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Wood identification of Dalbergia nigra (CITES Appendix I) using quantitative
wood anatomy, principal components analysis and naı
¨ve Bayes classification
Peter Gasson1,*, Regis Miller2, Dov J. Stekel3, Frances Whinder1and Kasia Ziemin
´ska1
1
Jodrell Laboratory, Royal Botanic Gardens Kew, Kew Road, Richmond, Surrey TW9 3DS, UK,
2
Forest Products Laboratory,
One Gifford Pinchot Drive, Madison, WI 53726-2398, USA and
3
School of Biosciences, University of Nottingham, Sutton
Bonington Campus, Loughborough, Leics LE12 5RD, UK
Received: 17 August 2009 Returned for revision: 14 September 2009 Accepted: 5 October 2009 Published electronically: 2 November 2009
†Background and Aims Dalbergia nigra is one of the most valuable timber species of its genus, having been
traded for over 300 years. Due to over-exploitation it is facing extinction and trade has been banned under
CITES Appendix I since 1992. Current methods, primarily comparative wood anatomy, are inadequate for con-
clusive species identification. This study aims to find a set of anatomical characters that distinguish the wood of
D. nigra from other commercially important species of Dalbergia from Latin America.
†Methods Qualitative and quantitative wood anatomy, principal components analysis and naı
¨ve Bayes classifi-
cation were conducted on 43 specimens of Dalbergia, eight D. nigra and 35 from six other Latin American
species.
†Key Results Dalbergia cearensis and D. miscolobium can be distinguished from D. nigra on the basis of vessel
frequency for the former, and ray frequency for the latter. Principal components analysis was unable to provide
any further basis for separating the species. Naı
¨ve Bayes classification using the four characters: minimum vessel
diameter; frequency of solitary vessels; mean ray width; and frequency of axially fused rays, classified all eight
D. nigra correctly with no false negatives, but there was a false positive rate of 36.36 %.
†Conclusions Wood anatomy alone cannot distinguish D. nigra from all other commercially important Dalbergia
species likely to be encountered by customs officials, but can be used to reduce the number of specimens that
would need further study.
Key words: Dalbergia nigra, Brazilian rosewood, CITES, wood anatomy, PCA, naı
¨ve Bayes analysis.
INTRODUCTION
Dalbergia is a pantropical genus belonging to the
Leguminosae– Papilionoideae and comprises approx. 250
species (Klitgaard and Lavin, 2005). Of these, around 10– 15
species (Record and Hess, 1943; Richter et al., 1996) are
known for the beauty and quality of their timber, making
them of particular economic importance.
The Brazilian species Dalbergia nigra, commonly known as
Brazilian rosewood, has been traded for over 300 years
(Record and Hess, 1943). It grows in the Atlantic forest, pri-
marily from southern Bahia to northern Espı
´rito Santo, but
also on the coast of Sa
˜o Paulo state and inland in Minas
Gerais (Carvalho, 1989, 1997). Specimens in the coastal
Atlantic forest may reach approx. 125 ft (38 m) in height
(Record and Hess, 1943), whereas inland specimens are
shorter (Carvalho, 1989). The heartwood of D. nigra is slow-
forming, ranging in colour from brown to purplish-black, and
is durable, resisting fungal and insect attack (Flynn and
Holder, 2001; Wiedenhoeft and Miller, 2005). Due to the strik-
ing colour and grain pattern, early explorers brought the wood
back to Spain and Portugal where it fast became a most desir-
able wood. Its timber has been used in the manufacture of fur-
niture and cabinets and is also highly prized for musical
instruments, particularly guitars, as it is believed to have
superior acoustic qualities to other woods (Flynn and Holder,
2001).
Because of its popularity, D. nigra has been overexploited
and has been listed as ‘vulnerable’ on the Red List of the
International Union for the Conservation of Nature (IUCN)
since 1998 (Varty, 1998), meaning it is considered at a high
risk of extinction in the medium-term if efforts are not made
to conserve it. In 1992 it was listed on Appendix I of the
Convention on International Trade in Endangered Species of
Wild Fauna and Flora (CITES). All international trade is pro-
hibited, except for the purposes of scientific research.
CITES legislation makes it necessary to be able to dis-
tinguish the wood of D. nigra from species that are not pro-
tected by trade laws. Experienced wood anatomists recognize
a specimen as belonging to the genus Dalbergia by the pres-
ence of short, regularly storied rays, typically large vessels,
abundance of diffuse and diffuse-in-aggregate axial parench-
yma, and vestured pits. North Carolina State University main-
tains an online database for the identification of hardwoods,
InsideWood (2004 onwards), which uses the International
Association of Wood Anatomists (IAWA) List of
Microscopic Features for Hardwood Identification (IAWA
Committee, 1989) to provide descriptions of the wood
anatomy of over 5700 specimens. Of these, the characters
‘all rays storied’, ‘diffuse-in-aggregate axial parenchyma’
and ‘vestured pits’ are listed in only 38 records. Careful com-
parison with reference material is usually enough to make a
* For correspondence. E-mail P.Gasson@kew.org
#The Author 2009. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved.
For Permissions, please email: journals.permissions@oxfordjournals.org
Annals of Botany 105: 45– 56, 2010
doi:10.1093/aob/mcp270, available online at www.aob.oxfordjournals.org
distinction to genus level. However, for CITES legislation to
be implemented, the identification must be to species level,
and this is much more difficult.
The Brazilian species D. spruceana is not protected by trade
laws and is often confused with D. nigra (Kribs, 1959; Miller
and Wiemann, 2006, and other sources). Although InsideWood
(2004 onwards) suggests a possible separation between these
two species, the features showing differences are variable or
not sufficient to make an accurate separation.
Currently, there aretwo resources relating to the identification
of D. nigra for CITES purposes. The first is the CITES
Identification Guide – Tropical Woods (CITES, 2002), which
provides a general key to identifying CITES species, together
with low magnification photographs of the transverse surface
of D. nigra and similar species. This book is aimed primarily
at non-experts such as customs inspectors to allow most
species to pass through inspection. For Dalbergia species,
inspectors need to send wood samples to experts for positive
identification. The second is the more recent CITESwoodID
CD-ROM (Richter et al., 2008), which includes an interactive
key, providing more detail than the printed guide. However, it
is still not possible to distinguish D. nigra from D. spruceana
because recorded differences between the species are variable;
for example, D. spruceana does not have distinct growth rings,
but D. nigra may have distinct growth rings, or they may be
indistinct or absent. Another potential resource for identification
is the InsideWood database (InsideWood 2004 onwards), but
whereas it can be readily used to identify Dalbergia, the discri-
minatory power of the individual character states is insufficient
for species identification.
Miller and Wiemann (2006) developed a system for separ-
ating heartwood specimens of D. nigra and D. spruceana
based on measurements of density and water and ethanol flu-
orescence of extracts. However, the results only apply if the
specimen being examined is definitely one of these two
species from Brazil, and it is not always appropriate to make
this assumption. Guzman et al. (2008) applied this technique
to three Mexican species including D. granadillo and
D. stevensonii, but the latter ovelaps with D. nigra in the flu-
orescence of its ethanol extract. Future studies using non-
anatomical features including all commercial Latin American
species are planned.
Therefore, it is clear that, if CITES regulations are to be
implemented successfully, a more conclusive method for dis-
tinguishing D. nigra from other Dalbergia species is needed.
One way to investigate subtle wood anatomical differences
between species is to collect quantitative data relating to key
characteristics, and to subject these data to multivariate statisti-
cal classification. Broadly, such techniques fall into two cat-
egories: unsupervised and supervised. Unsupervised
classification techniques, such as cluster analysis and principal
components analysis (PCA) take the measurements associated
with a set of specimens and look for clusters of similarity
without taking into account any pre-existing categorization
of the specimens. Supervised classification techniques, such
as naı
¨ve Bayes classification or artificial neural networks
make use of categories to which (some of) the specimens
belong. These specimens are known as the training set. The
aim is to identify features in the measurements of the training
set that accurately predict the categories of the training set, so
that these features can be used to predict the category of future
specimens for which the category is unknown.
Angyalossy-Alfonso and Miller (2002) used average linkage
cluster analysis to investigate 51 species and varieties of
Swartzia together with eight other genera in the tribe
Swartzieae. The species and varieties of Swartzia became
four groups, whilst the tribe Swartzieae became seven
groups. Canonical discriminant analysis showed that the char-
acters – storied structure, number of tiers per millimetre, axial
parenchyma strand length, ray height, intervascular pit size
and exclusively uniseriate rays – could be used to discriminate
between the groups.
Hellberg and Carcaillet (2003) used PCA to investigate car-
bonized samples of four Betula species from Western Europe:
B. nana, a shrub, and B. pendula,B. pubescens and
B. pubescens ssp. tortuosa, which are all trees. Their PCAs
placed B. nana in a cluster distinct from the three tree
species, with size of vessel groups, mean vessel abundance
and ray density being the key distinguishing characters. PCA
was not able to fully distinguish the tree species from each
other, although a further PCA of only B. pendula and
B. pubescens did give separation on the first PC, with the
key characters being; vessel diameter, ray density, ray width
and ray height.
MacLachlan and Gasson (2010) applied PCA to 21 speci-
mens from seven species in the genus Pterocarpus, with the
aim of distinguishing P. santalinus (CITES Appendix II),
from closely related taxa. Specimens of P. santalinus formed
a cluster that was discrete from the six other species studied,
with the key characters being specific gravity, fibre wall thick-
ness, parenchyma band width, and vessel density/mean tan-
gential vessel diameter. Two further species also formed
discrete clusters, and the authors concluded that when used
in conjunction with comparative anatomy, PCA has the poten-
tial to be a useful tool in identifying CITES species.
The use of supervised classification techniques in wood
identification has been largely confined to the development
of intelligent identification programs, such as that created by
Khalid et al. (2008) who used artificial neural networks to
train a computer to recognize the wood of 30 tropical
species from Malaysia. In addition, Esteban et al. (2009)
used artificial neural networks to differentiate between
Juniperus cedrus and J. phoenicea with a 92 % probability.
As far as we are aware, there is no record of naı
¨ve Bayes classi-
fication being used for wood identification.
The aim of this study was to use a combination of qualitative
and quantitative wood anatomy and statistical analysis to estab-
lish one set of characters that could be used to distinguish
D. nigra from similar species in the genus. This was
approached in three ways: (1) a comparison of qualitative
and quantitative wood anatomy; (2) using an unsupervised
classification technique, PCA; and (3) with a supervised classi-
fication technique, naı
¨ve Bayes classification. Ultimately we
intend to extend this study to include all the commercial
Dalbergia species from the Americas, Africa and Asia.
MATERIALS AND METHODS
In addition to Dalbergia nigra, six Latin American Dalbergia
species were studied because they have some degree of
Gasson et al. — Wood identification of Dalbergia nigra46
commercial importance and are similar enough to D. nigra
in terms of overall appearance and/or anatomy, to be mistaken
for it.
(1) D. cearensis (Brazilian kingwood) is a component of the
caatinga vegetation, occurring from Ceara
´and southern
Piauı
´to southern Bahia in Brazil. The timber is violet
brown to black and is used mostly for decorative purposes,
such as inlay work and marquetry (Carvalho, 1989).
(2) D. miscolobium (various common names, all Portuguese:
jacaranda
´-do-cerrado, jacaranda
´-do-campo, caviu
´na-do-
cerrado) is a typical component of the cerrado vegetation
in Brazil. Its timber is similar to D. nigra (Rizzini, 1978
cited by Carvalho, 1989) but the tree has a ‘tortuous
trunk’, unique amongst Dalbergia (Carvalho, 1989), limit-
ing its use to small products.
(3) D. spruceana (Amazon rosewood) is from Brazil, Bolivia
and Venezuela (Carvalho, 1989, 1997). According to
Kuhlmann (1977, cited by Carvalho, 1989) it is found in
both evergreen and deciduous Amazonian forests. It has
dark brown to black wood (Carvalho, 1989) and is used in
cabinet work, as well as for smaller items such as knife
and brush handles (Loureiro and Silva, 1968 cited by
Carvalho, 1989).
(4) D. granadillo (‘granadillo’) is native to Mexico and
traded as cocobolo, a name also applied to D. retusa
(from Mexico to Colombia). The timber is deep red to
black and is used extensively for the production of cutlery
handles as well as other small items, ranging from jewellery
boxes to musical instruments (Flynn and Holder, 2001).
(5) D. stevensonii (Honduras rosewood) occurs in Belize,
south-east Mexico and Guatemala and is commonly
known as Honduras rosewood (Richter et al., 1996). The
timber is brown to purplish, often with black markings
and is used primarily for the production of percussion bars
of xylophones and occasionally for veneers and cabinets
(Flynn and Holder, 2001). Guatemalan D. stevensonii is pro-
tected by CITES Appendix III.
(6) D. tucurensis (Guatemala rosewood) including
D. cubilquitensis is native to Honduras and Guatemala.
The wood is a relatively recent addition to the international
market and is exported mainly for use in decorative veneers
(Richter et al., 1996).
Another commercially important Brazilian species of
Dalbergia is D. decipularis (Brazilian tuplipwood). This
species was not included in the present study, since it produces
a yellow or pinkish wood, which is easily distinguished from
the brown to purplish black wood of D. nigra. Dalbergia
retusa (from Mexico to Panama) has similar coloured wood
to D. nigra but was not included in this study due to a lack
of specimens with reliable provenance. However, in the speci-
mens that are available, the pattern of diffuse-in-aggregate par-
enchyma is very regular, forming an almost reticulate pattern,
and this appears to be diagnostic.
Specimens were obtained from the Economic Botany col-
lection at RBG Kew (Kw), the Forest Products Laboratory,
Wisconsin, USA (MADw and SJRw), the Institute for
Technology Research of the Federal State, Sao Paulo, Brazil
(BCTw) and the Federal Research Centre for Forestry and
Forest Products, Hamburg, Germany (RBHw). Table 1
shows the identity of each specimen studied, together with
its current location and, where available, collection infor-
mation. Only heartwood specimens were used to increase the
validity of comparisons.
Preparation of slides
Sectioning blocks, approx. 1 cm
3
were softened in boiling
water or in a 5 % solution of ethylenediamine placed in a
vacuum oven at 1000 mb, 60 8C for up to 40 h (MacLachlan
and Gasson, 2010, modified from Kukachka, 1977).
Transverse, tangential and radial sections were cut (10 –
20 mm thick) using a Reichert sliding microtome. The sections
were stained in Safranin (1 % in 50 % ethanol) for 5 min and
Alcian Blue (1 % aqueous) for 2 min. Sections were then
rinsed with distilled water, before being dehydrated in a
series of alcohols, 3 min each in 50 %, 70 %, 95 % and 100
% (twice). Finally, sections were rinsed in histoclear and
mounted in Euparal on a microscope slide.
Choice of characters
All of the species studied have the following characters from
the IAWA List (IAWA Committee, 1989), characters not listed
are either absent or do not apply:
5. Wood diffuse porous
13. Simple perforation plates
22. Intervessel pits alternate
29. Vestured pits
30. Vessel-ray pits with distinct borders; similar to interves-
sel pits in size and shape throughout the ray cell
58. Gums and other deposits in heartwood vessels
61. Fibres with simple to minutely bordered pits
66. Non-septate fibres present
76. Axial parenchyma diffuse
77. Axial parenchyma diffuse-in-aggregates
97. Ray width one to three cells
104. All ray cells procumbent
118. All rays storied
120. Axial parenchyma and/or vessel elements storied
136. Prismatic crystals present.
In D. cearensis,D. granadillo and D. stevensonii, growth rings
are always distinct (character 1), but in the rest of the species
studied, growth boundaries may be distinct, or indistinct/
absent (character 2).
Dalbergia cearensis and D. miscolobium have a mean tan-
gential vessel diameter of 50 – 100 mm (character 41),
whereas, D. tucurensis has a mean tangential vessel diameter
of 200 mm (character 43). In the four other species
studied, mean tangential vessel diameter is between 100 and
200 mm (character 42).
All of the species studied, except D. cearensis,have5
vessels mm
22
(character 46). Dalbergia cearensis has 20– 40
vessels mm
22
(character 48).
Dalbergia cearensis and D. stevensonii lack abundant vasi-
centric parenchyma (character 79), whereas in D. miscolobium,
parenchyma is always associated with the vessels and frequently
Gasson et al. — Wood identification of Dalbergia nigra 47
forms winged-aliform (character 82) or confluent patterns (char-
acter 83). In the remaining species, parenchyma pattern is more
variable, although there is a tendency towards winged-aliform
parenchyma in D. tucurensis.
This illustrates the difficulties of using only the IAWA List
to distinguish between members of the same genus; the char-
acters are too broad to allow subtle distinctions to be made.
Therefore, for this study, characters were chosen using the
IAWA List as a guide, but with modifications to allow for
increased discrimination accuracy and subsequently species
level separation.
Quantitative characters:
(1) Mean vessel lumen diameter (mm)
(2) Maximum vessel lumen diameter (mm)
(3) Minimum vessel lumen diameter (mm)
(4) Vessel frequency (1 mm
22
)
(5) Solitary vessel frequency (1 mm
22
)
(6) Mean ray height (cells)
(7) Maximum ray height (cells)
(8) Minimum ray height (cells)
(9) Mean ray width (cells)
(10) Ray frequency (10 mm
22
)
(11) Frequency of axially fused rays (10 mm
22
)
(12) Frequency of uniseriate rays (10 mm
22
).
Qualitative characters:
(13) Growth ring boundaries distinct or indistinct/absent
(14) Paratracheal axial parenchyma scanty or abundant
vasicentric
(15) Axial parenchyma winged aliform present or absent
(16) Axial parenchyma confluent present or absent
TABLE 1. Collection details of the specimens examined
Specimen no. Species Xylarium no. Collector and no. Country
1D. cearensis BCTw 18817 A. Mattos Filho, C. T. Rizzini Brazil
2D. cearensis MADw 31946* A. Dahlgren Brazil
3D. cearensis SJRw 44336
†
A. Ducke; wood 422 Brazil
4D. cearensis SJRw 45480 A. Ducke Brazil
5D. miscolobium BCTw 1588 Brazil
6D. miscolobium BCTw 3659 Brazil
7D. miscolobium BCTw 4133 Brazil
8D. miscolobium BCTw 15956 Brazil
9D. miscolobium RBHw 10922 E.P. Heringer 3349 Brazil
10 D. nigra Kw 6269 Brazil
11 D. nigra Kw 6270 F.H. Pierpont 85 Brazil
12 D. nigra Kw 6272 Brazil
13 D. nigra Kw 72717 A. Euponino, F. Souza Santo 414 Brazil
14 D. nigra SJRw 3273 (MADw 10769)
†
H.N. Whitford 76 Brazil
15 D. nigra SJRw 4146 (MADw 7017)
†
H.M. Curran 6 Brazil
16 D. nigra SJRw 4230 Brazil
17 D. nigra SJRw 5990 Brazil
18 D. spruceana BCTw 13349 Brazil
19 D. spruceana BCTw 14315 Brazil
20 D. spruceana BCTw 16612 Brazil
21 D. spruceana Kw 21266 M. Bastos Brazil
22 D. spruceana MADw 18588
‡
B. A. Krukoff 4921 Brazil
23 D. spruceana RBHw 7118 Brazil
24 D. spruceana RBHw 13049 Brazil
25 D. spruceana SJRw 22610 (MADw 31968)
†
A. Ducke 150 Brazil
26 D. tucurensis BCTw 12619
27 D. tucurensis RBHw 13987 Honduras
28 D. tucurensis RBHw 18255 Honduras
29 D. tucurensis RBHw 16326
30 D. tucurensis RBHw 18277
31 D. tucurensis SJRw 3738 (MADw 31973)
†
H. N. Whitford, L. R. Stadtmiller 79 Guatemala/Honduras
32 D. tucurensis SJRw 10729 (MADw 31975)
†
C. Galusser 9 Guatemala
33 D. tucurensis SJRw 3721 (MADw 10836)
†
H. N. Whitford, L. R. Stadtmiller 61 Guatemala/Honduras
34 D. granadillo BCTw 18439
35 D. granadillo RBHw 24160 Mexico
36 D. granadillo MADw 3781 L. E. Harthan Mexico
37 D. granadillo SJRw 38303 Nicaragua
38 D. granadillo SJRw 4410
†
J. A. Gamon Mexico
39 D. stevensonii RBHw 13048 Honduras
40 D. stevensonii RBHw 10840 Belize
41 D. stevensonii RBHw none
42 D. stevensonii RBHw none
43 D. stevensonii Kw 6303 F. H. Pierpont Honduras
Voucher is held by: *the Field Museum in Chicago,
†
MAD (housed in the University of Wisconsin) or
‡
the New York Botanical Garden.
Gasson et al. — Wood identification of Dalbergia nigra48
Characters 1– 3 correspond to characters 40–44 on the
IAWA List, but use actual measurements rather than size
classes.
Character 4 corresponds to IAWA characters 46– 51, and
character 5 is equivalent to characters 9– 11. Actual frequen-
cies were used instead of frequency classes to increase accu-
racy, and an area of 10 mm
2
was examined, as there were
often only 3 or 4 vessels mm
22
, which did not give enough
variation to separate the species.
Characters 6– 8 replace IAWA List characters 102 and 103,
with height in cells providing more opportunity for distinction
than recording rays taller than 1 mm (in the Dalbergia species
studied, the majority of rays were under 200 mm high, and
none were .500 mm tall), or of two distinct heights.
Characters 9 and 12 replace IAWA List characters 96 and
97; since all rays were 1– 3 cells wide, mean ray width indi-
cates whether more uniseriate (mean ray width close to 1) or
tri-seriate rays were present (mean ray width over 2).
Characters 10 – 12, which measure ray frequency, are not
covered by the IAWA List. Rays per 10 mm
2
was used
instead of the more conventional area of 1 mm
2
,because
axially fused and uniseriate rays were rare in some specimens,
and would have been missed altogether using the smaller
area.
Character 13 is IAWA List characters 1 and 2. Character 14
is IAWA List characters 78 and 79. Characters 15 and 16 are
IAWA characters 82 and 83.
Microscopy and measurements
Qualitative characters were determined by examining trans-
verse sections, approx. 10 mm
2
,at40 magnification, using
light microscopy. Presence of a qualitative character was
scored as 1 and absence as 0. For each species, the percentage
of specimens in which each character was present was calculated.
Quantitative characters were measured using a light micro-
scope with a drawing tube attached. Measurements were super-
imposed onto grids corresponding to 10 mm
2
on the slide to
facilitate counting. Vessels were observed using transverse
sections at 40 magnification, whereas rays were viewed
using tangential longitudinal sections at 100 magnification.
Rays bisecting the bottom or left-hand side of the grid were
included in the count; rays bisecting the top or right-hand
side of the grid were excluded. The measurements for vessel
frequency were then divided by 10 to give vessels per square
millimetre, which is the IAWA List character. For mean
vessel diameter, mean ray height and mean ray width, 30
measurements were made at random. The mean and standard
TABLE 2. Qualitative and quantitative wood anatomy measurements, summarized by species
D. cearensis D. miscolobium D. nigra D. spruceana D. tucurensis D. granadillo D. stevensonii
QUALITATIVE CHARACTERS (%)
Growth rings distinct 100 80 71 50 63 100 100
Parenchyma abundant vasicentric 0 100 71 38 75 20 0
Axial parenchyma winged aliform 0 20 14 63 75 0 0
Axial parenchyma confluent 0 40 29 50 38 0 0
QUANTITATIVE CHARACTERS
Vessel diameter (mm)
Mean +s.d. 76.7+41.398
.7+48.3 138.2+76.6 130.6+77.9 205.2+73.1 167.4+67.5 153.4+53.7
Maximum 248 224 656 344 440 299 374
Minimum 16 24 16 16 66 42 50
Vessel frequency (mm
22
)
Mean +s.d. 22.0+7.94
.5+2.12
.8+1.13
.5+1.22
.8+1.43
.8+2.63
.1+1.0
Maximum 29 8 5 5 6 8 4
Minimum 11 2 2 2 1 1 2
Frequency solitary vessels (mm
22
)
Mean +s.d. 5.1+2.61
.4+0.51
.5+0.61
.0+0.51
.3+0.61
.9+1.52
.0+1.0
Maximum 9 2 3 2 2 5 4
Minimum 3 1 0 0 0 0 0
Ray height (cells)
Mean +s.d. 6.3+1.75
.2+1.37
.1+1.57
.1+1.76
.1+1.46
.2+1.26
.7+1.3
Maximum 10 8 12 11 9 9 10
Minimum 2 2 3 2 3 3 3
Ray width (cells)
Mean +s.d. 1.7+0.61
.5+0.62
.0+0.41
.7+0.72
.0+0.51
.5+0.51
.9+0.5
Ray frequency (10 mm
22
)
Mean +s.d. 64.5+14.597
.6+11.364
.6+15.656
.0+9.955
.6+8.069
.6+8.957
.0+13.3
Maximum 80 113 84 68 67 83 77
Minimum 47 83 42 39 41 59 40
Frequency axially fused rays (10 mm
22
)
Mean +s.d. 1.5+1.96
.4+5.80
.1+0.40
.9+1.10
.5+1.12
.2+2.90
.2+0.4
Maximum 4 14 1 3 3 7 1
Minimum 0 1 0 0 0 0 0
Frequency uniseriate rays (10 mm
22
)
Mean +s.d. 25.25 +19.752
.8+16.75
.9+3.823
.4+13.97
.8+7.037
.0+20.86
.4+4.3
Maximum 48 75 11 40 22 61 13
Minimum 6 37 2 4 0 14 2
Gasson et al. — Wood identification of Dalbergia nigra 49
deviation were calculated for each character for each species.
Maximum and minimum values are the single highest or
lowest measurement recorded within a species.
PCA
PCA was conducted according to the methodology of Mo
¨ller
et al. (2007) using the multivariate statistical package R-pack
Le Progiciel R.4.0d10 (http://www.bio.umontreal.ca/Casgrain
/en/labo/R/v4/progress.html). Since PCA works by finding
the directions of maximum variability in the data, and qualitat-
ive characters were scored only as 0 or 1, they were not
included in the PCAs.
Two PCAs were carried out, one on the four Brazilian
species – D. nigra,D. spruceana,D. miscolobium and
D. cearensis – and a second on all seven species together.
1
mm 1
mm
1
mm 1
mm
1
mm 1
mm
1
mm 1
mm
A B
CD
EF
GH
FIG. 1. Transverse sections: (A) D. cearensis (MADw 31946); (B) D. miscolobium (BCTw 1588); (C) D. nigra (Kw 6270); (D) D. spruceana (RBHw 7118); (E)
D. tucurensis (SJRw 3738); (F) D. granadillo (SJRw 4410); (G) D. stevensonii (RBHw trade specimen); (H) D. nigra (SJRw 5990). Note the differences in vessel diameter.
Gasson et al. — Wood identification of Dalbergia nigra50
Naı
¨ve Bayes classification
Class prediction was carried out using naı
¨ve Bayes classifi-
cation (Hand and Yu, 2001) on the four characters: minimum
vessel diameter; frequency of solitary vessels; mean ray width;
and frequency of axially fused rays. Minimum vessel diameter
and mean ray width were modelled as normal distributions; the
two measurements of frequency were modelled as binomial
distributions with pseudocounts of 0.25 included to compen-
sate for under-sampling (Durbin et al., 1998). Equal priors
were used for two classes to avoid bias, and marginal pos-
teriors were used to compute the joint posterior in order to
ensure equal weight to the four characters. The model was
200 µm 200 µm
200 µm 200 µm
200 µm 200 µm
200 µm 200 µm
AB
CD
EF
GH
FIG. 2 . Tangential longitudinal sections: (A) D. cearensis (MADw 31946); (B) D. miscolobium (BCTw 3659); (C) D. nigra (Kw 6269); (D) D. spruceana
(BCTw 14315); (E) D. tucurensis (SJRw 3738); (F) D. granadillo (SJRw 38303); (G) D. stevensonii (Kw 6303); (H) D. nigra (SJRw 5990).
Gasson et al. — Wood identification of Dalbergia nigra 51
validated using leave-one-out cross-validation (Miller, 1974).
The four characters were selected on the basis of: biological
relevance; reasonable distinction of the classes in univariate
analyses; having low correlation with each other (the largest
R
2
between selected characters is 0.178); and statistical
expediency.
Miller and Wiemann (2006) report that specimen 17,
D. nigra SJRw 5990 has been misidentified and this was con-
firmed by chemical analysis of extracts (Geoffrey Kite, RBG
Kew, pers. comm.). This specimen was therefore not included
when the quantitative and qualitative wood anatomy data were
analysed according to species, and was also excluded from all
training sets used in the naı
¨ve Bayes classification, but was
used as a test specimen. It was left in the PCAs as this test
treats each specimen individually.
RESULTS AND DISCUSSION
Qualitative wood anatomy
No one species was consistent in, and unique for, any single
discrete character or combination of discrete characters
studied. Therefore these characters on their own are not
enough to identify these species, or to identify D. nigra.
Dalbergia cearensis and D. stevensonii had identical results
and showed the most consistency across the specimens studied,
with all specimens from both species showing distinct growth
rings and lacking abundant vasicentric, or axial parenchyma in
winged-aliform or confluent patterns (see Table 2).
Distinct growth rings were also recorded in all specimens
of D. granadillo and in four out of five specimens of
D. miscolobium.ForD. nigra,D. spruceana and
D. tucurensis, the presence or absence of growth ring bound-
aries was more variable with a maximum of 71 % (D. nigra)
of specimens having distinct growth rings. However, the
presence or absence of growth rings is most likely to be a
reflection of the environment in which the tree grew, may
vary in a given tree, and is therefore not a good character for
determining species identity.
Abundant vasicentric parenchyma was present in all
the specimens of D. miscolobium and in six out of eight
specimens of D. tucurensis. However, D. nigra,D. spruceana
and D. granadillo showed more variability in this character,
with a maximum of 75 % (D. tucurensis)ofspecimens
having parenchyma that was abundant vasicentric. In addition
to D. cearensis and D. stevensonii, no specimens of
D. granadillo showed a winged-aliform or confluent pattern of
axial parenchyma. In the four remaining species, these patterns
were sometimes present, but never for all specimens of one
species. Patel and Shah (1980) also found inconsistency in the
patterns of axial parenchyma present in specimens of
Dalbergia latifolia from India. Although a more comprehensive
study might show consistent trends that have not been observed
here, it appears that parenchyma patterns are inconsistent and
are not diagnostic for these Dalbergia species.
Quantitative wood anatomy
Dalbergia cearensis is one of the most easily identifiable of
the Latin American species of Dalbergia due to the presence
of numerous small vessels (Kribs, 1959; Miller and
Wiemann, 2006). The results of this study show D. cearensis
can be separated from the other species by vessel frequency,
with all specimens of D. cearensis, and no specimens from
other species, having a vessel frequency of over 10 vessels
mm
22
(see Fig. 1). Dalbergia miscolobium, had the second
highest vessel frequency at 4.48 vessels mm
22
and
D. tucurensis had the lowest vessel frequency at 2.75 vessels
mm
22
(see Table 2 and Fig. 1).
Dalbergia miscolobium also stands out in terms of ray
characteristics, with the shortest ray height, greatest ray
frequency, highest frequency of uniseriate rays and highest
frequency of axially fused rays. A ray frequency of over
90 rays 10 mm
22
distinguishes four out of five of the
D. miscolobium specimens from all the other specimens
studied (see Fig. 2). This may reflect its twisted habit when
compared with the other species which have straighter trunks
(Carvalho, 1997). Dalbergia granadillo has the second
highest ray frequency at 69.6rays10mm
22
. The difference
between D. granadillo and the species with the lowest ray fre-
quency, D. tucurensis is just 14 rays 10 mm
22
.
Quantitative characters cannot be used to distinguish
D. nigra from D. spruceana,D. tucurensis,D.granadillo or
D. stevensonii, as the results for D. nigra fall within the
range of these species. In particular, D. nigra and
D. spruceana had the most similar mean vessel diameter,
with vessels of D. nigra being, on average, just 7.6mm
larger than those of D. spruceana, and identical mean ray
heights.
PCA
Analysis of the Brazilian species alone and then all
seven species together yielded cumulative variance for PC1
and PC2 at 58.74 % for the former and 51.25 % for the
TABLE 3. PCA eigenvectors
Brazilian species All species
Character
Axis
1
Axis
2
Axis
3
Axis
1
Axis
2
Axis
3
Mean vessel
diameter
0.31 20.36 20.02 0.36 20.33 20.01
Maximum vessel
diameter
0.30 20.24 20.19 0.33 20.24 20.06
Minimum vessel
diameter
0.22 20.16 0.49 0.27 20.26 20.11
Vessel frequency 20.13 0.55 0.05 20.20 0.42 20.34
Solitary vessel
frequency
20.12 0.54 0.06 20.17 0.39 20.36
Mean ray height 0.39 0.18 0.16 0.29 0.40 0.33
Maximum ray height 0.37 0.17 0.28 0.24 0.42 0.34
Minimum ray height 0.27 0.09 20.34 0.27 0.13 20.01
Mean ray width 0.22 0.12 20.54 0.28 0.02 20.56
Ray frequency 20.35 20.23 20.17 20.34 20.24 20.05
Frequency uniseriate
rays
20.28 20.07 20.24 20.28 20.11 20.15
Frequency axially
fused rays
20.34 20.21 0.35 20.37 20.14 0.41
Percentage of
variation explained
38.07 20.67 12.50 35.81 21.44 11.48
Gasson et al. — Wood identification of Dalbergia nigra52
latter group. This is considerably less than the cumulative PC1
and PC2 score of 70– 90 % considered appropriate by Everitt
and Dunn (2001). The outcomes of the PCAs conducted for
this study are therefore treated with caution. However, when
the cumulative value of PCs 1–3 is considered, 71.23 % for
the Brazilian species and 68.73 % for all seven species were
achieved, which are higher than the 60 % reported by
MacLachlan and Gasson (2010) and thus some discussion of
the results produced by these PCAs is appropriate.
When considering the four Brazilian species alone, PC1
accounted for 38.07 % of the variance and was most highly cor-
related with ray characteristics (see Table 3). Figure 3A shows
that PC1 splits the species into two groups, with eight out of
nine specimens of D. cearensis and D. miscolobium being
located on the negative side of the axis, and 13 out of 15 speci-
mens of D. nigra and D. spruceana on the positive side. PC2,
which correlated most strongly with vessel characteristics can
then be used to separate D. miscolobium, with all specimens
17
–4
–2
0
2
4
6A
B
C
D
E
F
–5 –4 –3 –2 –1 0 1 2 3
D. cearensis
D. miscolobium
D. nigra
D. spruceana
17
–4
–2
0
2
4
PC1 PC1
PC3
PC3
17
–3
–2
–1
0
1
2
3
4
6420
–2–4
PC2
PC3 PC2
17
–4
–2
0
2
4
6
PC2
D. cearensis
D. miscolobium
D. nigra
D. spruceana
D. tucurensis
D. granadillo
D. stevensonii
17
–4
–2
0
2
4
5
31–1–3–5
–5 –4 –3 –2 –1 0 1 2 3
PC1 PC1
5
31–1–3–5
17
–4
–2
0
2
4
6420–2–4
PC2
PC3
FIG. 3 . Principal component axes score plots: (A) Brazilian species PC1 vs. PC2; (B) Brazilian species, PC1 vs. PC3; (C) Brazilian species PC2 vs. PC3; (D) all
species PC1 vs. PC2; (E) all species PC1 vs. PC3; (F) all species PC2 vs. PC3.
Gasson et al. — Wood identification of Dalbergia nigra 53
on the negative side, fromD. cearensis on the positive side of the
axis. However, these distinctions were apparent directly from the
raw data and PC1 and PC2 are still unable to split D. nigra from
D. spruceana. Figure 3B and C shows PC3 plotted against PC1
and PC2, respectively. PC3 accounted for only 12.40 % of the
variance, and the specimens were not separated by this axis.
When all seven species were considered together, the same
eight characters made the biggest contributions to PC1 and
PC2, but the distinction between vessel and ray characters
did not exist because D. tucurensis had the largest mean
vessel diameter and lowest vessel frequency and D. cearensis
had the smallest mean vessel diameter and the highest vessel
frequency, thus, when these two species are considered
together, the variability in vessel characteristics is increased
(see Table 3). However, there is no difference in terms of
the ability of the PCs to separate the species. Specimens of
D. nigra,D. spruceana,D. tucurensis and D. stevensonii
tend to be located on the positive side of PC1, with
D. miscolobium,D cearensis and D. granadillo on the negative
side, but PC2 is unable to separate the species any further (see
Fig. 3D– F). As was the case when only the Brazilian species
were considered, PC3 does not separate the species (see
Fig. 3E, F).
In all cases, specimen 17, D. nigra SJRw 5990, is the furth-
est D. nigra outlier on PC1, supporting Geoffrey Kite’s chemi-
cal analysis which suggests it has been misidentified.
PCA identifies the characters with the maximum variability
in the data, and the fact that the PCs are unable to separate the
species in this study means that the variation is largely
between members of the same species, rather than between
members of different species.
Naı
¨ve Bayes classification
Table 4 shows the outcome of the naı
¨ve Bayes classification,
with each specimen classified using the other 41 specimens as
TABLE 4. Results of naı
¨ve Bayes classification
Specimen no. Species Overall outcome Minimum vessel diameter Solitary vessel frequency Mean ray width Frequency axially fused rays
1D. cearensis 2þ22 þ
2D. cearensis 2þ2þ2
3D. cearensis 2þ22 þ
4D. cearensis 2þ22 2
5D. miscolobium 2þþ22
6D. miscolobium 2þ22 2
7D. miscolobium 2þ22 2
8D. miscolobium 2þ22 2
9D. miscolobium 2þ22 2
10 D. nigra þþ þþ þ
11 D. nigra þþ 2þ2
12 D. nigra þþ þþ þ
13 D. nigra þþ þþ þ
14 D. nigra þþ þþ þ
15 D. nigra þþ þ2þ
16 D. nigra þþ 2þþ
18 D. spruceana 2þ22 2
19 D. spruceana 2þþ2þ
20 D. spruceana 2þþ22
21 D. spruceana þþ þþ þ
22 D. spruceana 2þ22 þ
23 D. spruceana 22 þ22
24 D. spruceana 2þ22 þ
25 D. spruceana 2þ2þþ
26 D. tucurensis 22 þ2þ
27 D. tucurensis 22 2þþ
28 D. tucurensis 22 2þþ
29 D. tucurensis 22 þþ þ
30 D. tucurensis 22 2þþ
31 D. tucurensis 22 þ2þ
32 D. tucurensis 22 þþ 2
33 D. tucurensis þ2þþ þ
34 D. granadillo 22 þ2þ
35 D. granadillo 2þ22 2
36 D. granadillo 22 þ2þ
37 D. granadillo 22 þ2þ
38 D. granadillo 22 þ22
39 D. stevensonii þþ þ2þ
40 D. stevensonii 22 2þþ
41 D. stevensonii 22 þþ þ
42 D. stevensonii 22 þþ þ
43 D. stevensonii þ2þþ þ
þ¼‘nigra’; 2¼‘not nigra’.
Gasson et al. — Wood identification of Dalbergia nigra54
a training set. Mean ray width was the most successful
character, correctly identifying 27 out of 41 specimens, with
six out of seven D. nigra correctly identified. Frequency of
axially fused rays was the character least good at classifying
the specimens when used alone, correctly identifying only
18 out of 41 specimens, but again six out of seven D. nigra
were classified correctly using just this character.
Overall, 39 out of 42 specimens were correctly identified,
with all seven D. nigra specimens being classified as ‘nigra’.
Of the 34 remaining specimens, 30 were classified correctly
as ‘not nigra’, with four classified incorrectly as ‘nigra’. This
gives a false negative rate of 0 out of 7 and a false positive
rate of 4 out of 11, or 36.36 %. When developing the model,
the false negative rate was minimized because any unknown
specimen being tested under CITES regulations and classified
as ‘not-nigra’ can then be confidently assumed to be another
species, and thus can be imported or exported legally. Any
specimen that is classified as ‘nigra’, which form a minority,
would need further investigation.
Specimen 17, D. nigra SJRw 5990, was classified as ‘not
nigra’, which is in line with the chemical analysis of
Geoffrey Kite and strongly suggests that this specimen has
been misidentified.
One limitation of statistical analysis is that the results cannot
be generalized to apply to species that were not used in the
original analysis. Therefore, the naı
¨ve Bayes classification
developed for the present study should only be applied when
the specimen under examination is strongly believed to
belong to one of the seven species used to develop the model.
CONCLUSIONS
This study confirms that D. cearensis can be reliably distin-
guished from D. nigra and the other species in this study by
its small, numerous vessels, with any specimen with a vessel
frequency in excess of 10 vessels mm
22
being D. cearensis.
Specimens with a high ray frequency, over 100 rays
10 mm
22
, and a high number of axially fused rays are likely
to be D. miscolobium.
PCA is not able to provide a set of characters that dis-
tinguishes D. nigra from other commercially important Latin
American members of the genus, because the majority of vari-
ation is within members of the same species, not between
species.
However, using the four characters – minimum vessel diam-
eter, frequency of solitary vessels, mean ray width, and fre-
quency of axially fused rays in a naı
¨ve Bayes classification
– unidentified specimens can be determined as ‘not nigra’
with no false negatives. Specimens determined as ‘nigra’ are
twice as likely to be genuine D. nigra as not. This suggests
that whilst wood anatomy alone is unlikely to provide the
level of identification certainty needed by legislation such as
CITES, it can be used as a relatively inexpensive and straight-
forward way of reducing the number of specimens that would
need more comprehensive study.
The success of naı
¨ve Bayes classification over PCA indi-
cates that such supervised classification techniques could be
used more widely to solve problems in wood anatomy.
Indeed it is a promising technique for helping to solve other
intractable biological identification problems.
ACKNOWLEDGEMENTS
We would like to thank H. G. Richter and G. Koch (Hamburg)
and R. Pigozzo (IPT, Sao Paulo) for providing several wood
samples. Ian MacLachlan helped with our initial methodology
and commented on the manuscript.
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