Research Articles

2018  |  Vol: 4(4)  |  Issue: 4 (July- August)  |  https://doi.org/10.31024/ajpp.2018.4.4.10
Isolation and identification of endophytic fungi from Gymnema sylvestre R. Br.

Ambika Vasantakumar, G. M. Vidyasagar*

Medicinal Plants and Microbiology Research Laboratory, Department of P G Studies and Research in Botany, Gulbarga University, Kalaburagi-585106, Karnataka, India.

*Address for Corresponding Author

G. M. Vidyasagar

Medicinal Plants and Microbiology Research Laboratory,Department of P G Studies and Research in Botany, Gulbarga University, Kalaburagi-585106, Karnataka, India.


Abstract

Objective: Endophytes act as major reservoir of biologically active compounds for health care applications and an increasing number of novel bioactive compounds are being extracted from fungal endophytes.so the present study is isolation and identification of endophytic fungi from Gymnema sylvestreMaterial and methods: Fresh leaves and stem were washed thoroughly in running water and with distilled water to remove dust and debris. The material was surface sterilized by mercury chloride.  The leaf segments were placed in petri dishes (2–3 segments/petri dish) containing Potato Dextrose Agar (PDA) medium. The petri dishes were sealed using parafilm and incubated at 26±2 °C in the light chamber for 3 weeks. The fungi grown were transferred to PDA plate and identified based on their morphology and other features. Results: A total of 60 endophytic isolates were isolated from Gymnema sylvestre and among them 10 endophytes belonging to seven genera were identified. The other fungal isolates need further efforts to identify. Conclusion: A total of 10 endophytic fungi belongs to seven genera were isolated and identified from Gymnema sylvestre and they may be very much useful in the extraction of potent active compounds that the plant is known far. The study may be useful in protecting the highly useful medicinal plant by giving the alternate source for drug.

Keywords: Gymnema sylvestre , endophytes


Introduction

Gymnema sylvestre R.Br  is  a  medicinal  herb  native  to  central  and  western  India,  tropical Africa  and Australia.  It  is  often  called  “gurmar”  (destroyer  of  sugar), as  chewing  the leaves causes a  loss of  sweet  taste  (Yeh et al., 2003). G. sylvestre  containing the triterpenoid  gymnemagenin and Gymnemic acid, extracted  from  leaves and  roots is  mainly  used  in  India  and  parts  of  Asia  as  a  natural  treatment  for  diabetes  as  it helps to lower the blood sugar(Duke   et al., 1997). In addition, it possesses antimicrobial, antihyper  cholesterolemic  (Bishayee et al., 1994),  sweet  suppressing  (Kurihara et al., 1992) and  hepatoprotective  (Rana et al., 1992)  activities.  It  also acts  as  feeding  deterrents  to  caterpillar,  prodenia  eridania (Granich  et al., 1974),  prevent  dental  caries  caused by Streptococcus mutans (Hiji  et al., 1990)  and skin cosmetics (Komalavalli  et al., 2000). Besides, it is also used in the treatment of asthma, eye complaints, inflammations, snake bite (Kini et al., 1982). Gymnemagenin, the wonder drug for diabetes (Liu et al., 1992,) and obesity (Kanetkar et al 2007), also possess antiviral properties (Sinsheimer et al., 1971). They are being isolated from the leaves and roots of Gymnema sylvestre, which leads to the destruction of natural resources. Alternatively, gymnemagenin-producing endophytic fungi Penicillium oxalicum  is isolated from Gymnema sylvestre, so the fungus can serve as a potential material for the production of gymnemagenin. As G. sylvestre has been used in the treatment  of several ailments, it is assumed to have several active compounds such as, Betaine, Beta-Carotene, Choline, Gymnemic-Acid, Niacin, Ascorbic-Acid. Hence the present study was carried out to identify the fungal endophytes capable of producing the active compounds reported from G. sylvestre.

Figure 1. Gymnema sylvestre

 

 

 

 

Materials and methods

Collection of the plant material

G. sylvestre collected from Botanical garden of Gulbarga University, Kalaburagi, Karnataka and was authenticated with the help of flora of Gulbarga district (Seetharam et al., 2000). A specimen is deposited in the herbarium, Dept. of Botany, Gulbarga University Kalaburagi (Voucher speciemen No HGUG-58).

Isolation and identification of endophytic fungi

Fresh leaves and stem were washed thoroughly in running water and with distilled water to remove dust and debris. The material was surface sterilized by mercury chloride.  Leaves and stem samples were cut into 2–3 cm segments under aseptic conditions, The leaf segments were placed in petri dishes (2–3 segments/petri dish) containing Potato Dextrose Agar (PDA) medium. The petri dishes were sealed using parafilm and incubated at 26±2 °C in the light chamber for 3 weeks. The plates were monitored every day to check the growth of endophytic fungal colonies on leaf segments. The fungi grown were transferred to PDA plate and identified based on their morphology and other features.

Results and discussion

Isolation and identification of endophytic fungi

In the present study, a total of 60 isolates were isolated from the leaf and stem materials of G. sylvestre. Of which, ten fungal species belonging to seven genera were isolated and identified. Cylindrocladium parvum, Cladosporium variabile, cochliobolus geniculatus were recorded during September 2015, Alternaria brassicicola, Colletotrichum capsici, colletotrichum dematium were recovered during August and Aspergillus fumigates, Aspergillus niger, Fusarium oxysporum and Alternaria alternate were recorded during February month of 2015 (Figure 2).Aspergillus niger and Alternaria alternata  were also reported from Calotropis gigantean (Srimathi et al., 2011) Calotropis procera (Rezwana et al., 2007), Withania somnifera (Rezwana et al., 2010), Azadirachta indica (Tenguria et al., 2011) and Melia azadirachta (Kaushal et al., 2010). Ascomycota forms have been reported as endophytes (Blodgett et al., 2000; Suryanarayanan et al., 1998) and natural products from endophytes have a broad spectrum of biological activity (Zhang et al., 2006).  

Identification of endophytes

Endophytic fungi were identified according to their microscopic and macroscopic characteristics such as the morphology of fruiting structures and spore morphology under a bright-field microscope (10X and 40X) using Motic image microscope. Fungal features have been recorded with the help of a hand book of soil fungi by Nagamani (Nagamani et al., 2006) and illustrated genera of imperfect fungi by Barnet HL and Hunter BB (Barnet et al., 1972).

Table 1. Fungal endophytes isolated from leaf and stem parts of Gymnema sylvestre

S. No.

Name of the endophyte

Plant part used

Growth in diameter

1

Cylindrocladium parvum

Stem

2.5

2

cladosporium variabile

Stem

2

3

cochliobolus geniculatus

Leaf

4.32

4

Alternaria brassicicola

Leaf

3.72

5

Colletotrichum capsici

Leaf

3.9

6

colletotrichum dematium

Stem

2.38

7

Aspergillus fumigates

Stem

4.16

8

Aspergillus niger

Stem

2.03

9

Fusarium oxysporum

Stem

2.06

10

Alternaria alternata

Stem

3.27

Figgure 2. Endophytic fungi isolated from Gymnema sylvestre: (a) Cylindrocladium parvum, (b) Cladosporium variabile, (c) Cochliobolus geniculatus, (d) Alternaria brassicicola, (e) Colletotrichum capsici, (f) Colletotrichum dematium, (g) Aspergillus fumigates, (h) Aspergillus niger, (i) Fusarium oxysporum, (j) Alternaria alternate

 

 

 

 

 

Conclusion

A total of 10 endophytic fungi belongs to seven genera were isolated and identified from Gymnema sylvestre and they may be very much useful in the extraction of potent active compounds that the plant is known far. The study may be useful in protecting the highly useful medicinal plant by giving the alternate source for drug.

Conflicts of interest: Nil

References

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