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In Vitro Assessment of Lactobacillus crispatus UBLCp01, Lactobacillus gasseri UBLG36, and Lactobacillus johnsonii UBLJ01 as a Potential Vaginal Probiotic Candidate

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Abstract

In this study, Lactobacillus crispatus UBLCp01, Lactobacillus gasseri UBLG36, and Lactobacillus johnsonii UBLJ01 isolated from the vagina of healthy reproductive age Indian women were screened for beneficial probiotic properties. These strains showed the ability to survive acidic and simulated vaginal fluid conditions and could adhere to mucin. Lact. gasseri UBLG36, and Lact. johnsonii UBLJ01 produced d- and l-lactic acid, whereas Lact. crispatus UBLCp01 produced hydrogen peroxide and d- and l-lactic acid. All strains inhibited the growth of pathogens (Escherichia coli, Gardnerella vaginalis, Proteus mirabilis, and Candida albicans) and were capable of co-aggregating with them with varying degrees. Strains secreted exopolysaccharides and formed biofilms under in vitro conditions. Safety assessment showed that these strains had a usual antibiotic susceptibility profile, did not produce hemolysins, gelatinases, and mucin degrading enzymes. Based on strain characteristics and beneficial properties, we believe that these strains are promising candidates for human trials to confirm their ability to prevent/treat vaginal dysbiosis and maintain a healthy vaginal eco-system.

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All data is included in the text; however, the raw data of this article will be made available by the authors, without undue reservation, to any qualified researcher.

References

  1. Smith SB, Ravel J (2017) The vaginal microbiota, host defence and reproductive physiology. J Physiol 595:451–463. https://doi.org/10.1113/JP271694

    Article  CAS  PubMed  Google Scholar 

  2. Ravel J, Gajer P, Abdo Z, Schneider GM, Koenig SS, McCulle SL, Karlebach S, Gorle R, Russell J, Tacket CO, Brotman RM, Davis CC, Ault K, Peralta L, Forney LJ (2011) Vaginal microbiome of reproductive-age women. Proc Natl Acad Sci USA 108S1:4680–4687. https://doi.org/10.1073/pnas.1002611107

    Article  Google Scholar 

  3. Amabebe E, Anumba DOC (2018) The vaginal microenvironment: the physiologic role of Lactobacilli. Front Med (Lausanne) 5:181. https://doi.org/10.3389/fmed.2018.00181

    Article  PubMed  Google Scholar 

  4. Macklaim JM, Fernandes AD, Di Bella JM, Hammond JA, Reid G, Gloor GB (2013) Comparative meta-RNA-seq of the vaginal microbiota and differential expression by Lactobacillus iners in health and dysbiosis. Microbiome 1:12. https://doi.org/10.1186/2049-2618-1-12

    Article  PubMed  PubMed Central  Google Scholar 

  5. Stafford GP, Parker JL, Amabebe E, Kistler J, Reynolds S, Stern V, Paley M, Anumba DOC (2017) Spontaneous preterm birth is associated with differential expression of vaginal metabolites by Lactobacilli-dominated microflora. Front Physiol 8:615. https://doi.org/10.3389/fphys.2017.00615

    Article  PubMed  PubMed Central  Google Scholar 

  6. Chee WJY, Chew SY, Than LTL (2020) Vaginal microbiota and the potential of Lactobacillus derivatives in maintaining vaginal health. Microb Cell Fact 19:203. https://doi.org/10.1186/s12934-020-01464-4

    Article  PubMed  PubMed Central  Google Scholar 

  7. Song SD, Acharya KD, Zhu JE, Deveney CM, Walther-Antonio MRS, Tetel MJ, Chia N (2020) Daily vaginal microbiota fluctuations associated with natural hormonal cycle, contraceptives, diet, and exercise. mSphere 5:e00593-e620. https://doi.org/10.1128/mSphere.00593-20

    Article  PubMed  PubMed Central  Google Scholar 

  8. Chen X, Lu Y, Chen T, Li R (2021) The female vaginal microbiome in health and bacterial vaginosis. Front Cell Infect Microbiol 11:631972. https://doi.org/10.3389/fcimb.2021.631972

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Peebles K, Velloza J, Balkus JE, McClelland RS, Barnabas RV (2019) High global burden and costs of bacterial vaginosis: a systematic review and meta-analysis. Sex Transm Dis 46:304–311. https://doi.org/10.1097/OLQ.0000000000000972

    Article  PubMed  Google Scholar 

  10. Machado D, Castro J, Palmeira-de-Oliveira A, Martinez-de-Oliveira J, Cerca N (2016) Bacterial vaginosis biofilms: challenges to current therapies and emerging solutions. Front Microbiol 6:1528. https://doi.org/10.3389/fmicb.2015.01528

    Article  PubMed  PubMed Central  Google Scholar 

  11. Jeng HS, Yan TR, Chen JY (2020) Treating vaginitis with probiotics in non-pregnant females: a systematic review and meta-analysis. Exp Ther Med 20:3749–3765. https://doi.org/10.3892/etm.2020.9090

    Article  PubMed  PubMed Central  Google Scholar 

  12. Bohbot JM, Daraï E, Bretelle F, Brami G, Daniel C, Cardot JM (2018) Efficacy and safety of vaginally administered lyophilized Lactobacillus crispatus IP 174178 in the prevention of bacterial vaginosis recurrence. J Gynecol Obstet Hum Reprod 47:81–86. https://doi.org/10.1016/j.jogoh.2017.11.005

    Article  CAS  PubMed  Google Scholar 

  13. Geshnizgani AM, Onderdonk AB (1992) Defined medium simulating genital tract secretions for growth of vaginal microflora. J Clin Microbiol 30:1323–1326. https://doi.org/10.1128/JCM.30.5.1323-1326.1992

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Pedersen C, Jonsson H, Lindberg JE, Roos S (2004) Microbiological characterization of wet wheat distillers’ grain, with focus on isolation of lactobacilli with potential as probiotics. Appl Environ Microbiol 70:1522–1527. https://doi.org/10.1128/aem.70.3.1522-1527.2004

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Ahire JJ, Jakkamsetty C, Kashikar MS, Lakshmi SG, Madempudi RS (2021) In vitro evaluation of probiotic properties of Lactobacillus plantarum UBLP40 isolated from traditional indigenous fermented food. Probiotics Antimicrob Proteins. https://doi.org/10.1007/s12602-021-09775-7

    Article  PubMed  Google Scholar 

  16. Zhou JS, Gopal PK, Gill HS (2001) Potential probiotic lactic acid bacteria Lactobacillus rhamnosus (HN001), Lactobacillus acidophilus (HN017) and Bifidobacterium lactis (HN019) do not degrade gastric mucin in vitro. Int J Food Microbiol 63:81–90. https://doi.org/10.1016/s0168-1605(00)00398-6

    Article  CAS  PubMed  Google Scholar 

  17. Ahire JJ, Dicks LM (2014) 2,3-dihydroxybenzoic acid-containing nanofiber wound dressings inhibit biofilm formation by Pseudomonas aeruginosa. Antimicrob Agents Chemother 58:2098–2104. https://doi.org/10.1128/AAC.02397-13

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Rosenberg M, Gutnick D, Rosenberg E (1980) Adherence of bacteria to hydrocarbons: a simple method for measuring cell-surface hydrophobicity. FEMS Microbiol Lett 9:29–33. https://doi.org/10.1111/j.1574-6968.1980.tb05599.x

    Article  CAS  Google Scholar 

  19. Ahire JJ, Kashikar MS, Lakshmi SG, Madempudi R (2020) Identification and characterization of antimicrobial peptide produced by indigenously isolated Bacillus paralicheniformis UBBLi30 strain. 3 Biotech 10:112. https://doi.org/10.1007/s13205-020-2109-6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Del Re B, Sgorbati B, Miglioli M, Palenzona D (2000) Adhesion, autoaggregation and hydrophobicity of 13 strains of Bifidobacterium longum. Lett Appl Microbiol 31:438–442. https://doi.org/10.1046/j.1365-2672.2000.00845.x

    Article  PubMed  Google Scholar 

  21. Campana R, van Hemert S, Baffone W (2017) Strain-specific probiotic properties of lactic acid bacteria and their interference with human intestinal pathogens invasion. Gut Pathog 9:12. https://doi.org/10.1186/s13099-017-0162-4

    Article  PubMed  PubMed Central  Google Scholar 

  22. Ahire JJ (2012) Studies on probiotic microorganism(s) and its biogenic metabolite(s). Ph.D. Dissertation, North Maharashtra University, India

  23. Matuschek E, Brown DF, Kahlmeter G (2014) Development of the EUCAST disk diffusion antimicrobial susceptibility testing method and its implementation in routine microbiology laboratories. Clin Microbiol Infect 20:O255–O266. https://doi.org/10.1111/1469-0691.12373

    Article  CAS  PubMed  Google Scholar 

  24. Clinical and Laboratory Standards Institute (2020) Performance standards for antimicrobial susceptibility testing. 30th ed. CSLI supplement M100.Wayne, PA. https://clsi.org/media/3481/m100ed30_sample.Pdf

  25. Andrews JM (2001) Determination of minimum inhibitory concentrations. J Antimicrob Chemother 48:5–16. https://doi.org/10.1093/jac/48.suppl_1.5

    Article  CAS  PubMed  Google Scholar 

  26. EFSA (2018) Guidance on the characterisation of microorganisms used as feed additives or as production organisms. EFSA J 16:e05206. https://doi.org/10.2903/j.efsa.2018.5206

    Article  Google Scholar 

  27. Pino A, Bartolo E, Caggia C, Cianci A, Randazzo CL (2019) Detection of vaginal lactobacilli as probiotic candidates. Sci Rep 9:3355. https://doi.org/10.1038/s41598-019-40304-3

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Stingele F, Neeser JR, Mollet B (1996) Identification and characterization of the eps (Exopolysaccharide) gene cluster from Streptococcus thermophilus Sfi6. J Bacteriol 178:1680–1690. https://doi.org/10.1128/jb.178.6.1680-1690.1996

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Neveling DP, Ahire JJ, Laubscher W, Rautenbach M, Dicks LMT (2020) Genetic and phenotypic characteristics of a multi-strain probiotic for broilers. Curr Microbiol 77:369–387. https://doi.org/10.1007/s00284-019-01797-3

    Article  CAS  PubMed  Google Scholar 

  30. Ahire JJ, Dicks LM (2015) Nisin incorporated with 2,3-dihydroxybenzoic acid in nanofibers inhibits biofilm formation by a methicillin-resistant strain of Staphylococcus aureus. Probiotics Antimicrob Proteins 7:52–59. https://doi.org/10.1007/s12602-014-9171-5

    Article  CAS  PubMed  Google Scholar 

  31. López-Moreno A, Aguilera M (2021) Vaginal probiotics for reproductive health and related dysbiosis: systematic review and meta-analysis. J Clin Med 10:1461. https://doi.org/10.3390/jcm10071461

    Article  PubMed  PubMed Central  Google Scholar 

  32. Massicotte R, Mafu AA, Ahmad D, Deshaies F, Pichette G, Belhumeur P (2017) Comparison between flow cytometry and traditional culture methods for efficacy assessment of six disinfectant agents against nosocomial bacterial species. Front Microbiol 8:112. https://doi.org/10.3389/fmicb.2017.00112

    Article  PubMed  PubMed Central  Google Scholar 

  33. Pan M, Hidalgo-Cantabrana C, Goh YJ, Sanozky-Dawes R, Barrangou R (2020) Comparative analysis of Lactobacillus gasseri and Lactobacillus crispatus isolated from human urogenital and gastrointestinal tracts. Front Microbiol 10:3146. https://doi.org/10.3389/fmicb.2019.03146

    Article  PubMed  PubMed Central  Google Scholar 

  34. Paavonen J (1983) Physiology and ecology of the vagina. Scand J Infect Dis Suppl 40:31–35

    CAS  PubMed  Google Scholar 

  35. Tomás MSJ, Nader-Macías ME (2007) Effect of a medium simulating vaginal fluid on the growth and expression of beneficial characteristics of potentially probiotic lactobacilli. In: Mendez-Vilas A (ed) Communicating Current Research and Educational Topics and Trends in Applied Microbiology, 1st edn. Badajoz, Formatex, Spain, pp 732–739

  36. Nader-Macías ME, Juárez Tomás MS (2015) Profiles and technological requirements of urogenital probiotics. Adv Drug Deliv Rev 92:84–104. https://doi.org/10.1016/j.addr.2015.03.016

    Article  CAS  PubMed  Google Scholar 

  37. Weerkamp AH, Uyen HM, Busscher HJ (1988) Effect of zeta potential and surface energy on bacterial adhesion to uncoated and saliva-coated human enamel and dentin. J Dental Res 67:1483–1487. https://doi.org/10.1177/00220345880670120801

    Article  CAS  Google Scholar 

  38. Zeng Z, Zuo F, Marcotte H (2019) Putative adhesion factors in vaginal Lactobacillus gasseri DSM 14869: functional characterization. Appl Environ Microbiol 85:e00800-e819. https://doi.org/10.1128/AEM.00800-19

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. He Y, Niu X, Wang B, Na R, Xiao B, Yang H (2020) Evaluation of the inhibitory effects of Lactobacillus gasseri and Lactobacillus crispatus on the adhesion of seven common lower genital tract infection-causing pathogens to vaginal epithelial cells. Front Med (Lausanne) 7:284. https://doi.org/10.3389/fmed.2020.00284

    Article  PubMed  Google Scholar 

  40. Ocaña VS, Nader-Macías ME (2002) Vaginal lactobacilli: self- and co-aggregating ability. Br J Biomed Sci 59:183–190. https://doi.org/10.1080/09674845.2002.11783657

    Article  PubMed  Google Scholar 

  41. Tachedjian G, Aldunate M, Bradshaw CS, Cone RA (2017) The role of lactic acid production by probiotic Lactobacillus species in vaginal health. Res Microbiol 168:782–792. https://doi.org/10.1016/j.resmic.2017.04.001

    Article  CAS  PubMed  Google Scholar 

  42. Nasioudis D, Beghini J, Bongiovanni AM, Giraldo PC, Linhares IM, Witkin SS (2015) α-Amylase in vaginal fluid: association with conditions favorable to dominance of Lactobacillus. Reprod Sci 22:1393–1398. https://doi.org/10.1177/1933719115581000

    Article  CAS  PubMed  Google Scholar 

  43. O’Hanlon DE, Moench TR, Cone RA (2011) In vaginal fluid, bacteria associated with bacterial vaginosis can be suppressed with lactic acid but not hydrogen peroxide. BMC Infect Dis 11:200. https://doi.org/10.1186/1471-2334-11-200

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Scillato M, Spitale A, Mongelli G, Privitera GF, Mangano K, Cianci A, Stefani S, Santagati M (2021) Antimicrobial properties of Lactobacillus cell-free supernatants against multidrug-resistant urogenital pathogens. MicrobiologyOpen 10:e1173. https://doi.org/10.1002/mbo3.1173

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Terraf MC, Juárez Tomás MS, Nader-Macías ME, Silva C (2012) Screening of biofilm formation by beneficial vaginal lactobacilli and influence of culture media components. J Appl Microbiol 113:1517–1529. https://doi.org/10.1111/j.1365-2672.2012.05429.x

    Article  PubMed  Google Scholar 

  46. Verstraelen H, Verhelst R, Claeys G, De Backer E, Temmerman M, Vaneechoutte M (2009) Longitudinal analysis of the vaginal microflora in pregnancy suggests that L. crispatus promotes the stability of the normal vaginal microflora and that L. gasseri and/or L. iners are more conducive to the occurrence of abnormal vaginal microflora. BMC Microbiol 9:116. https://doi.org/10.1186/1471-2180-9-116

    Article  PubMed  PubMed Central  Google Scholar 

  47. Angelin J, Kavitha M (2020) Exopolysaccharides from probiotic bacteria and their health potential. Int J Biol Macromol 162:853–865. https://doi.org/10.1016/j.ijbiomac.2020.06.190

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Donnarumma G, Molinaro A, Cimini D, De Castro C, Valli V, De Gregorio V, De Rosa M, Schiraldi C (2014) Lactobacillus crispatus L1: high cell density cultivation and exopolysaccharide structure characterization to highlight potentially beneficial effects against vaginal pathogens. BMC Microbiol 14:137. https://doi.org/10.1186/1471-2180-14-137

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Li T, Teng D, Mao R, Hao Y, Wang X, Wang J (2020) A critical review of antibiotic resistance in probiotic bacteria. Food Res Int 19:109571. https://doi.org/10.1016/j.foodres.2020.109571

    Article  CAS  Google Scholar 

  50. Sirichoat A, Flórez AB, Vázquez L, Buppasiri P, Panya M, Lulitanond V, Mayo B (2020) Antibiotic susceptibility profiles of lactic acid bacteria from the human vagina and genetic basis of acquired resistances. Int J Mol Sci 21:2594. https://doi.org/10.3390/ijms21072594

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Funding

This work was supported by Unique Biotech Limited, Hyderabad, India.

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  1. Centre for Research and Development, Unique Biotech Limited, Plot No. 2, Phase II, MN Park, Hyderabad, Telangana, 500078, India

    J. J. Ahire, S. Sahoo, M. S. Kashikar, A. Heerekar, S. G. Lakshmi & R. S. Madempudi

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  1. J. J. Ahire
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  2. S. Sahoo
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  3. M. S. Kashikar
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Contributions

Ahire JJ, Lakshmi SG, and Madempudi RS contributed to the study conception, Ahire JJ design the study. Material preparation, data collection, and analysis were performed by Ahire JJ, Kashikar MS, Sahoo S. Heerekar A re-confirmed the strains by DNA analysis. The first draft of the manuscript was written by Ahire JJ and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

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Correspondence to J. J. Ahire.

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This study does not contain any work related with the participation of humans and/or animals.

Competing Interests

Ahire JJ, Kashikar MS, Heerekar A, Lakshmi SG were employed by Unique Biotech Limited. Sahoo S worked as a project intern. Madempudi RS is the Managing Director of Unique Biotech Limited. This does not alter our adherence to journal policies on sharing data and materials.

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Ahire, J.J., Sahoo, S., Kashikar, M.S. et al. In Vitro Assessment of Lactobacillus crispatus UBLCp01, Lactobacillus gasseri UBLG36, and Lactobacillus johnsonii UBLJ01 as a Potential Vaginal Probiotic Candidate. Probiotics & Antimicro. Prot. 15, 275–286 (2023). https://doi.org/10.1007/s12602-021-09838-9

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