Numerical analyses of seed morphology and its taxonomic significance in the genus Oxytropis DC. (Fabaceae) from northwestern China

Abstract

The lack of diagnostic taxonomic characteristics in some species complexes leave the species delimitation of Oxytropis DC. unresolved. Seed morphological features have proved to be useful diagnostic and taxonomic characteristics in Fabaceae. However, there are few systematic studies on the seed characteristics of Oxytropis . Here, we used scanning electron and stereoscopic microscopy to investigate the seed characteristics of 35 samples obtained from 21 Oxytropis species from northwest China. Our examination showed two main types of hilum positions, terminal and central, and five different types of seed shapes: prolonged semielliptic, reniform, prolonged reniform, quadratic, and cardiform. Seven different sculpturing patterns were identified: scaled, regulated, lophate with stellated testa cells, simple reticulate, rough, compound reticulate, and lophate with rounded testa cells. The seeds ranged from 1.27 to 2.57 mm in length and from 1.18 to 2.02 mm in width, and the length-to-width ratio ranged from 0.89 to 1.55 mm. The seed shape was constant within species and was useful for species delimitation within the genus Oxytropis when combined with other macroscopic traits. In contrast, the sculpturing patterns were highly variable at the species level and could not be used for species identification. Results of the cluster analysis and principal component analysis (PCA) indicated that the seed traits of Oxytropis species are useful for taxa identification at the species level, but have low taxonomic value at the section level.

Full text

Numerical analyses of seed morphology and its
taxonomic significance in the genus Oxytropis DC.
(Fabaceae) from northwestern China
Xiang Zhao1, Yingying Liu1, Jigang Li1, Hui Zhang1,
Lingyun Jia1, Qinzheng Hou1, Kun Sun1
1College of Life Sciences, Northwest Normal University, Lanzhou 730070, Gansu, China
Corresponding authors: Kun Sun (kunsun@nwnu.edu.cn); Qinzheng Hou (hou_qzh@nwnu.edu.cn)
Academic editor: C. Morden|Received 1 November 2022|Accepted 16 February 2023|Published 21 March 2023
Citation: Zhao X, Liu Y, Li J, Zhang H, Jia L, Hou Q, Sun K (2023) Numerical analyses of seed morphology and
its taxonomic signicance in the genus Oxytropis DC. (Fabaceae) from northwestern China. PhytoKeys 222: 49–67.
https://doi.org/10.3897/phytokeys.222.96990
Abstract
e lack of diagnostic taxonomic characteristics in some species complexes leave the species delimitation
of Oxytropis DC. unresolved. Seed morphological features have proved to be useful diagnostic and taxo-
nomic characteristics in Fabaceae. However, there are few systematic studies on the seed characteristics of
Oxytropis. Here, we used scanning electron and stereoscopic microscopy to investigate the seed character-
istics of 35 samples obtained from 21 Oxytropis species from northwest China. Our examination showed
two main types of hilum positions, terminal and central, and ve dierent types of seed shapes: prolonged
semielliptic, reniform, prolonged reniform, quadratic, and cardiform. Seven dierent sculpturing patterns
were identied: scaled, regulated, lophate with stellated testa cells, simple reticulate, rough, compound
reticulate, and lophate with rounded testa cells. e seeds ranged from 1.27 to 2.57 mm in length and
from 1.18 to 2.02 mm in width, and the length-to-width ratio ranged from 0.89 to 1.55 mm. e seed
shape was constant within species and was useful for species delimitation within the genus Oxytropis when
combined with other macroscopic traits. In contrast, the sculpturing patterns were highly variable at the
species level and could not be used for species identication. Results of the cluster analysis and principal
component analysis (PCA) indicated that the seed traits of Oxytropis species are useful for taxa identica-
tion at the species level, but have low taxonomic value at the section level.
Keywords
China, cluster analysis, Oxytropis, PCA, seed morphology, SEM, taxonomy
Copyright Xiang Zhao et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0),
which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
PhytoKeys 222: 49–67 (2023)
doi: 10.3897/phytokeys.222.96990
https://phytokeys.pensoft.net
RESEARCH ARTICLE
Launched to accelerate biodiversity research
A peer-reviewed open-access journal

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
50
Introduction
e genus Oxytropis DC. belongs to the tribe Galegeae (Fabaceae: Papilionoideae). It
has been reported to be one of the largest groups of angiosperms, comprising approxi-
mately 330 species. e genus is distributed mainly in the cold mountainous regions of
Asia, Europe, and North America (Polhill 1981; Zhu et al. 2010). It is thought to have
been derived from Astragalus L. approximately 12–16 Ma, with which it shares many
morphological features (Wojciechowski 2005). e genus Oxytropis is distinguished
from Astragalus by beaked keels, asymmetrical leaets, and acaulescent habit (Barneby
1952). Likely because of its relatively recent diversication, many taxonomic relation-
ships within Oxytropis remain problematic (reviewed in Welsh 2001).
e genus Oxytropis was rst established in 1802 by De Candolle (De Candolle
1802). It included 33 species, and he divided them into three groups according to
whether stipules are adherent to stems or not and whether leaets are opposite, verticil-
late, or neither. Bunge’s (1874) comprehensive treatment of Oxytropis species in Eurasia
identied four subgenera, 19 sections, and 181 species. His research also marked the
beginning of modern Oxytropis research. Vassilczenko (1948) revised the work on Oxy-
tropis in the Flora of USSR and separated the genus into six subgenera, 21 sections, and
276 species. Pavlov (1961) divided Oxytropis into four subgenera, 15 sections, and 124
species in the Flora of Kazakhstan. Leins and Merxmüller (1968) compiled 24 species,
three subspecies, one variety, and two suspected species from Europe and divided them
into two groups. Zhang (1998) recognized six subgenera, 146 species, 12 varieties,
and three forms in Flora Reipublicae Popularis Sinicae. In contrast, Zhu and Ohashi
(2000) recognized 125 species and four varieties in China. Welsh (2001) revised the
genus Oxytropis in North America to include 57 taxa in only 22 species. Later, Zhu
et al. (2010) taxonomically revised the genus in China and reported that it comprises
three subgenera and 20 sections containing 133 species. ese previous treatments of
Oxytropis claried many taxonomic problems. However, the lack of diagnostic taxo-
nomic characteristics in some Oxytropis complexes has led to diculties and dierences
in species delimitation, leaving the internal classication of Oxytropis unresolved.
Seed morphological features, such as seed shape, hilum shape, sculpturing pat-
tern, and size, have been proven to be useful diagnostic and taxonomic characteristics
in some genera of Fabaceae and other families (Lersten and Gunn 1981; Solum and
Lockerman 1991; López et al. 2000; Al-Gohary and Mohamed 2007; Salimpour et al.
2007; Vural et al. 2008; Venora et al. 2009; Zorić et al. 2010; Celep et al. 2012; Kaya
and Dirmenci 2012; Lantieri et al. 2013; Kamala and Aydin 2018; Rashid et al. 2018;
Shemetova et al. 2018; Rashid et al. 2020). In the genus Trifolium L., Salimpour et
al. (2007) reported that seed characteristics such as sculpturing pattern, shape, size,
and hilum position, can be used as taxonomic markers within the section Lotoidea.
In contrast, Zorić et al. (2010) concluded that the seed characteristics do not support
infrageneric classication of Trifolium. Similarly, Shemetova et al. (2018) reported that
seed shapes, colours, sizes, surface sculptures, and hilum positions are very diverse in
Astragalus, and they emphasized that the systematic importance of seed characteristics

Seed morphology of Oxytropis DC. 51
needs to be evaluated in a phylogenetic context. However, Vural et al. (2008) found
that Astragalus seed sculpturing pattern and seed shape can be used as taxonomically
signicant characteristics at the species level, if supported by other macromorphologi-
cal characteristics. López et al. (2000) found that seed colour, weight, shape, and size,
presence of an aril, and hilum position can be used as diagnostic characteristics for
segregating two subtribes and delimiting lower taxonomic levels in the tribe Genisteae.
Similarly, Rashid et al. (2020) concluded that seed shape, sculpturing pattern, and
size are valuable characteristics for the identication and delimitation of species in the
tribes Astragaleae and Trifolieae. Kamala and Aydin (2018) and Rashid et al. (2018)
also reported that seed characteristics (coat, shape, colour, seed size, etc.) can be used
to identify taxa in the tribe Vicieae.
Seeds of Oxytropis species were rst studied by Solum and Lockerman (1991), who
documented the seed coat patterns of Oxytropis riparia Litv. and Oxytropis campestris (L.)
DC. Bojňanský and Fargašová (2007) studied the seeds of four European Oxytropis spe-
cies and recorded their size, colour, and other information. Farrington et al. (2008) stud-
ied the morphological properties of seeds of 15 Alaskan Oxytropis taxa and found that
seed coat micromorphology and anatomy can distinguish it from the genus Astragalus.
Meyers et al. (2013) analysed the seed characteristics of 22 Oxytropis species in Alaska
and concluded that seed coat types are highly variable at the species level and cannot be
used for species identication. Erkul et al. (2015) studied the morphological properties
of seeds of 13 Turkish Oxytropis taxa and found that seed characteristics have low taxo-
nomic value in distinguishing subgenera, sections, and species. e infraspecic variation
in seed traits has not been well addressed in most of the abovementioned studies because
of sampling limitations. Only Meyers et al. (2013) studied whether seed traits were stable
within species, along with studying the correlation between seed traits and the environ-
ment; however, they did not conduct any systematic analyses, such as cluster analysis.
Numerical taxonomy, also known as phenetics, mathematical taxonomy and mul-
tivariate morphometrics (Singh 2019), is mainly based on the overall anity (similar-
ity) at any taxonomic level. Quantitative traits have long been overlooked in taxo-
nomic studies until numerical methodologies, such as cluster analysis, started to be
widely applied in species delimitation (ien et al. 1975). Recently, dendrograms and
cladograms have been used instead of subjective analyses in many studies on the seed
morphology of Fabaceae (Erkul et al. 2015; Fayed et al. 2019; Abusaief and Boasoul
2021). However, quantitative seed traits of the genus Oxytropis, such as length, width,
length/width ratio, and weight, have not received much attention in taxonomic stud-
ies, possibly because these traits are considered uctuating, and this uctuation is ran-
dom or excessive.
Northwest China is one of the main distribution regions of the genus Oxytropis
(Zhang 1998; Zhu et al. 2010), but there is little research on the seed characteristics of
Oxytropis in this area. Here, we carried out the rst numerical analysis and microscopic
investigation of 35 samples belonging to 21 Oxytropis species from northwest China
using scanning electron and stereoscopic microscopy to elucidate the taxonomic sig-
nicance of their seed micromorphology.

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
52
Materials and methods
e present study was mainly based on seeds collected in the eld, with only a few
seeds obtained from herbarium vouchers housed at the herbarium of Northwest Nor-
mal University (HWTC; Table 1). Voucher specimens collected from wild seeds are
also kept at HWTC. e investigated species and their sources are listed in Table 1,
and the classication of genera by Zhu et al. (2010) was adopted. Seed morphology
was examined using a stereoscopic microscope (Leica M205 FA). For measuring seed
length and width among the samples from the eld, 80 mature and representative
seeds per population were measured, while among the samples from herbarium speci-
mens, 30 seeds per specimen were measured. e minimum–maximum range, mean,
standard deviations in seed length and width, and length/width ratio were calculated.
For SEM, the selected representative material was directly mounted onto aluminium
stubs with double adhesive tape and coated with gold prior to observation with a
HITA-CHIS-450 scanning electron microscope (NWNU University) at 25 kV.
Seed shapes and surface sculpturing were classied according to previous studies on
the microscopic morphology of Fabaceae seeds (Bojňanský and Fargašová 2007; Vural
et al. 2008; Al-Ghamdi 2011; Meyers et al. 2013; Erkul et al. 2015). Based on previous
studies and observations of seed morphology in the genus Oxytropis, seven seed traits,
including four quantitative and three qualitative traits, were selected for morphometric
analysis in the present study (Erkul et al. 2015). e selected characteristics and their
states for cluster analysis were as follows: 1. seed length (mm); 2. seed width (mm);
3. seed length/width ratio; 4. seed shape: cardiform (0), prolonged (1), reniform (2),
quadratic (3), prolonged semi-elliptic (4); 5. seed surface sculpturing: scaled (0), ru-
gulate (1), lophate with stellated testa cells (2), simple reticulate (3), rough (4), com-
pound reticulate (5), lophate with rounded testa cells (6); 6. hilum position: central
(0), terminal (1); 7. seed weight (g). For the seeds collected in the eld, 300 mature
and full seeds were randomly selected and their 100-seed weight was determined. For
the seeds collected from a few specimens, we randomly selected 30 seeds and weighed
the 10-seeds. e 100-seed weights determined from the seeds of the specimens in the
cluster analysis were expressed as 10-seed weights multiplied by 10.
Numerical analysis
Cluster analysis and principal component analysis (PCA) were performed using the
Origin 2022 software (OriginLab Corporation 2022). e raw data matrix included
quantitative traits, such as length, width, L/W ratio and weight, and qualitative char-
acteristics, such as shape, sculpturing, and hilum position. e qualitative character-
istics were coded using a presence/absence (0/1) matrix. Ward’s method was used for
cluster analysis using Euclidean distance to interpret the morphological similarities
among species. In the cluster analysis, Euclidean distance is one of the most commonly
used distance measurements in hierarchical clustering, which can reect the absolute

Seed morphology of Oxytropis DC. 53
Table 1. List of examined taxa with collection details.
Section Code Species Locality Coordinates Voucher Collection
date
Section Xerobia 1O ciliata Turcz. Yueliang Mountain 36°25'41.85"N,
105°42'23.71"E
X. Zhao 1947 2019
Section Polyadena 2O. muricata (Pall.) DC. Maxian Mountain 35°47'46.48"N,
103°58'12.64"E
X. Zhao 1903 2019
3O. muricata (Pall.) DC. Tiemu Mountain 35°58'32.21"N,
104°46'31.40"E
X. Zhao 1970 2019
Section Falcicarpae 4O. falcata Bunge Awangcang wetland park 33°45'32.85"N,
101°41'6.58"E
X. Zhao 1842 2018
5O. falcata Bunge Beach of Maqu section
of Yellow River
Unknown Gannan Grassland Team 60B Unknown
Section Baicalia 6O. ochrantha Turcz. Xinglong Mountain 35°45'52.41"N,
104°2'21.66"E
X. Zhao 1813 2018
7O. ochrantha Turcz. North mountain of
Pingliang
35°33'49.11"N,
106°41'2.34"E
X. Zhao 1837 2018
8O. bicolor Bunge Unknown Unknown Unknown 790043 Unknown
9O. bicolor Bunge Tiemu Mountain 35°58'32.21"N,
104°46'31.40"E
X. Zhao 1927 2019
10 O. racemosa Turcz. Yanchi 37°43'52.02"N,
107°23'55.77"E
X. Zhao 1946 2019
11 O. myriophylla (Pall.) DC. Erdaogou 35°25'19.39"N,
106°40'6.25"E
X. Zhao 1831 2018
12 O. myriophylla (Pall.) DC. Anguo 35°38'49.75"N,
106°28'54.92"E
X. Zhao 1833 2018
13 O. myriophylla (Pall.) DC. Maxian Mountain 35°47'46.48"N,
103°58'12.64"E
X. Zhao 1836 2018
Section Neimonggolicae 14 O. neimonggolica
C.W.Chang & Y.Z.Zhao
Helan Mountain 38°39'37.76"N,
105°48'34.42"E
X. Zhao 1948 2019
Section Eumorpha 15 O. imbricata Kom. Liancheng National
Nature Reserve
36°36'24.65"N,
102°49'34.30"E
X. Zhao 1809 2018
16 O. imbricata Kom. Taohe River 34°33'28.66"N,
102°34'53.99"E
X. Zhao 1940 2019
17 O. coerulea (Pall.) DC. Taitong Mountain 35°30'8.94"N,
106°35'54.90"E
X. Zhao 1832 2018
18 O. coerulea (Pall.) DC. Erdaogou 35°25'19.39"N,
106°40'6.25"E
X. Zhao 1833 2018
19 O. holanshanensis H.C.Fu Helan Mountain 38°39'37.76"N,
105°48'34.42"E
X. Zhao 1949 2019
Section Mesogaea 20 O. xinglongshanica
C.W.Chang
Maxian Mountain 35°46'46.16"N,
103°59'19.19"E
X. Zhao 1913 2019
21 O. xinglongshanica
C.W.Chang
Xinglong Mountain 35°46'20.53"N,
104°1'2.49"E
X. Zhao 1910 2019
22 O. glabra (Lam.) DC. Rabah Lake National
Nature Reserve
37°42'3.19"N,
107°2'33.46"E
X. Zhao 1950 2019
23 O. kansuensis Bunge Azi Test Station of LZU 33°39'57.96"N,
101°52'22.44"E
X. Zhao 1819 2018
24 O. kansuensis Bunge Charlie temple 32°45'7.95"N,
102°3'26.83"E
X. Zhao 1820 2018
25 O. taochensis Kom. Liupan Mountain 35°33'21.81"N,
106°25'21.54"E
X. Zhao 1838 2018
26 O. ochrocephala Bunge Nanhuang Mountain 36°22'42.67"N,
105°39'26.20"E
X. Zhao 1952 2019
27 O. ochrocephala Bunge Xinglong Mountain 35°47'5.17"N,
104°0'0.67"E
X. Zhao 1828 2018
28 O. ochrocephala Bunge Maxian Mountain 35°46'46.60"N,
103°59'19.33"E
X. Zhao 1953 2019
29 O. ochrocephala Bunge Jinqiang River 37°13'36.45"N,
102°41'3.46"E
X. Zhao 1840 2018
30 O. ochrocephala Bunge Hougou Village 35°48'47.34"N,
103°57'53.83"E
X. Zhao 1954 2019
31 O. qinghaiensis Y.H.Wu Labrang Monastery 35°11'8.91"N,
102°30'37.00"E
X. Zhao 1822 2018

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
54
dierences of individual numerical characteristics, and were applied to analyze dier-
ences in the numerical size of dimensions (Raymond and Sylvia 1993; Farhana and
Safwana 2018). e Ward error sum of squares method applies the concept of ANO-
VA to classication, resulting in richer clustering information that is rarely aected
by abnormal data (Ward 1963; Szekely and Rizzo 2005). In the present study, to test
the validity of the seed macro-and micromorphological traits, PCA was used to select
taxonomically relevant qualitative and quantitative characteristics. It is usually used to
distinguish between species within a given genus.
Results
Seed morphology
e studied seeds, all from the genus Oxytropis, had two main types of hilum positions,
terminal and central, and ve dierent types of seed shapes: prolonged semielliptic,
reniform, prolonged reniform, quadratic, and cardiform (Table 2; Figs 1, 2). Hilum
position was observed as terminal in O. racemosa, O. neimonggolica, O. imbricata (LC,
TR), O. coerulea (TT, EDG), O. xinglongshanica (MX, XL), O. glabra, O. taochensis,
and O. ochrocephala (NH, XL, MX, JQ, and HG). Hilum position was observed as
central in O ciliata, O. muricata (MX, TM), O. falcata (AWC, MQ), O. ochrantha
(XL, NMP), O. bicolor (U, TM), O. myriophylla (EDG, AG, and MX), O. holan-
shanensis, O. kansuensis (AZ, CT), O. qinghaiensis, O. latibracteata, O. qilianshanica,
O.aciphylla, and O. squammulosa. In addition, seed shapes could be separated into
ve groups (Table 2): a cardiform seed was found in O ciliata, O. muricata (MX, TM),
O.falcata (AWC, MQ), O. ochrantha (XL, NMP), O. bicolor (U, TM), O. holanshan-
ensis, O.kansuensis (AZ, CT), and O. squammulosa (Table 2; Figs 1, 2); a prolonged
reniform seed was observed in O. racemosa, O. neimonggolica, O. imbricata (LC, TR),
O. coerulea (TT, EDG), O. xinglongshanica (MX, XL), O. glabra, and O. taochensis
(Table 2; Figs 1, 2); a reniform seed was found in O. myriophylla (EDG, AG, MX),
O.qinghaiensis, O. latibracteata, and O. qilianshanica; a quadratic seed was only found
in O. ochrocephala (NH, XL, MX, JQ, HG); and nally, a prolonged semielliptic seed
was only found in O. aciphylla (Table 2; Fig. 2).
e seeds ranged from 1.27 mm (O. kansuensis (AZ)) to 2.57 mm (O. coerulea
(EDG)) in length and from 1.18 mm (O. qinghaiensis) to 2.02 mm (O. holanshanensis)
Section Code Species Locality Coordinates Voucher Collection
date
Section Oxytropis 32 O. latibracteata Jurtz. Helan Mountain 38°39'46.59"E,
105°49'20.25"N
X. Zhao 1951 2019
33 O. qilianshanica
C.W.Chang & C.L.Zhang
Jinqiang River Unknown J.Q. Wang 710113 Unknown
Section Lycotriche 34 O. aciphylla Ledeb. Jiji Spring Nature
Reserve
38°59'43"N,
101°55'39"E
X. Zhao 1924 2019
Section Leucopodia 35 O. squammulosa Candolle Shaochagou 35°42'57.20"N,
105°2'21.20"E
X. Zhao 1928 2019

Seed morphology of Oxytropis DC. 55
Table 2. Seed morphological features of Oxytropis under scanning electron microscopy.
Section Code Species Shape of seed Sculpturing Hilum position
Section Xerobia 1O ciliata Cardiform Scaled Central
Section Polyadena 2O. muricata (MX) Cardiform Rugulate Central
3O. muricata (TM) Cardiform Rugulate Central
Section Falcicarpae 4O. falcata (AWC) Cardiform Rugulate Central
5O. falcata (MQ) Cardiform Rugulate Central
Section Baicalia 6O. ochrantha (XL) Cardiform Lophate with stellated testa cells Central
7O. ochrantha (NMP) Cardiform Rugulate Central
8O. bicolor (U) Cardiform Rough Central
9O. bicolor (TM) Cardiform Simple reticulate Central
10 O. racemosa Prolonged Reniform Rough Terminal
11 O. myriophylla (EDG) Reniform Rugulate Central
12 O. myriophylla (AG) Reniform Rough Central
13 O. myriophylla (MX) Reniform Rugulate Central
Section Neimonggolicae 14 O. neimonggolica Prolonged Reniform Scaled Terminal
Section Eumorpha 15 O. imbricata (LC) Prolonged Reniform Rugulate Terminal
16 O. imbricata (TR) Prolonged Reniform Rough Terminal
17 O. coerulea (TT) Prolonged Reniform Rugulate Terminal
18 O. coerulea (EDG) Prolonged Reniform Rugulate Terminal
19 O. holanshanensis Cardiform Compound reticulate Central
Section Mesogaea 20 O. xinglongshanica (MX) Prolonged Reniform Lophate with stellated testa cells Terminal
21 O. xinglongshanica (XL) Prolonged Reniform Lophate with stellated testa cells Terminal
22 O. glabra Prolonged Reniform Rugulate Terminal
23 O. kansuensis (AZ) Cardiform Rugulate Central
24 O. kansuensis (CT) Cardiform Rugulate Central
25 O. taochensis Prolonged Reniform Lophate with stellated testa cells Terminal
26 O. ochrocephala (NH) Quadratic Rugulate Terminal
27 O. ochrocephala (XL) Quadratic Rugulate Terminal
28 O. ochrocephala (MX) Quadratic Rugulate Terminal
29 O. ochrocephala (JQ) Quadratic Rugulate Terminal
30 O. ochrocephala (HG) Quadratic Rugulate Terminal
31 O. qinghaiensis Reniform Compound reticulate Central
Section Oxytropis 32 O. latibracteata Reniform Rugulate Central
33 O. qilianshanica Reniform Rough Central
Section Lycotriche 34 O. aciphylla Prolonged Semielliptic Simple reticulate Central
Section Leucopodia 35 O. squammulosa Cardiform Lophate with rounded testa cells Central
in width (Table 3). e lowest length/width ratio (0.89) was observed in O. ochroceph-
ala (JQ), while the highest (1.55) was found in O. imbricata (LC). e lightest seeds
were measured in O. qinghaiensis at 0.1058 g, while the heaviest seeds were measured
in O. ciliata at 0.3521 g (Table 3).
Surface sculpturing
Seven dierent seed surface sculpturing patterns were observed: scaled, regulate, lophate
with stellated testa cells, simple reticulate, rough, compound reticulate, and lophate
with rounded testa cells (Table 2; Figs 3, 4). e regulate sculpturing pattern was
common in most taxa and was predominant in O. muricata (MX, TM), O. falcata
(AWC, MQ), O. ochrantha (NMP), O. myriophylla (EDG, MX), O. kansuensis (AZ,
CT), O. latibracteata, O. imbricata (LC), O. coerulea (TT, EDG), O. glabra, and

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
56
Figure 1. Seed shape of the studied species A O. ciliata B O. muricata (MX) C O. muricata (TM)
DO.falcata (AWC) E O. falcata (MQ) F O. ochrantha (XL) G O. ochrantha (NMP) H O. bicolor (U)
IO.bicolor (TM) J O. racemosa K O. myriophylla (EDG) L O. myriophylla (AG) M O. myriophylla (MX)
NO. neimonggolica O O. imbricata (LC) P O. imbricata (TR). Scale bar: 500 μm.
O.ochrocephala (NH, XL, MX, JQ, and HG) (Table 2; Figs 3, 4). e simple reticulate
sculpturing pattern was predominant in O. bicolor (U) and O. aciphylla, while the
compound reticulate sculpturing pattern was predominant in O. holanshanensis and
O.qinghaiensis (Table 2; Figs 3, 4). e scaled sculpturing pattern was predominant in
O. ciliata and O.neimonggolica, while the rough sculpturing pattern was predominant

Seed morphology of Oxytropis DC. 57
Figure 2. Seed shape of the studied species A O. coerulea (TT) B O. coerulea (EDG) C O. holanshanensis
D O. xinglongshanica (MX) E O. xinglongshanica (XL) F O. glabra G O. kansuensis (AZ) H O. kansuensis
(CT) I O. taochensis J O. ochrocephala (NH) K O. ochrocephala (XL) L O. ochrocephala (MX) M O.ochro-
cephala (JQ) N O. ochrocephala (HG) O O. qinghaiensis P O. latibracteata Q O. qilianshanica R O.aci-
phylla S O. squammulosa. Scale bar: 500 μm.

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
58
Table 3. Seed morphological features of Oxytropis under stereoscopic microscopy.
Section Code Species Length Min. (mean
± SD) max./mm
Width Min. (mean ±
SD) max./mm
L/W ratio Seed weight/g
Section Xerobia 1O ciliata 1.54(2.43±0.36)3.28 1.34(2.05±0.28)2.8 1.19±0.07 0.3521±0.0236
ection Polyadena 2O. muricata (MX) 1.38(2.02±0.3)2.91 1.1(1.78±0.26)2.38 1.14±0.07 0.2627±0.0041
3O. muricata (TM) 1.41(2.04±0.29)2.66 1.15(1.66±0.23)2.03 1.24±0.18 0.248±0.013
Section Falcicarpae 4O. falcata (AWC) 1.59(2.14±0.25)2.79 1.42(1.87±0.18)2.35 1.15±0.13 0.2981± 0.0106
5O. falcata (MQ) 1.7(2.09±0.14)2.5 1.45(1.86±0.17)2.15 1.13±0.13 0.323±0.005
Section Baicalia 6O. ochrantha (XL) 1.29(1.65±0.17)2.09 1.18(1.53±0.16)2 1.07±0.07 0.2148±0.0091
7O. ochrantha (NMP) 1.16(1.49±0.17)1.84 1.15(1.35±0.12)1.6 1.1±0.07 0.1732±0.0021
8O. bicolor (U) 1.09(1.57±0.23)1.98 1.21(1.49±0.17)1.88 1.05±0.06 0.146±0.013
9O. bicolor (TM) 1.32(1.74±0.21)2.31 1.23(1.65±0.23)2.32 1.06±0.08 0.1326±0.0086
10 O. racemosa 1.24(1.71±0.17)2.19 0.77(1.23±0.14)1.5 1.4±0.12 0.1668±0.0128
11 O. myriophylla (EDG) 1.05(1.56±0.21)2.14 0.81(1.25±0.19)1.64 1.26±0.1 0.1290±0.0004
12 O. myriophylla (AG) 1.26(1.59±0.13)1.93 0.94(1.32±0.16)1.63 1.22±0.09 0.1231±0.0007
13 O. myriophylla (MX) 1.06(1.67±0.23)2.1 1.04(1.45±0.16)1.9 1.15±0.07 0.1349±0.0043
Section Neimonggolicae 14 O. neimonggolica 1.85(2.11±0.12)2.32 1.54(1.72±0.12)1.98 1.23±0.04 0.326±0.01
Section Eumorpha 15 O. imbricata (LC) 1.77(2.44±0.27)2.93 1.1(1.59±0.2)2.05 1.54±0.11 0.3188±0.0054
16 O. imbricata (TR) 1.79(2.36±0.31)3.13 1.08(1.56±0.27)2.34 1.52±0.12 0.3264±0.0112
17 O. coerulea (TT) 1.66(2.39±0.25)2.96 1.22(1.69±0.21)2.11 1.43±0.12 0.2799±0.0016
18 O. coerulea (EDG) 1.99(2.57±0.16)2.92 1.44(1.86±0.15)2.2 1.39±0.09 0.2986± 0.0013
19 O. holanshanensis 1.81(2.21±0.19)2.66 1.43(2.02±0.22)2.58 1.1±0.09 0.3264±0.0062
Section Mesogaea 20 O. xinglongshanica (MX) 1.56(2.32±0.32)3.22 1.29(1.93±0.29)2.69 1.21±0.1 0.2914±0.0038
21 O. xinglongshanica (XL) 1.47(2.23±0.23)2.7 1.42(1.77±0.16)2.24 1.26±0.11 0.2763±0.0103
22 O. glabra 0.93(1.78±0.33)2.63 0.84(1.53±0.27)2.23 1.16±0.09 0.1892±0.0066
23 O. kansuensis (AZ) 0.87(1.27±0.2)1.9 0.91(1.28±0.17)1.69 0.99±0.09 0.1074±0.0057
24 O. kansuensis (CT) 0.95(1.38±0.13)1.65 1.05(1.44±0.17)1.77 0.97±0.11 0.1260±0.0044
25 O. taochensis 1.54(2.09±0.25)2.73 1.08(1.55±0.18)1.89 1.36±0.11 0.2236±0.0134
26 O. ochrocephala (NH) 1.22(1.73±0.22)2.23 1.35(1.91±0.23)2.53 0.9±0.06 0.2719±0.0043
27 O. ochrocephala (XL) 1.1(1.63±0.17)2.03 1.23(1.77±0.24)2.39 0.92±0.07 0.2517±0.0103
28 O. ochrocephala (MX) 1.28(1.64±0.17)2.01 1.56(1.82±0.12)2.14 0.9±0.07 0.2417±0.0065
29 O. ochrocephala (JQ) 0.92(1.56±0.23)2.06 1.23(1.75±0.21)2.43 0.89±0.08 0.2506±0.0098
30 O. ochrocephala (HG) 1.02(1.66±0.26)2.28 1.43(1.8±0.14)2.27 0.92±0.11 0.2854±0.0123
31 O. qinghaiensis 1.2(1.56±0.18)1.99 0.93(1.18±0.11)1.56 1.33±0.1 0.1058±0.0087
Section Oxytropis 32 O. latibracteata 1.5(2.05±0.25)2.64 1.2(1.69±0.21)2.19 1.22±0.1 0.2368±0.0106
33 O. qilianshanica 1.38(1.57±0.08)1.71 1.09(1.31±0.1)1.49 1.2±0.05 0.112±0.008
Section Lycotriche 34 O. aciphylla 1.36(1.99±0.28)2.81 1.01(1.43±0.21)1.98 1.39±0.12 0.1822±0.0094
Section Leucopodia 35 O. squammulosa 1.22(1.81±0.25)2.61 0.95(1.62±0.29)2.37 1.13±0.09 0.2070±0.0117
in O. bicolor, O.myriophylla (AG), O. qilianshanica, O. racemosa, and O. imbricata (TR)
(Table 2; Figs 3, 4). Lastly, the lophate pattern with stellated testa cells was predominant
in O.ochrantha (XL), O.xinglongshanica (MX, XL), and O. taochensis, while the lophate
pattern with rounded testa cells was only found in O. squammulosa (Table 2; Figs 3, 4).
Numerical analysis
In the present study, principal components analysis (PCA) indicates three groups of
traits, which explain 82.81% of the total variation (Table 4). e rst principal compo-
nent (PC1) exhibited 41.51% of the variability, which had a high loading component
of the seed length, width, and weight. e second PC (PC2) accounted for 22.18% of
the variation and was strongly associated with L/W ratio and sculpturing, whereas the

Seed morphology of Oxytropis DC. 59
Figure 3. Seed surface sculpturing of the studied species A O. ciliata B O. muricata (MX) C O. muricata
(TM) D O. falcata (AWC) E O. falcata (MQ) F O. ochrantha (XL) G O. ochrantha (NMP) H O. bicolor
(U) I O. bicolor (TM) J O. racemosa K O. myriophylla (EDG) L O. myriophylla (AG) M O. myriophylla
(MX) N O. neimonggolica O O. imbricata (LC) P O. imbricata (TR). Scale bar: 5 μm.
third PC (PC3) contained 19.12% of the variability in which hilum position and seed
shape were important. As shown in Fig. 5, the scatter points for the same species are
closely aggregated, such as the ve samples of O. ochrocephala (NH, XL, MX, JQ, and
HG), indicating that samples from dierent populations within the same species had

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
60
Figure 4. Seed surface sculpturing of the studied species A O. coerulea (TT) B O. coerulea (EDG)
CO.holanshanensis D O. xinglongshanica (MX) E O. xinglongshanica (XL) F O. glabra G O. kansuensis
(AZ) H O. kansuensis (CT) I O. taochensis J O. ochrocephala (NH) K O. ochrocephala (XL) L O. ochro-
cephala (MX) M O. ochrocephala (JQ) N O. ochrocephala (HG) O O. qinghaiensis P O. latibracteata
QO.qilianshanica R O. aciphylla S O. squammulosa. Scale bar: 5 μm.

Seed morphology of Oxytropis DC. 61
Table 4. PCA variable loading characters of seed micro-morphology of studied Oxytropis species.
PCA variable loadings PC1 PC2 PC3
Shape of seed -0.01682 -0.32175 0.7042
Sculpturing -0.24237 0.44835 0.07685
Hilum position 0.33478 -0.18679 0.5441
Length 0.50744 0.37518 0.00087
Width 0.48498 -0.23663 -0.28718
L/W ratio 0.14335 0.68091 0.32905
Seed weight 0.56183 -0.01324 -0.10682
Eigenvalue 2.90597 1.55275 1.33831
Variability/% 41.51384 22.18208 19.11869
Cumulative/% 41.51384 63.69592 82.81461
Figure 5. PCA for 35 samples belonging to 21 Oxytropis species based on seed morphological characters.
Dots of dierent colors represent dierent species, and dots of the same color represent dierent popula-
tions of the same species.
similar characteristics. However, the arrangement of 21 species belonging to 10 sec-
tions does not show a certain regularity. For example, species belonging to dierent sec-
tions are also arranged together, indicating that the seed morphological characteristics
of Oxytropis species does not have regularity within the section. Cluster analysis reects
the similarity among species based on the anatomical characteristics and delimitation

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
62
of these groups. Our phenograms of the quantitative and qualitative data showed three
primary clusters (Fig. 6). e rst cluster included O ciliata, O.muricata (MX, TM),
O. falcata (AWC, MQ), O. holanshanensis, O. neimonggolica, O.xinglongshanica (MX,
XL), O. imbricata (LC), O. coerulea (TT, EDG), and O. imbricata (TR). e second
cluster only contained O. ochrocephala (NH, XL, MX, JQ, and HG). e third clus-
ter included O. ochrantha (XL, NMP), O. kansuensis (AZ, CT), O. bicolor (U, TM),
O.squammulosa, O. racemosa, O. myriophylla (AG), O. qilianshanica, O.qinghaiensis,
O. myriophylla (EDG, MX), O. glabra, O. taochensis, O. latibracteata, and O. aciphylla.
Discussion
Seed morphology of the investigated species was determined for the rst time in the
present study. Seed characteristics, such as coat pattern, shape, and size, have been
shown to be important for the classication within genera of Fabaceae species (Lersten
and Gunn 1981; Solum and Lockerman 1991; López et al. 2000; Al-Gohary and
Mohamed 2007; Salimpour et al. 2007; Vural et al. 2008; Venora et al. 2009; Zorić et
al. 2010; Celep et al. 2012; De-Paula and Oliveira 2012; Kaya and Dirmenci 2012;
Lantieri et al. 2013). Previous studies have shown that seed shape and hilum position
are taxonomically signicant and can therefore be used for the classication of taxa
at the genus or even species level (López et al. 2000; Salimpour et al. 2007; Vural
et al. 2008). e ve main types of seed shapes observed in the present study were
consistent with previous ndings on Oxytropis (Erkul et al 2015). e seed shapes of
Figure 6. Phenogram for 35 samples belonging to 21 Oxytropis species based on based on seed morpho-
logical characters.

Seed morphology of Oxytropis DC. 63
dierent populations of the same Oxytropis species were highly consistent, indicating
that they were relatively constant within species. Particularly, O. ochrocephala and O.
kansuensis are easily confused, as they are morphologically dicult to distinguish and
are both abundant in the northwest China (Zhu et al. 2010). However, our obser-
vations demonstrate that these two species can be distinguished based on their seed
shape; O. ochrocephala has a quadratic seed, whereas O. kansuensis has a cardiform
seed. ese results indicate that seed shape might be a useful taxonomic marker for
some Oxytropis species. However, similar seed shapes exist in other species of the genus
Oxytropis and other groups of Fabaceae (Erkul et al. 2015). us, they should be con-
sidered in combination with other macro-morphological characteristics when applied
to species identication within the genus Oxytropis.
e sculpturing pattern of seeds is thought to provide useful information for the
infrageneric classication of some genera of Fabaceae (Salimpour et al. 2007; Vural
et al. 2008; Kamala and Aydin 2018; Rashid et al. 2020). Farrington et al. (2008)
proposed that Oxytropis seed coat micromorphology and anatomy can be used to dis-
tinguish Oxytropis from its sister taxon, Astragalus. However, studies have shown that
the taxonomic value of seed sculpturing patterns in Astragalus and Oxytropis species
is limited. For example, a study that examined 48 species of Turkish Astragalus found
only two distinct seed coat morphological types (rugulate and rugulate-reticulate) (Vu-
ral et al. 2008). Similarly, Shemetova et al. (2018) recognised two main types of seed
surface in the genus Astragalus: reticulate and indistinct primary sculpture. However,
these seed sculpturing patterns have also been observed in the genus Oxytropis. Far-
rington et al. (2008) found that Alaskan Oxytropis (15 taxa) has rugulate, rugulate-
reticulate, and lophate sculpturing patterns. Consistently, Erkul et al. (2015) reported
three types of seed sculpturing patterns in Oxytropis, namely rugulate, rugulate-retic-
ulate, and lophate, and proposed that seed characteristics are not useful for separating
the genera Oxytropis and Astragalus. Furthermore, Meyers et al. (2013) proposed that
seed coat types among the Alaskan members of Oxytropis are highly variable at the spe-
cies level and cannot be used for species identication. Our results supported this hy-
pothesis because seed sculpturing patterns are variable within some species, including
O.ochrantha (XL, NMP), O. bicolor (U, TM), O. myriophylla (EDG, AG, and MX),
and O.imbricata (LC, TR), suggesting that seed sculpturing pattern has a limited taxo-
nomic value. Interestingly, in the present study, the seed sculpturing pattern appeared
to be conserved dierently within dierent sections. Seed coat patterns were stable
within some species in the section Mesogaea, such as O. ochrocephala, O. kansuensis,
and O. xinglongshanica, but highly variable in the species of the sections Baicalia and
Eumorpha. erefore, the taxonomic signicance of seed sculpturing pattern should be
comprehensively analysed using a broader sample.
Previous studies on Oxytropis have suggested that seed characteristics, such as size
(length, width, and length/width ratio), shape, surface sculpturing, and weight have
low taxonomic value at the infrageneric level (Solum and Lockerman 1991; Bojňanský
and Fargašová 2007; Farrington et al. 2008; Meyers et al. 2013; Erkul et al. 2015).
However, most of these studies only subjectively compared their quantitative traits
without a systematic analysis such as a cluster analysis. Only Erkul et al. (2015)

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
64
systematically analysed the seed traits in 13 Oxytropis species from Turkey, but they did
not explore the variation in seed traits at the species level because of sampling limita-
tions. In the present study, the results of the cluster analysis showed that, except for
O. myriophylla, dierent populations of the same species were clustered into one clade,
indicating that the seed traits of Oxytropis are useful for the identication of taxa at the
species level. However, species belonging to dierent sections were present in the same
clade, indicating that seed characteristics have low taxonomic value at the section level.
e results of the PCA also supported the former view that populations within the
same species cluster together, while the distribution of samples of dierent species does
not show a certain regularity. Furthermore, the rst PC of the PCA provided a highly
dominant variability of 41.51%, the characteristics with major scores that contributed
to the formation of the groups were quantitative characteristics, such as length, width,
and weight of seed. e second and third PCs are mainly qualitative characteristics,
accounting for 41.3% of the total variance. ese results suggest that even though
quantitative traits and some qualitative traits, such as seed sculpturing patterns, are
highly variable within species, these traits still play an important role in systematic
analysis. erefore, it is necessary to comprehensively analyse qualitative and quantita-
tive characteristics in future research into Oxytropis seed morphology.
To date, a comprehensive phylogenetic study of the genus Oxytropis has not been
carried out. Moreover, even though several studies have utilized DNA barcodes such
as ITS, trnL-F, and psbA-trnH to investigate the molecular phylogeny of Oxytropis in
northwest China, the low genetic dierence of these barcodes among species makes it
dicult to distinguish species within this genus and solve the phylogenetic relation-
ship among its species (Li et al. 2011; Gao et al. 2013; Lu et al. 2014). erefore, the
phylogenetic reliability of seed traits in Oxytropis cannot be conrmed. More detailed
molecular phylogenetic studies and more extensive taxon sampling are needed to dis-
cover the correlation between seed features and genus taxonomy.
Conclusions
Our results suggest that the seed traits of Oxytropis are helpful for identifying taxa at
the species level, but have low taxonomic value at the section level. Seed shape was
constant within species and was useful for species delimitation in the genus Oxytropis
when combined with other macroscopic traits. e seed sculpturing patterns were
highly variable at the species level and could not be used for species identication.
Although quantitative traits and some qualitative traits, such as seed sculpturing pat-
terns, are highly variable within species, these traits still play an important role in
PCA and cluster analysis. e results of the PCA and cluster analysis showed that
dierent populations of the same species were clustered into one clade, indicating
that in Oxytropis, seed traits are useful for the identication of taxa at the species level.
However, species belonging to dierent sections also clustered into the same clade,
indicating that seed characteristics have low taxonomic value at the section level.

Seed morphology of Oxytropis DC. 65
Acknowledgements
is work was supported by the Gansu Key Research and Development Project-Agricul-
ture (grant number 18YF1NA051), Gansu Provincal Talent development Project (grant
number 20220401), and National Natural Science Foundation of China (32260054).
e authors declare that they have no known competing nancial interests or per-
sonal relationships that could have appeared to inuence the work reported in this paper.
References
Abusaief H, Boasoul SH (2021) A taxonomic study of twelve wild forage species of Fabaceae.
Heliyon 7(2): e06077. https://doi.org/10.1016/j.heliyon.2021.e06077
Al-Ghamdi FA (2011) Seed Morphology of Some Species of Indigofera (Fabaceae) from Saudi
Arabia (Identication of Species and Systematic Signicance). American Journal of Plant
Sciences 02(3): 484–495. https://doi.org/10.4236/ajps.2011.23057
Al-Gohary IH, Mohamed AH (2007) Seed morphology of Acacia in Egypt and its taxonomic
signicance. International Journal of Agriculture and Biology 9: 435–438.
Barneby RC (1952) A revision of the North American species of Oxytropis DC. San Francisco.
Proceedings of the California Academy of Sciences 4: 177–312.
Bojňanský V, Fargašová A (2007) Atlas of Seeds and Fruits of Central and East-European Flora:
e Carpathian Mountains region. Springer, e Netherlands.
Bunge A (1874) Species generis Oxytropis, DC. Mémoires de l’Académie impériale des sciences
de St.-Pétersbourg ser. 7, 22(1): 160.
Celep F, Koyuncu MA, Fritsch RM, Kahraman AC, Doǧan M (2012) Taxonomic importance
of seed morphology in Allium (Amaryllidaceae). Systematic Botany 37(4): 893–912.
https://doi.org/10.1600/036364412X656563
De Candolle P (1802) Astragalogia Nempe Astragali, Biserrulae et Oxytropidis, nec non Pha-
cae, Colutae et Lessertiae Historia Iconibus Illustrate [Astragalogia, namely Astragalus,
Biserrula and Oxytropis, or not, Phaca, Colutea and Lessertia, History icons illustared].
Joann Bapt Garnery, Paris, 324 pp. https://doi.org/10.5962/bhl.title.517
De-Paula OC, Oliveira DMT (2012) Seed ontogeny of Chamaecrista and its systematic im-
plications in Cassiinae (Leguminosae, Caesalpinioideae). Plant Systematics and Evolution
298(9): 1659–1669. https://doi.org/10.1007/s00606-012-0667-8
Erkul SK, Celep F, Aytaç, Z (2015) Seed morphology and its systematic implications for genus
Oxytropis DC. (Fabaceae). Plant Biosystems – An International Journal Dealing with all As-
pects of Plant Biology 149(5): 875–883. https://doi.org/10.1080/11263504.2014.969353
Farhana H, Safwana H (2018) Plant recognition system using leaf shape features and Minimum
Euclidean distance. Journal on Image and Video Processing 9(2): 1919–1925. https://doi.
org/10.21917/ijivp.2018.0272
Farrington R, Meyers Z, Ickert-Bond S (2008) A Comparative Study of Seed Coat Micromor-
phology and Seed Anatomy in Alaskan Oxytropis (Loco-weed, Fabaceae) using SEM, and
their taxonomic signicance. Poster Presentation. Vancouver, Canada.

Xiang Zhao et al. / PhytoKeys 222: 49–67 (2023)
66
Fayed AA, El-Hadidy AF, Faried A, Olwey A (2019) Taxonomic implications of multivariate
analyses of Egyptian Ononis L. (Fabaceae) based on morphological traits. Korean Journal of
Plant Taxonomy 49(1): 13–27. https://doi.org/10.11110/kjpt.2019.49.1.13
Gao LX, Lu P, Jin F, Gao J (2013) trnL-F sequences and molecular phylogeny of 10 species of
Oxytropis. Xibei Zhiwu Xuebao 33(2): 266–271. [In Chinese with English abstract]
Kamala VA, Aydin MA (2018) Micromorphological structure of seeds in some taxa of Vicia
L. (Fabaceae) occurring in Azerbaijan. Plant & Fungal Research 1(1): 43–51. https://doi.
org/10.29228/plantfungalres.34
Kaya A, Dirmenci T (2012) Nutlet morphology of Turkish Ziziphora L. (Lamiaceae). Plant Bio-
systems – An International Journal Dealing with all Aspects of Plant Biology 146: 560–563.
Lantieri A, Guglielmo A, Pavone P, Salmeri C (2013) Seed germination in Sarcopoterium spi-
nosum (L.) Spach from South-Eastern Sicily. Plant Biosystems – An International Journal
Dealing with all Aspects of Plant Biology 147: 60–63. https://doi.org/10.1080/1126350
4.2012.717549
Leins P, Merxmüller H (1968) Oxytropis DC. In: Tutin TG, Heywood VH, Burges NA, Moore
DM, Valentine DH, Walters SM, Webb DA, Chater AO, Ball PW, DeFilipps RA, Ferguson
IK, Richardson IBK (Eds) Flora Europaea. Cambridge University Press, Cambridge, 124–126.
Lersten NR, Gunn CR (1981) Seed Morphology and Testa Topography in Cicer (Fabaceae:
Faboideae). Systematic Botany 6(3): e223. https://doi.org/10.2307/2418283
Li YF, Lan FR, Chang ZY, Guo ZK (2011) Molecular phylogeny of Oxytropis of Qinghai-
Tibetan Plateau by ITS and trnL-F sequences. Xibei Nong-Lin Keji Daxue Xuebao. Ziran
Kexue Ban 39(11): 188–193. [In Chinese with English abstract]
López J, Devesa JA, Ortega‐Olivencia A, Ruiz T (2000) Production and morphology of fruit
and seeds in Genisteae (Fabaceae) of south-west Spain. Botanical Journal of the Linnean
Society 132(2): 97–120. https://doi.org/10.1111/j.1095-8339.2000.tb01208.x
Lu P, Gao LX, Jin F (2014) Molecular Phylogeny of Ten Species of Oxytropis Based on psbA-trnH
Sequences. Plant Diversity and Resources 36(3): 279–284. [In Chinese with English abstract]
Meyers ZJ, Ickert-Bond SM, LaMesjerant R (2013) A survey of seed coat morphology in Oxy-
tropis, Sects. Arctobia, Baicaua, Glaeocephala, Mesogaea, and Orobia (FABACEAE) from
Alaska. Journal of the Botanical Research Institute of Texas 7(1): 391–404. http://www.
jstor.org/stable/24621090
OriginLab Corporation (2022) Origin (Pro) (Version 2022, Learning edition). Northampton.
Pavlov NV (1961) Flora Kazakhstana. Nauka Kazakhskoĭ SSR. Alma-Ata 5: 330–441.
Polhill RM (1981) Tribe 16. Galegeae (Bronn). In: Polhill RM, Raven PH (Eds) Advances in
Legume Systematics, part 1. e Royal Botanic Gardens, Kew, 357–363.
Rashid N, Zafar M, Ahmad M, Malik K, Haq IU, Shah SN, Mateen A, Ahmed T (2018)
Intraspecic variation in seed morphology of tribe vicieae (Papilionoidae) using scanning
electron microscopy techniques. Microscopy Research and Technique 81(3): 298–307.
https://doi.org/10.1002/jemt.22979
Rashid N, Zafar M, Ahmad M, Memon RA, Akhter MS, Malik K, Malik NZ, Sultana S, Shah
SN (2020) Seed morphology: An addition to the taxonomy of Astragaleae and Trifolieae
(Leguminosae: Papilionoidae) from Pakistan. Microscopy Research and Technique 84(5):
1053–1062. https://doi.org/10.1002/jemt.23666

Seed morphology of Oxytropis DC. 67
Raymond DR, Sylvia RM (1993) Squared Euclidean Distance: a Statistical Test to Evaluate
Plant Community Change. Research Note, 5 pp. https://doi.org/10.2737/PSW-RN-416
Salimpour F, Mostafavi G, Sharifnia F (2007) Micromorphologic study of the seed of the
genus Trifolium, section Lotoidea, in Iran. Pakistan Journal of Biological Sciences 10(3):
378–382. https://doi.org/10.3923/pjbs.2007.378.382
Shemetova T, Erst AS, Wang W, Xiang K, Vural C, Aytaç Z (2018) Seed morphology of the
genus Astragalus L. from North Asia. Turkish Journal of Botany 42(6): 710–721. https://
doi.org/10.3906/bot-1802-25
Singh G (2019) Numerical and Cladistic Methods. In: Singh G (Ed.) Plant Systematics: An
Integrated Approach, Fourth Edition. CRC Press, New Delhi, 163 pp.
Solum DJ, Lockerman RH (1991) Seed coat surface patterns and structures of Oxytropis ri-
paria, Oxytropis campestris, Medicago sativa, and Astragalus cicer. Scanning Microscopy 5:
779–786.
Szekely G, Rizzo M (2005) Hierarchical Clustering via Joint Between-Within Distances: Ex-
tending Ward’s Minimum Variance Method. Journal of Classication 22(2): 151–183.
https://doi.org/10.1007/s00357-005-0012-9
ien LB, Heimermann WH, Holman RT (1975) Floral odors and quantitative taxonomy of
Magnolia and Liriodendron. Taxon 24(5–6): 557–568. https://doi.org/10.2307/1220717
Vassilczenko IT (1948) Oxytropis DC. In: Komarova VL, Boris KS (Eds) Flora of the USSR.
Izd-vo Akademii nauk SSSR, Moskow-Leningrad, 229 pp.
Venora G, Grillo O, Ravalli C, Cremonini R (2009) Identication of Italian landraces of bean
(Phaseolus vulgaris L.) using an image analysis system. Scientia Horticulturae 121(4): 410–
418. https://doi.org/10.1016/j.scienta.2009.03.014
Vural C, Ekici M, Akan H, Aytaç Z (2008) Seed morphology and its systematic implications for
genus Astragalus L. sections Onobrychoidei DC., Uliginosi Gray and Ornithopodium Bunge
(Fabaceae). Plant Systematics and Evolution 274(3–4): 255–263. https://doi.org/10.1007/
s00606-008-0025-z
Ward Jr JH (1963) Hierarchical Grouping to Optimize an Objective Function. Journal of the
American Statistical Association 58(301): 236–244. https://doi.org/10.1080/01621459.1
963.10500845
Welsh SL (2001) Revision of North American species of Oxytropis de Candolle (Leguminosae).
EPS Inc., Orem.
Wojciechowski MF (2005) Astragalus (Fabaceae): A molecular phylogenetic perspective. Brittonia
57(4): 382–396. https://doi.org/10.1663/0007-196X(2005)057[0382:AFAMPP]2.0.CO;2
Zhang CW (1998) Oxytropis DC. In: Editing Committee of Chinese Flora (Eds) Flora Reipub-
licae Polularis Sinicae. Science Press, Beijing, 146 pp. [In Chinese]
Zhu XY, Ohashi H (2000) Systematics of Chinese Oxytropis DC. (Leguminosae). In: Hong DY
(Ed.) Cathaya 11–12. International Academic Publishers, Beijing, 218 pp.
Zhu XY, Welsh SL, Ohashi H (2010) Oxytropis DC. In: Editorial Committee of Flora of China
(Eds) Flora of China. Science Press & Missouri Botanical Garden Press, Beijing & St.
Louis, 453–500.
Zorić L, Merkulov L, Luković J, Boza PP (2010) Comparative seed morphology of Trifolium L.
Species (Fabaceae). Periodicum Biologorum 112: 263–272.