Research field

1. Microscopic
observation of in
vitro Aquilaria
malaccensis after
wounding treatment
Subasini Anamulai
168246
Forest Production,Faculty of
Forestry, UPM
Supervisor
Assoc. Prof. Dr. Rozi Mohamed

2. Aquilaria malaccensis is a species of plant in the
Thymelaeaceae family.
It is found in Bangladesh, Bhutan, India,
Indonesia, Laos, Malaysia, Myanmar, the
Philippines, Singapore, and Thailand. It is
threatened by habitat loss (Angela, 2000)
Aquilaria malaccensis is the major source of
agarwood, a resinous heartwood, used for perfume
and incense (Chang, 2005)
Introduction

3. Problem and justification
 Previous study showed that there was no
differences in anatomical features between
juvenile and matured trees after wounding
treatment in natural environment led to gaharu
induction.
 In this study, anatomical characteristics of in vitro
plants will be investigated
 So, this experiment to find the anatomical
changes of in vitro of Aquilaria malaccensis under
microscopic observation

4. To characterize anatomical features of
in vitro Aquilaria malaccensis under
microscopic observation
To determine any changes in the
anatomical features after wounding
treatment
Objectives

5. • Samples of in vitro plantlets
• Fungal inoculum – Fusarium solani
• Chemical elicitor – Methyl jasmonate (MeJa)
• Epoxy embedding kit
Epoxy Embedding kit, contains DDSA,NMA, DMP -30
Materials and method

6. Wounding treatment
Wounding+ fungus
wounding + chemical elicitor
Non-wounding + fungus
non-wounding + chemical elicitor
Wounded 24 hours
Embedding (Epoxy Embedding)
Sectioning
Microscopic study

7. In vitro plantlets after treatments for 2
months.
The anatomical were observed under light
microscopy
Samples are embedded
using Epoxy
Results

8. CP
VC
Controlled Aquilaria malaccensis (Thymelaceae) plantlets. PF= Phloem
Fibre, P= Phloem, C=Cortex, VC= Vascular Cambium, Scale =100µm.
C
PX

9. cp
x
ph
hb
ca
c
scl
Traverse section of stem of plantlets with F.solani inoculation and
wounding treatment showing the formation of haustorium bridge.
Abbreviations: c=cortex,ph=phloem,hb= haustarium bridge,
sph=secondary phloem, x=xylem, ca=vascular cambium. Scale
=100µm
Ip

10. P
sx
hb
sph
c
ph
Traverse section of stem of plantlets with F.solani inoculation and non
wounding treatment showing the formation of haustorium bridge.
Abbreviations: c=cortex,ph=phloem,hb= haustarium bridge,
sph=secondary phloem, sx= secondry xylem, p=pith, ip=included
phloem, Scale =100µm
Ip

11. Showing the extended parts of plantlets with F.solani inoculation and non wounding
treatment. Scale =100µm
Ep

12. p
sx
sp
cp
Ip
TS of plantlets treated with MeJa and non wounded. Ip=
Included phloem, sp= secondry phloem. Sx=secondry xylem,
p=pith, cp=

13. Lenticels
Deposit in
pores
Showing the lenticels and deposit in pore under
scale=100µm

14. Ip
TS of plantlets treated with MeJa and wounded. Shows
the changes in distribution of phloem compare to
controlled and the higher number of deposit in pores Ip=
Included phloem. Scale =100µm

15. Ip
TS of plantlets treated with MeJa and wounded. The
distribution of included phloem. Scale =30µm

16. SX
P
VC
RP
pe
TS of plantlets wounded for 24 hours. Pe= Pholem elements,
rp=ray parencyhma, sx=secondry xylem, p=pith, vc=vascular
cambium. Scale =100µm

17. Ip
Ts of wounded 24 hours plantlets included phloem.
Scale =30µm

18. • It shows, when Tissue culture of A.malaccensis interact with
MeJa as a chemical elicitor, the deposit in pores become high
and showing that potential to induce production fragnant
compound.
• The results shows similarly with study by Ito in wild Aquilaria
sp. (Ito et al. 200)
• There is haustorium bridge development in xylem that
extended to new growth in plants tissue, assumed that the
tissue culture’s defense of these agarwood producing species
is triggerd by F. solani attack, as Turjaman,2011 mention the
same statement in his journal.
• Treatments done chemically, fungus inoculation and
technically, the reaction of the tissues occur in chemical
treatment and fungus inoculation are faster.
Discussion

19. The anatomical changes in in vitro A.malaccensis
can be seen clearly in xylem and phloem tissues
after undergo treatments chemically, fungus
inoculation and technically.
Methyl Jasmonate can be potential chemical
solution that can trigger the induction of
agarwood.
 F. solani induce the reaction by enchance the
plantlets defense and change the tissues shape
and structure.
Conclusion

20. • Rahman, M. A., & Khisa, S. K. (1984). Agar production in
agar tree by artificial inoculation and wounding II. Bano
Biggyan Patrika, 13,57-63.
• Yaacob, S. (1999). Agarwood: Trade and CITES
Implementation in Malaysia. Unpublished report prepared
for TRAFFIC Southeast Asia, Malaysia
• Mohamed, R.1*, Wong, M. T.1 and Halis, R.2 1Forest
Biotech Laboratory, Department of Forest Management,
Faculty of Forestry, Universiti Putra Malaysia, 43400
Serdang, Selangor, Malaysia
References

21. Thank you