Download Actinomycetes are fungus-like Gram-positive bacteria having high

Isolation and Screening of Endophytic
Actinomycetes from Herb of Chanthaburi
Province and Their Bioactivities
Winyou Puckdee1,2* and Thongchai Taechowisan1
1
Department of Microbiology, Faculty of Science, Silpakorn University, Nakorn Pathom 73000, Thailand
2
Department of Biology, Faculty of Science and Technology, Rambhai Barni Rajabhat University, Chanthaburi 22000, Thailand
*Corresponding/first author’s e-mail: [email protected]
Introduction :: Actinomycetes are fungus-like Gram-positive bacteria having high
guanine and cytosine (> 55 %) content in their DNA. They are in the order Actinomycetales and are free living and saprophytic bacteria. The majority of Actinomycetes are
a widely distributed group of microorganisms in nature found in soil, water, air and
tissues of animals or plants. The actinomycetes that reside within living plant tissues
A
C
B
Fig. 1 Macroscopic (A and B) and microscopic
(C) characterization of endophytic actinomycete w14.
actinomycetes. They have the capability to produce numerous bioactive compounds such as antibiotics, antioxidant, anti-cancer, immunomodulating agents, pesticides, herbicides and enzymes and are of considerable
importance in medical, agriculture, and the veterinary.
Objectives :: To isolate and screen the active endophytic actinomycetes from herb of Chanthaburi province and
to evaluate crude extract of endophytic actinomycetes.
Methods ::
Isolation of endophytic actinomycetes from the herb of
Chanthaburi province were carried out by surface sterilization technique and
B
C
A
cultured on humic acid-vitamin (HV) agar. Double layer technique, agar Fig. 2 Endophytic actinomycete on humic acid-vitamin
- (HV) agar (A) and double layer technique for antimicrobacterial activity of all isolated actinomycetes. Crude fraction was extracted with bial activity of endophytic actinomycete w14 (B and C).
.
ethyl acetate and then isolated with thin layer chromatography (TLC). And 2,2-diphenyl-1-picrylhydrazyl (DPPH ) scavenging assay was used to determine their anti-oxidant activity.
Results :: Twenty-seven isolates of endophytic actinomycetes were isolated
Fig. 3 Crude extracte of endophytic actinomycete w14.
Fig. 5 Scavenging activity
(DPPH.) of w14 crude
extracte.
from Amomum uliginosum K. D. Koenig (Reo-hom), Amomum testaceum Ridl
(Kra-waan) and Zingiber zerumbet (L) Smith (Kra-thue). Five isolates (w03, w08,
w13, w14 and mw07) were active against 7 species of tested microorganisms and Fig. 4 Broth microdilumaximum inhibition index was found in w14 against Staphylococcus aureus and tion method.
Bacillus cereus at 12.12 ± 1.14 and 9.5 ± 0.44, w08 at 6.30 ± 0.39 and 5.89 ± 0.20, respectively.
Therefore endophytic actinomycetes code w14 was used to extraction with ethyl acetate. The
crude extract was dark brown weigh 1.13 g in 3 times of extraction of 1.5 L of culture. The minimum
inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against S. aureus,
B. cereus and B. subtilis
fractions were isolated by TLC with acetone and hexane at 2 : 3 ratio and antibacterial against
S. aureus, B. cereus and B. subtilis were tested by bioautography technique have retardation factor
.
(Rf) at 0.67. The 50% DPPH
and brown spore production and microscopic
characterized by slide culture show the small
with continues chain and coil. Isolation of bioactive
compounds in the crude extract should be studied
further.
A
B
C
Fig. 6 Bioautography technique for antibacterial activity of
endophytic actinomycete w14 crude extract against B. subtilis (A),
B. cereus (B) and S. aureus (C).
Conclusion :: Twenty-seven isolates of endophytic actinomycetes were isolated from herb of
Chanthaburi province. Five isolates have antibacterial activity and maximum inhibition index was
.
found in w14. Ethyl acetate crude extract of w14 was dark brown with antibacterial and DPPH
scavenging activities.
Reference :: Taechowisan T, Peberdy J & Lumyong S (2003) Acknowladgements ::
This work was supported by Higher
Isolation of endophytic actinomycetes from selected plants Education Research Promotion (HERP), National Research Univerand their antifungal activity. World Journal of Microbiology and
Biotechnology 19: 381-385.
and the Faculty of Science, Silpakorn University, Nakorn Pathom,
Thailand.