173: 103941.
34. Tran TC, Panesso D, Zhao A, Khan A, Axell-House DB, Shamoo Y, et al. LiaX, a Member of the LiaFSR System, is Essential for Cell Envelope Adaptation System in Enterococcus faecium. Open Forum Infect Dis 2022; 9(Suppl 2): ofac492.207.
35. Tymoszewska A, Szylińska M, Aleksandrzak-Piekarczyk T. The LiaFSR-LiaX System Mediates Resistance of Enterococcus faecium to Peptide Antibiotics and to Aureocin A53- and Enterocin L50-Like Bacteriocins. Microbiol Spectr 2023; 11(3): e0034323.
36. Jumper J, Evans R, Pritzel A, Green T, Figurnov M, Ronneberger O, et al. Highly accurate protein structure prediction with AlphaFold. Nature 2021; 596: 583-589.
37. Zhang Y, Li P, Pan F, Liu H, Hong P, Liu X, et al. Applications of AlphaFold beyond Protein Structure Prediction [Internet]. 2021 [cited 2024 Jun 30]. Available from: http://biorxiv.org/lookup/doi/10.1101/2021.11.03.467194 doi:10.1101/2021.11.03.467194
38. Laskowski RA, MacArthur MW, Thornton JM. PROCHECK: validation of protein-structure coordinates. In: International Tables for Crystallography. Vol F 2012: 684-687.
39. Tsaban T, Varga JK, Avraham O, Ben-Aharon Z, Khramushin A, Schueler-Furman O. Harnessing protein folding neural networks for peptide-protein docking. Nat Commun 2022; 13: 176.
40. de Been M, Pinholt M, Top J, Bletz S, Mellmann A, van Schaik W, et al. Core Genome Multilocus Sequence Typing Scheme for High- Resolution Typing of Enterococcus faecium. J Clin Microbiol 2015; 53: 3788-3797.
| Files | ||
| Issue | Vol 18 No 5 (2026) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v18i5.22866 | |
| Keywords | ||
| Enterococcus faecium Biofilms Drug resistance Microbial Genome Bacterial Catheter-related infections | ||
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How to Cite
1.
Rahayu A, Aminy S, Umaroh D, Gusnaniar N, Atikana A, Lisdiyanti P, Perraud E, Imbert C, Nuryastuti T. Genomic insight into Enterococcus faecium biofilm formation isolated from catheter associated urinary tract infection. Iran J Microbiol. 2026;18(5):752-764.
