Isolation and Characterization of Endophytic Bacteria from Piper longum

  • Mubashar Nazir Mintoo
  • Sushma Mishra
  • Prem Kumar DantuEmail author
Research Article


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.


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



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.

Compliance with Ethical Standards

Conflict of interest

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


  1. 1.
    Bacon CW, White J (2000) Microbial endophytes. Marcel Dekker Inc, New YorkGoogle 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. CrossRefGoogle Scholar
  3. 3.
    Leifert C, Cassells AC (2001) Microbial hazards in plant tissue and cell cultures. In Vitro Cell Dev Biol Plant 37:133–138CrossRefGoogle 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 321:305–339CrossRefGoogle 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 353:409–417CrossRefGoogle 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 132:469–479CrossRefGoogle 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 58:288–293CrossRefGoogle 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 39:441–445CrossRefGoogle 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):214–216CrossRefGoogle Scholar
  10. 10.
    Brader G, Compant S, Mitter B, Trognitz F, Sessitsch A (2014) Metabolic potential of endophytic bacteria. Curr Opin Biotechnol 27:30–37CrossRefGoogle 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. CrossRefGoogle 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 109:9–17CrossRefGoogle 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 48:58–64CrossRefGoogle Scholar
  14. 14.
    Gram C (1884) Ueber die isolirte Farbung der Schizomyceten in Schnitt-und Trockenpraparaten. Fortschritte der Medicine 2:185–189Google Scholar
  15. 15.
    Graham PH, Parker CA (1964) Diagnostic features in the characterization of the root-nodule bacteria of legumes. Plant Soil 20:383–396CrossRefGoogle Scholar
  16. 16.
    Dubey RC, Maheshwari DK (2012) Practical microbiology. S. Chand and Company Ltd, New DelhiGoogle 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 20:545–550CrossRefGoogle Scholar
  18. 18.
    Gordon SA, Weber RP (1951) Colorimetric estimation of indoleacetic acid. Plant Physiol 26:192CrossRefGoogle 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 125–179Google Scholar
  20. 20.
    Lorck H (1948) Production of hydrocyanic acid by bacteria. Physiol Plant 1:142–146CrossRefGoogle 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 69–104Google Scholar
  22. 22.
    Cappucino JG (1983) Microbiology: a laboratory manual. Addison Wesley Publishing Company, BostonGoogle Scholar
  23. 23.
    Koser SA (1923) Utilization of the salts of organic acids by the colon-aerogenes group. J Bacteriol 8:493Google 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 50:277–287CrossRefGoogle Scholar
  25. 25.
    Kovacs N (1956) Identification of Pseudomonas pyocyanea by the oxidase reaction. Nature 178:703CrossRefGoogle Scholar
  26. 26.
    Clarke PH, Cowman ST (1952) Biochemical methods for bacteriology. Microbiology 6:187–197Google 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 80:165–173CrossRefGoogle Scholar
  28. 28.
    Pelczar MJ, Reid RD (1965) Microbiology. McGraw-Hill, New YorkGoogle Scholar
  29. 29.
    Kersters K, Vancanneyt M (2005) Bergey’s manual of systematic bacteriology. Springer, BerlinGoogle Scholar
  30. 30.
    Nejad P, Johnson PA (2000) Endophytic bacteria induce growth promotion and wilt disease suppression in oilseed rape and tomato. Biol Control 18:208–215CrossRefGoogle 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. CrossRefGoogle 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 41:1768–1774CrossRefGoogle 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 4:261–265Google 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 66:485CrossRefGoogle 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 31:199–208CrossRefGoogle Scholar
  36. 36.
    Pham VHT, Kim J (2012) Cultivation of unculturable soil bacteria. Trends Biotechnol 30:475–484CrossRefGoogle Scholar

Copyright information

© The National Academy of Sciences, India 2019

Authors and Affiliations

  • Mubashar Nazir Mintoo
    • 1
  • Sushma Mishra
    • 1
  • Prem Kumar Dantu
    • 1
    Email author
  1. 1.Plant Biotechnology Lab, Department of BotanyDayalbagh Educational InstituteDayalbagh, AgraIndia

Personalised recommendations