Journal of Threatened
Taxa | www.threatenedtaxa.org | 26 September 2025 | 17(9): 27488–27495
ISSN 0974-7907 (Online) | ISSN 0974-7893 (Print)
https://doi.org/10.11609/jott.9625.17.9.27488-27495
#9625 | Received 16 January 2025 | Final received 21 August 2025 |
Finally accepted 03 September 2025
Leaf architecture of threatened Aquilaria cumingiana
(Decne.) Ridley and Aquilaria
malaccensis Lam. (Thymelaeales:
Thymelaeaceae) using morphometrics analysis
Rhea Lou R. Germo
1, Christian C. Estrologo 2 & Gindol Rey A. Limbaro 3
1 Forestry Department, College of
Agriculture and Related Sciences, University of Southeastern Philippines Tagum
– Mabini, Mabini Unit 8807, Davao de Oro, Philippines.
2,3 College of Forestry and
Environmental Studies, Mindanao State University-Maguindanao, Dalican, Datu Odin Sinsuat 9601, Maguindanao del Norte, Philippines.
1 rlrgermo@usep.edu.ph, 2 ccestrologo@msumaguindanao.edu.ph,
3 galimbaro@msumaguindanao.edu.ph (corresponding author),
Editor: A.J. Solomon Raju,
Andhra University, Visakhapatnam, India. Date of publication: 26 September 2025 (online & print)
Citation: Germo, R.L.R., C.C. Estrologo
& G.R.A. Limbaro (2025). Leaf
architecture of threatened Aquilaria cumingiana (Decne.) Ridley
and Aquilaria malaccensis
Lam. (Thymelaeales: Thymelaeaceae)
using morphometrics analysis. Journal of
Threatened Taxa 17(9):
27488–27495. https://doi.org/10.11609/jott.9625.17.9.27488-27495
Copyright: © Germo et al. 2025. Creative Commons Attribution 4.0
International License. JoTT allows unrestricted use,
reproduction, and distribution of this article in any medium by providing
adequate credit to the author(s) and the source of publication.
Funding: None.
Competing interests: The authors declare no competing interests.
Author details: Rhea Lou R. Germo is affiliated with the University of Southeastern Philippines-Mabini Campus, Davao de Oro, Philippines. A PhD student in Forest Biological Sciences at University of the Philippines Los Baños. Her research focus in forest biology and resource conservation. Christian C. Estrologo is affiliated with Mindanao State University-Maguindanao, Maguindanao del Norte, Philippines. A PhD student in Forest Biological Sciences at University of the Philippines Los Baños. His research focuses on Forest Genetics and Natural Resources Conservation and Management. Gindol Rey A. Limbaro is a licensed forester and affiliated with Mindanao State University-Maguindanao, Maguindanao del Norte, Philippines. A PhD student in Forest Industry Engineering at Kastamonu University, Türkiye. His studies primarily focus on Dendrology, Conservation Biology, and Sustainable Forest Products Utilization..
Author contributions: RLRG—research design, paper conceptualization, data collection, data analysis, writing and editing the manuscript. CCE— paper conceptualization, data analysis and writing the manuscript. GRAL—research design, paper conceptualization, data analysis, writing and editing the manuscript, and corresponding journal submission.
Acknowledgments: The authors acknowledge Dr. Nympha Branzuela the provider of Aquilaria species material for the various measurements in order to identify the different variations of the two genera of Aquilaria. The affiliated institutions: University of Southeastern Philippines Tagum-Mabini campus, and Mindanao State University, Maguindanao campus, are acknowledged for providing an avenue to do the research. The authors also acknowledge the emotional support from family, friends, and above all, the Almighty God for the strength and motivation to pursue this study.
Abstract: Due to a very limited number of
scientific studies on the morphology of the very closely related threatened
species, Aquilaria cumingiana
and Aquilaria malaccensis,
it is very challenging to identify them thoroughly. The leaf architecture was
studied in A. cumingiana and A. malaccensis of the family Thymelaeaceae.
Quantitative and descriptive methods were used to assess 21 leaf-traits of A.
cumingiana and A. malaccensis. The study indicated that 10 leaf traits, such
as base shape, apex shape, secondary vein spacing, tertiary vein angle
category, tertiary vein angle to primary, quaternary vein, venation pattern,
laminar shape, base angle, and apex angle, are important for identifying, and
distinguishing the leaf architecture of A. cumingiana,
and A. malaccensis. This study highlights the
importance of leaf morphology and venation patterns in identifying and
differentiating A. cumingiana and A. malaccensis.
Keywords: Dendrogram, leaf apex, leaf
base, leaf blade, leaf morphology, leaf shape, leaf venation, trichomes.
INTRODUCTION
Agarwood-producing species,
specifically the Aquilaria spp. in the Thymelaeaceae family, are primarily distributed in the
Asian region (Li et al. 2023; Xie et al. 2024; Bora
et al. 2025). The Aquilaria genus has 21
species, of which 13 species are reported to be agarwood producers (Lee &
Mohamed 2016; Xie et al. 2024). They produce agarwood
in their trunks and primary branches due to wounding by worms, lightning or
wind-broken branches, natural microbial or fungal infections, or infections
that are artificially induced by drilling holes, cutting the bark, and injecting
chemicals (Jim 2015; Azren et al. 2019; Wang et al.
2020).
The infection court of the fungal
infection of Aquilaria spp. is in the
heartwood, where Aquilaria spp. would generate
a high commercial value (Zhang et al. 2024). The increase in levels of trade
over the past decade has resulted in overexploitation throughout the range of
this species (Chowdhury et al. 2024; Xie et al.
2024). Despite the challenges, such as illegal harvesting in the wild, it is
difficult to cultivate A. malaccensis due to
its sensitivity index in terms of survival rate and environmental conditions
where this species is compatible (Kharnaior &
Thomas 2021; Latifah et al. 2024).
Aquilaria cumingiana and A. malaccensis
are two closely related species of the family Thymelaeaceae.
Globally, A. malaccensis was categorized as
‘Critically Endangered’ while A. cumingiana
was categorized as ‘Vulnerable’ in the IUCN Red List (Harvey-Brown 2018). These
Aquilaria sp. are considered as a
problematic species in terms of species identification due to lack of
scientific studies on species identification. Using leaf architecture is one
way of baseline identification of the species (Mercado et al. 2024). Leaf
architecture refers to the form and position of elements in leaf structure,
including venation pattern, marginal configuration, and leaf shape. Maulia & Susandarini (2019)
reported that venation patterns show significant differences in leaf
architecture that distinguish the closely related species of Aquilaria.
In the present study, the leaf
architecture in Aquilaria cumingiana and A. malaccensis
was examined. This study aimed to evaluate the role of leaf architecture in
species identification of A. cumingiana and A.
malaccensis growing in Mindanao areas. To date, there is no published report on the
characterization of leaf architecture of A. cumingiana
and A. malaccensis as useful taxonomic
evidence, especially for species identification.
MATERIALS AND METHODS
Study area: Samples of plant materials were
obtained from two provinces in Mindanao, Philippines. A. cumingiana
leaf samples were collected from Davao Oriental, while A. malaccensis leaf samples were collected from Agusan del Sur (Image 1). These two species were later
propagated in a backyard nursery situated in Makar, Baliok,
Toril, Davao City, Davao del Sur, Philippines (Figure
1). Laboratory analysis of collected leaf material was performed at the
Forestry Laboratory of the University of Mindanao, Matina
Campus, Davao City, Davao del Sur, Philippines (Image 2). Data were analyzed on
01 August 2022.
Material collection
Materials used in this study were
leaves from seedlings of A. cumingiana and A.
malaccensis collected from the two provinces,
Davao Oriental, and Agusan del Sur. There were 30
juvenile leaves of each species of A. cumingiana and
A. malaccensis collected for the
statistical data analysis. Some of the leaves was added to the herbaria
collection for the taxonomical evidence. The leaves of each species were
collected from different provenances. Foresters and a local parataxonomist
confirmed the identification of tree species. The herbaria were deposited in
the Department of Forestry of the University of Southeastern Philippines –
Mabini Campus.
Leaf architecture traits
There were 21 leaf architectural
traits employed in this study, covering both general morphological traits and
detailed venation features. Traits such as base shape, apex shape, laminar
shape, and angles (base and apex) describe the overall form of the leaf, while
traits like tooth apex, lobation, marginal
development, and leaf margin account for edge modifications. Venation–related
traits, including primary to quaternary vein categories, vein spacing, and
venation pattern, provide critical information on vascular architecture, which
is highly diagnostic in distinguishing species. Additionally, the areole and
laminar blade contribute to identifying structural variations at finer scales.
These traits follow the standardized classification of leaf architecture
proposed by Hickey (1973) and further refined in the Manual of Leaf
Architecture by Ellis et al. (2009).
Measurement
The leaf architecture data were
recorded based on manual leaf architecture (Table 1) with modifications and
several additional traits developed by the Smithsonian Institution (1999). The general morphological traits (laminar
shape, base, apex, margin, lobation, leaf size, and
area) of A. cumingiana and A. malaccensis
were measured using ruler, calipers, and image analysis (Hickey 1979).
Venation traits were examined under compound OptiLab
microscope camera for digital image capturing.
Analysis: Evaluating the leaf architecture
in A. cumingiana and A. malaccensis
was analyzed to cluster analysis using the PAST (Paleontological Statistics)
software version 3.23 to determine the hierarchical relationships among the
different species variations.
RESULTS AND DISCUSSION
Leaf architecture of Aquilaria cumingiana
Leaves of A. cumingiana were alternate and simple in terms of leaf
attachments (Image 3a). Laminar shape was lanceolate, with laminar size varying
754–5,600 mm (Image 3b). The leaves are symmetrical, glabrous, cuneate, entire,
acute both in leaf shape, base angle, apex shape, and apex angle (Image 3a–e).
The leaf texture was smooth and shiny, light green in colour,
while the leaf margin was untoothed, and no distinguished lobation
(Image 3). The leaf venation was pinnate, weak in primary vein size, regular
polygonal reticulate, vein spacing increasing towards the base (Image 3). The
primary venation is straight to slightly curved (Image 3f–g,i), the secondary venation is festooned semi-craspedodromous, secondary vein angle uniform (Image 3g),
and the tertiary venation is opposite percurrent (Image 3h–m). The areolation and the quaternary venation
were not observed. The marginal development was arranged in a looped formation
(Image 3i). There were variations in
midrib width, marginal vein width, and the blade class. Trichomes in the laminar area were observed,
but strong evidence is required (Image 3i–j).
Leaf architecture of Aquilaria malaccensis
Aquilaria malaccensis displays its variation in terms
of leaf architecture as compared to A. cumingiana.
The leaves of A. malaccensis were alternate,
simple, lanceolate, symmetrical, acute, obtuse, acuminate, entire, glabrous,
untoothed, and no lobation (Image 4a–e). The venation
characteristics of A. malaccensis are pinnate,
weak, reticulodromous, straight to slightly curved
for the primary vein course, with irregular venation spacing (Image 4f). The
secondary vein category is semi-craspedodromous, the
tertiary vein is categorized as random, while the quaternary vein is
dichotomizing (Image 4). The areolation was not observed, while the marginal
development was looped (Image 4h–k). There was a notable occurrence of
trichomes in the below leaf surface (Image 4i). This result has a
similarity assessment to the study of Maulia & Susandarini (2019) on the leaf architecture of A. malaccensis.
Variations between Aquilaria cumingiana and
Aquilaria cumingiana
The dendrogram (Figure 1) clearly
distinguishes A. cumingiana from A. malaccensis based on 21 leaf architectural
characteristics, with A. cumingiana forming a
compact cluster that reflects its morphological uniformity, while A. malaccensis displays broader sub-clustering, indicative
of greater intraspecific variation. The correlation (Figure 2) further shows
that only 10 traits strongly influenced this clustering, particularly base
shape, apex shape, and venation-related traits such as secondary, tertiary, and
quaternary vein categories, while other traits like leaf margin, lobation, and tooth apex contributed little to species
identification. These results highlight that venation and lamina form are the
most reliable diagnostic features for separating the two Aquilaria
sp.
Summary of key findings
The comparative study of leaf
architecture in A. cumingiana and A. malaccensis is important for their morphological and
taxonomic identification. These species have smooth texture and pinnate
venation that includes festooned semi-craspedodromous
secondary veins, and a symmetrical, and lanceolate lamina. The stable morphological
profile suggested by the invariant features in the sample over different times
could be the result of the adaptation to an ecological niche.
Aquilaria malaccensis has higher leaf variability. The
secondary venation, mostly dichotomous, but there is also random tertiary
venation with possible irregular spacing, arc venation, and other morphological
plasticity, is a testament to its greater morphological plasticity. The
presence of trichomes
under the A. malaccensis leaves
(as opposed to the smooth surface of A. cumingiana)
could also be an adaptation to different environmental pressures.
Cluster analysis of 21 traits of
leaf form revealed clear taxonomic separation between A. cumngiana
and A. malaccensis. From this, it could be
concluded that the variation in A. malaccensis
is driven to a greater extent, suggesting that genetics or environment has a
greater effect on the morphology of these specimens. These findings underscore
the importance of leaf architecture in distinguishing closely related species,
particularly where morphological similarities blur taxonomic boundaries.
The fixed differences were
observed in 10 characteristics, including laminar blade, base angle, and apex
angle between both species. These dissimilarities suggest that these
characteristics could serve as diagnostic markers for taxa identification.
While the fixed nature of other traits reinforces the genealogical relationship
among these species, morphological divergence may result from ecological
divergence but may reflect genetic divergence.
CONCLUSION
This study underscores the relevance
of a comprehensive leaf architectural study toward the identification of
closely related species in the genus Aquilaria.
The study suggests that A. malaccensis is more
morphologically variable compared to A. cumingiana
and is likely to have a broader ecological amplitude or population genetic
diversity. In contrast, the stable morphology observed in A. cumingiana suggests a stable taxonomic relationship
that may be dictated by particular environmental demands. These findings serve
as original data for taxonomic identification and for the conservation and
sustainable management of these economically valuable agarwood-producing
species. These morphological differences should be further explored in terms of
their ecological and genetic basis using more molecular approaches and by
sampling more habitat types in the future. Indeed, exploring the environment
where trichome and venation patterns develop could also help in deciphering the
adaptive strategies of these species.
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REFERENCES
Azren, P. D., S. Y. Lee, D. Emang, & R Mohamed (2019). History and perspectives of
induction technology for agarwood production from cultivated Aquilaria in Asia: a review. Journal of Forestry
Research 30(1): 1–11. https://doi.org/10.1007/s11676-018-0627-4
Bora, S.S.,
R. Ronghang, P. Das, R.S. Naorem,
D.J. Hazarika, R. Gogoi & M. Barooah
(2025). Endophytic
microbial community structure and dynamics influence agarwood formation in Aquilaria malaccensis Lam.
Current Microbiology 82(2): 66. https://doi.org/10.1007/s00284-024-04048-2
Chowdhury,
B.D., A. Bhattacharjee & B. Debnath (2024). Endophytic microbes in agarwood
oil production from Aquilaria malaccensis Lam. Engendering bio-resources for
socioeconomic development, pp. 168–195. In: Advanced Green Technology for
Environmental Sustainability and Circular Economy. CRC Press, 278 pp. https://doi.org/10.1201/9781003517108
Ellis, B.,
D.C. Daly, L.J. Hickey, K.R. Johnson, J.D. Mitchell, P. Wilf & S.L. Wing
(2009). Manual of
Leaf Architecture. Cornell University Press, 201 pp.
Harvey-Brown,
Y. (2018). Aquilaria cumingiana. The IUCN Red List of Threatened
Species 2018: e.T38068A88301841. en.
Accessed on 11.i.2025. https://doi.org/10.2305/IUCN.UK.2018-1.RLTS.T38068A88301841
Harvey-Brown,
Y. (2018). Aquilaria malaccensis. The IUCN Red List of Threatened
Species 2018: e.T32056A2810130. en.
Accessed on 11.i.2025. https://doi.org/10.2305/IUCN.UK.2018-1.RLTS.T32056A2810130
Hickey, L.J.
(1979). A revised
classification of the architecture of dicotyledonous leaves, pp. 25–39. In:
Metcalfe, C.R. & L. Chalk (eds.). Anatomy of the Dicotyledons—Volume
1. Clarendon Press, 800 pp.
Jim, C. Y.
(2015). Cross-border
itinerant poaching of agarwood in Hong Kong’s peri-urban forests. Urban
Forestry & Urban Greening 14(2): 420–431. https://doi.org/10.1016/j.ufug.2015.04.007
Kharnaior, S. & S.C. Thomas (2021). A review of Aquilaria
malaccensis propagation and production of the
secondary metabolite from callus. Grassroots Journal of Natural
Resources 4(4): 85–94. https://doi.org/10.33002/nr2581.6853.040407
Latifah, S.,
A.L. Codilan, O.H. Syahputra,
A. Kustanti, G.R.N.B. Sembiring,
T.C. Ningrum & N.I.M. Daulay
(2024). Study of the
existence of cultivated agarwood plants Aquilaria
malacensis as an effort to preserve the environment
around the forest. In: E3S Web of Conferences, Volume 519, 2014. 5th
Talenta Conference on Engineering, Science and
Technology (TALENTA CEST-5 2024). Article number - 03003. https://doi.org/10.1051/e3sconf/202451903003
Lee, S.Y.
& R. Mohamed (2016). The origin and domestication of Aquilaria,
an important agarwood-producing genus, pp. 1–20. In: Mohamed, R. (ed.). Agarwood:
Science Behind the Fragrance.
Springer, Singapore, 167 pp. https://doi.org/10.1007/978-981-10-0833-7_1
Li, T., Z. Qiu, S.Y. Lee, X. Li, J. Gao, C. Jiang & J. Liu (2023). Biodiversity and application
prospects of fungal endophytes in the agarwood-producing genera, Aquilaria and Gyrinops (Thymelaeaceae): a review. Arabian Journal of Chemistry
16(1): 104435. https://doi.org/10.1016/j.arabjc.2022.104435
Maulia, Z. & R. Susandarini
(2019). Role of
Leaf Architecture for the Identification of agarwood — producing species Aquilaria malaccensis
Lam. and Gyrinops versteegii
(Gilg.) Domke at
Vegetative Stage. Journal of Biological Sciences 19(6): 396–406. https://doi.org/10.3923/jbs.2019.396.406
Mercado,
M.I., M.D.H.S. Matías, C.M. Jimenez, M.S.B. Sampietro, M.A. Sgariglia, J.R. Soberón & D.A. Sampietro
(2024). Comparative
Analysis of Leaf Architecture and Histochemistry in Schinus
fasciculatus and S. gracilipes
(Anacardiaceae). Brazilian Archives of Biology and
Technology 67: e24230088. https://doi.org/10.1590/1678-4324-2024230088
Wang, Z.F.,
H.L. Cao, C.X. Cai & Z.M. Wang (2020). Using genetic markers to identify
the origin of illegally traded agarwood producing Aquilaria
sinensis trees. Global Ecology and
Conservation 22: e00958. https://doi.org/10.1016/j.gecco.2020.e00958
Xie, Z.Q., J.Y. Xu, M. Rafiq & C.S. Cheng (2024). An analysis
of agarwood trade patterns,
historical perspectives, and species
identification challenges: repercussions
for importing nations. TMR Modern Herbal Medicine 7(1): 1–10. https://doi.org/10.53388/MHM2024001
Zhang, X., L.X. Wang, R. Hao,
J.J. Huang, M. Zargar, M.X. Chen & H.F. Dai
(2024). Sesquiterpenoids in agarwood: biosynthesis, microbial
induction, and pharmacological activities. Journal of Agricultural and Food
Chemistry 72(42): 23039–23052. https://doi.org/10.1021/acs.jafc.4c06383