Isolation and Characterization of Endophytic Bacteria from Piper longum


Endophytes are non-pathogenic microorganisms that reside within internal tissues of plant, without causing any apparent symptoms of infection. In an attempt to identify endophytes from Piper longum, the Indian long pepper, a combination of conventional and molecular approaches was used. Using culture-dependent approach, six different bacterial isolates were obtained from various surface-sterilized parts (roots, nodes, internodes, petioles, leaves and spikes) of the plant. In general, roots harboured maximum concentration of endophytic bacterial isolates, while leaves contained the minimum levels. These endophytes were analyzed on the basis of colony morphology and biochemical characteristics. Based on the results obtained after BLAST search of 16S rDNA sequence in NCBI database, the endophytes isolated from Piper longum showed highest similarity to Endophytic bacteria 135L-3, Enterobacter sp. SQ6-43, Bacillus casamancensis strain TN3, Alishewanella sp. JS-30, Bacterium B28 and Enterobacter ludwigii strain g45. Most of the identified bacteria belong to the phylum Proteobacteria and Firmicutes, whose members have been reported to act as growth-promoting bacteria in other plant species. Some of these endophytes tested positive for the ability to produce indole-3-acetic acid and hydrogen cyanide, suggesting their potential roles in plant growth promotion and biological control against pathogens, respectively.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2


  1. 1.

    Bacon CW, White J (2000) Microbial endophytes. Marcel Dekker Inc, New York

    Google Scholar 

  2. 2.

    Jia M, Chen L, Xin HL, Zheng CJ, Rahman K, Han T, Qin LP (2016) A friendly relationship between endophytic fungi and medicinal plants: a systematic review. Front Microbiol 7:906.

    Article  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Leifert C, Cassells AC (2001) Microbial hazards in plant tissue and cell cultures. In Vitro Cell Dev Biol Plant 37:133–138

    Article  Google Scholar 

  4. 4.

    Richardson AE, Barea JM, McNeill AM, Prigent-Combaret C (2009) Acquisition of phosphorus and nitrogen in the rhizosphere and plant growth promotion by microorganisms. Plant Soil 321:305–339

    CAS  Article  Google Scholar 

  5. 5.

    deSanti Ferrara FI, Oliveira ZM, Gonzales HHS, Floh EIS, Barbosa HRB (2012) Endophytic and rhizospheric enterobacteria isolated from sugar cane have different potentials for producing plant growth-promoting substances. Plant Soil 353:409–417

    Article  Google Scholar 

  6. 6.

    Saravanakumar D, Lavanya N, Muthumeena B, Raguchander T, Suresh S, Samiyappan R (2008) Pseudomonas fluorescens enhances resistance and natural enemy population in rice plants against leaf folder pest. J Appl Entomol 132:469–479

    Article  Google Scholar 

  7. 7.

    Senthilkumar M, Swarnalakshmi K, Govindasamy V, Lee YK, Annapurna K (2009) Biocontrol potential of soybean bacterial endophytes against charcoal rot fungus, Rhizoctonia bataticola. Curr Microbiol 58:288–293

    CAS  Article  Google Scholar 

  8. 8.

    Stajner D, Kevresan S, Gasic O, Mimica-Dukic N, Zongli H (1997) Nitrogen and Azotobacter chroococcum enhance oxidative stress tolerance in sugar beet. Biol Plant 39:441–445

    CAS  Article  Google Scholar 

  9. 9.

    Stierle A, Strobel G, Stierle D (1993) Taxol and taxane production by Taxomyces andreanae, an endophytic fungus of Pacific yew. Science 260(5105):214–216

    CAS  Article  Google Scholar 

  10. 10.

    Brader G, Compant S, Mitter B, Trognitz F, Sessitsch A (2014) Metabolic potential of endophytic bacteria. Curr Opin Biotechnol 27:30–37

    CAS  Article  Google Scholar 

  11. 11.

    Gouda S, Das G, Sen SK, Shin HS, Patra JK (2016) Endophytes: a treasure house of bioactive compounds of medicinal importance. Front Microbiol 7:1538.

    Article  PubMed  PubMed Central  Google Scholar 

  12. 12.

    Rani D, Dantu PK (2012) Direct shoot regeneration from nodal, internodal and petiolar segments of Piper longum L. and in vitro conservation of indexed plantlets. Plant Cell Tissue Org 109:9–17

    CAS  Article  Google Scholar 

  13. 13.

    Aravind R, Kumar A, Eapen SJ, Ramana KV (2009) Endophytic bacterial flora in root and stem tissues of black pepper (Piper nigrum L.) genotype, isolation, identification and evaluation against Phytophthora capsici. Lett Appl Microbiol 48:58–64

    CAS  Article  Google Scholar 

  14. 14.

    Gram C (1884) Ueber die isolirte Farbung der Schizomyceten in Schnitt-und Trockenpraparaten. Fortschritte der Medicine 2:185–189

    Google Scholar 

  15. 15.

    Graham PH, Parker CA (1964) Diagnostic features in the characterization of the root-nodule bacteria of legumes. Plant Soil 20:383–396

    Article  Google Scholar 

  16. 16.

    Dubey RC, Maheshwari DK (2012) Practical microbiology. S. Chand and Company Ltd, New Delhi

    Google Scholar 

  17. 17.

    Adedayo O, Javadpour S, Taylor C, Anderson WA, Moo-Young M (2004) Decolourization and detoxification of methyl red by aerobic bacteria from a wastewater treatment plant. World J Microbiol Biotechnol 20:545–550

    CAS  Article  Google Scholar 

  18. 18.

    Gordon SA, Weber RP (1951) Colorimetric estimation of indoleacetic acid. Plant Physiol 26:192

    CAS  Article  Google Scholar 

  19. 19.

    Cappuccino JC, Sherman N (1992) Negative staining. In: Cappuccino JC, Sherman N (eds) Microbiology: a laboratory manual. Redwood City, CA, Benjamin, Cummings, pp 125–179

  20. 20.

    Lorck H (1948) Production of hydrocyanic acid by bacteria. Physiol Plant 1:142–146

    CAS  Article  Google Scholar 

  21. 21.

    O’Brien M, Colwell R (1988) Characterization tests for numerical taxonomy studies. In: Colwell RR, Grigorova R (eds) Met in microbiology, vol 19. Academic Press, Cambridge, pp 69–104

    Google Scholar 

  22. 22.

    Cappucino JG (1983) Microbiology: a laboratory manual. Addison Wesley Publishing Company, Boston

    Google Scholar 

  23. 23.

    Koser SA (1923) Utilization of the salts of organic acids by the colon-aerogenes group. J Bacteriol 8:493

    CAS  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Lindstrom K, Lehtomaki S (1988) Metabolic properties, maximum growth temperature and phage sensitivity of Rhizobium sp. (Galega) compared with other fast-growing rhizobia. FEMS Microbiol Lett 50:277–287

    Article  Google Scholar 

  25. 25.

    Kovacs N (1956) Identification of Pseudomonas pyocyanea by the oxidase reaction. Nature 178:703

    CAS  Article  Google Scholar 

  26. 26.

    Clarke PH, Cowman ST (1952) Biochemical methods for bacteriology. Microbiology 6:187–197

    CAS  Google Scholar 

  27. 27.

    El Idrissi MM, Aujjar N, Belabed A, Dessaux Y, Filali-Maltouf A (1996) Characterization of Rhizobia isolated from Carob tree (Ceratonia siliqua). J Appl Bacteriol 80:165–173

    Article  Google Scholar 

  28. 28.

    Pelczar MJ, Reid RD (1965) Microbiology. McGraw-Hill, New York

    Google Scholar 

  29. 29.

    Kersters K, Vancanneyt M (2005) Bergey’s manual of systematic bacteriology. Springer, Berlin

    Google Scholar 

  30. 30.

    Nejad P, Johnson PA (2000) Endophytic bacteria induce growth promotion and wilt disease suppression in oilseed rape and tomato. Biol Control 18:208–215

    Article  Google Scholar 

  31. 31.

    Rijavec T, Lapanje A (2016) Hydrogen cyanide in the rhizosphere: not suppressing plant pathogens, but rather regulating availability of phosphate. Front Microbiol 7:1785.

    Article  PubMed  PubMed Central  Google Scholar 

  32. 32.

    Shoebitz M, Ribaudo CM, Pardo MA, Cantore ML, Ciampi L, Cura JA (2009) Plant growth promoting properties of a strain of Enterobacter ludwigii isolated from Lolium perenne rhizosphere. Soil Biol Biochem 41:1768–1774

    CAS  Article  Google Scholar 

  33. 33.

    Singh RP (2013) Isolation and characterization of multifarious plant growth promoting bacteria Enterobacter ludwigii PGP 19 isolated from pearl millet. Int J Sci Res 4:261–265

    Google Scholar 

  34. 34.

    Vardhan S, Yadav AK, Pandey AK, Arora DK (2013) Diversity analysis of biocontrol Bacillus isolated from rhizospheric soil of rice–wheat (Oryza sativaTriticum aestivum L.) at India. J Antibiot 66:485

    CAS  Article  Google Scholar 

  35. 35.

    Zheng XW, Yan Z, Nout R, Boekhout T, Han B, Zwietering MH, Smid EJ (2015) Characterization of the microbial community in different types of Daqu samples as revealed by 16S rRNA and 26S rRNA gene clone libraries. World J Microbiol Biotechnol 31:199–208

    CAS  Article  Google Scholar 

  36. 36.

    Pham VHT, Kim J (2012) Cultivation of unculturable soil bacteria. Trends Biotechnol 30:475–484

    CAS  Article  Google Scholar 

Download references


One of the authors wishes to acknowledge the University Grants Commission for non-SAP Fellowship. The authors also acknowledge the Director, Dayalbagh Educational Institute, for the financial support to carry out this work.

Author information



Corresponding author

Correspondence to Prem Kumar Dantu.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest to publish this manuscript.

Additional information

Significance Statement

The endophytic bacteria of Piper longum have plant growth-promoting and biocontrol properties. Further studies in this direction would unravel their mode of transmission, co-evolution and bioprospecting for economically important compounds.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Mintoo, M.N., Mishra, S. & Dantu, P.K. Isolation and Characterization of Endophytic Bacteria from Piper longum. Proc. Natl. Acad. Sci., India, Sect. B Biol. Sci. 89, 1447–1454 (2019).

Download citation


  • Piper longum
  • Endophytic bacteria
  • Medicinal plant
  • 16S rRNA sequencing
  • Microbial diversity