African Journal of
Plant Science

  • Abbreviation: Afr. J. Plant Sci.
  • Language: English
  • ISSN: 1996-0824
  • DOI: 10.5897/AJPS
  • Start Year: 2007
  • Published Articles: 779

Full Length Research Paper

Morphological studies on legume and hilum in seven species of Vigna Savi (Fabaceae)

Ali El Saied Abdelatif Gaafar
  • Ali El Saied Abdelatif Gaafar
  • Department of Botany, Faculty of Science, New Valley University, 72511 El- Kharga, Egypt.
  • Google Scholar


  •  Received: 26 September 2020
  •  Accepted: 19 November 2020
  •  Published: 31 December 2020

 ABSTRACT

In this study of different taxa of genus Vigna the macro and micromorphological characters were investigated. These taxa collected from different countries (3 cultivated and 8 annual or perennial herbs) representing 7 species of this genus, namely: Vigna radiate, Vigna trilobata, Vigna vexillata, Vigna caracalla, Vigna pubescens, Vigna unguiculata, and Vigna luteola. Legume features such as shape, colour, size, surface texture, number of locules, number of legumes per peduncle seed set percentage, orientation. Moreover, by using scanning electron microscopy, additional details were obtained for stomatal shape, size, type, cuticular and wax ornamentation of the surface of the mature pod, including details of the seed such as hilum shape, size, position, ornamentation based on scanning electron microscope evidence. The usefulness of the macro and micromorphological features as criteria for taxonomic identification was emphasized. 

 

Key words: Vigna, Fabaceae, morphological characters, pod, hilum, S.E.M., taxonomy.


 INTRODUCTION

Fabaceae Lindley is the largest of the third largest family of flowering plants, with ca. 19,500 species in 770 genera (Polhill and Raven, 1981; Christenhusz and Byng, 2016; LPWG, 2017). The genus Vigna Savi belongs to the tribe Phaseoleae DC., subtribe Phaseolinae Benth. (sub family Faboideae, family Fabaceae) widely cultivated with ca. 200 species distributed in tropical and sub-tropical regions (Fery, 2002; Gaafar, 2007; El-Ghamery et al., 2012). Previous studies of this genus based on morphological characters were made by Verdcourt (1970), Maréchal et al. (1978, 1981), Ng and Maréchal (1985), Pasquet (1993), Tomooka et al. (2002, 2003), Ng (1990), and GRIN (2005). The genus is divided into 7 subgenera: Ceratotropis (Piper) Verdcourt, Haydonia (R. Wilczek) Verdcourt, Plectrotropis (Schumach.) Baker., Sigmidotropis (Piper) Verdcourt, Lasiosporon (Piper) Benth., Macrodonta Verdcourt and Vigna with 16 sections and containing 81 species.
 
Economically Vigna is a source of plant protein for human food and animal feed in tropical, sub-tropical, arid and semi-arid regions, and also plays an important role in soil fertility by fixing nitrogen (Mbagwu and Endeoga, 2006; Pule-Meulenberg et al., 2010; Sprent et al., 2010; El-Ghamery et al., 2012; El-Gazzar et al., 2013; Popoola et al., 2015). Barthlott and Ehler (1977) maintained that the epidermal features are variable for angiosperm taxa and can be used to evaluate possible relationships. Seed morphological characters are useful in the analysis of of taxonomic distinguish inter and intra relationships in a wide variety of plant families (Esau, 1953; Shetler and Morin, 1986; Takhtajan, 1991; Buss et al., 2001; Zhang et al.,   2005;    Gontcharova   et  al.,  2009).  In addition to general characters of morphology of seeds, sculpturing details of outer seed coat are quite variable between different species and can be of a systematic importance (Chowdhury and Buth, 1970; Gohary and Mohammed, 2007). The importance of ultrastructural pattern analysis of the seed coat observed under the SEM has been well recognized as a reliable approach for assessing phenetic relationship and identification of species or taxa (Barthlott, 1981; Tobe et al., 1987; Koul et al., 2000; Yoshizaki, 2003; Javadi and Yamaguchi, 2004).
 
The micro-morphological characters of the seed and legume surface can be useful for delimiting taxa at various levels (Karcz et al., 2005; Akçin, 2008; Khafagi et al., 2018). Legume colour that is less affected by environmental variations has been used as a marker for the identification of species or varieties (Sangwan and Lodhi, 1998). In Vigna species, many taxonomists have used seed and hilum morphology and micromorphology to differentiate the species (Chandel et al., 1991; Nath and Dasgupta, 2015; Umdale et al., 2017). This work aims to describe the significance of macro and micromorphological characters of legume and hilum using SEM in 11 taxa of the genus Vigna and its implication in the classification of this plant.


 MATERIALS AND METHODS

Seeds of Vigna Savi accessions were obtained from the International Livestock Center for Africa (ILCA), Jazan Region of Saudia Arabia and from different localities in Egypt. Eleven taxa representing 11 species belonging to genus Vigna Savi were investigated. The sources and origins of these taxa are given in Table 1. Seeds were germinated in April 2019, at the Botanic Gardens of the Botany Department, Faculty of Science, New valley University, Egypt. Flowering began in June 2019. Samples of complete mature plants were collected, including fruits and seeds. Authentic specimens of all taxa were pressed, dried and deposited as herbarium specimens at the Botany Department, Faculty of Science, New Valley University. The morphological characters were examined,   which included  legume  shape,  colour,  size, texture, apex, shattering, seed per pod, locule per pod, seed set percentages, pod attachment, number of pods per peduncle, and stomata on the surface of a mature pod. Hilum shape, size, colour and position features of different species were also noted. The seed set percentage was calculated using the formula described by Popoola et al. (2015) as follows:
 
 
 
For preparation of legume and seeds of each taxon to scan the surface by using the scanning electron microscope (SEM), legumes and seeds from each taxon were divided into two groups. The first group was mounted on an adhesive surface and prepared for SEM investigation. The second group was washed thoroughly using distilled water to remove any impurities on the seed surfaces; then they were dried and soaked in 10% HCl for 6 h to remove the coat enveloping the seed (Ismail and El-Ghazaly, 1990). This was followed by washing the seeds with distilled water and then dried and prepared for SEM examination by mounting these dry legumes and seeds onto clean stubs using double-sided adhesive tape.  These clean dry legumes and seeds were gold coated using a JEOL JFC 1100E ion-sputtering device. Then, the coated seeds were viewed and photographed with a JOEL ISM-5500LV scanning electron microscope, operated at accelerating voltage of 15 KV at the Scanning Electron Microscopy Unit in the Regional Center for Mycology and Biotechnology, Al-Azhar University. Magnifications of images were denoted with scale bars either in mm or µm as appropriate. Measurements (L x B) of legumes and seeds were taken using a measuring scale. Seeds were uniformly scanned at the hilum, the surface surrounding both sides of the hilum, and the surface pattern was highlighted to observe the cellular and intercellular patterns. The terminology for describing seed coat patterns follows Barthlott (1981, 1990) and Stearn (1996). Details of the species used are presented in Table 1.


 RESULTS

Legume
 
The legume morphological characters of the studied taxa are given in Table 2, represented in Plate 1 (1a, 2a,  3a, 4a and 5a) and Plate 2 (6a, 7a, 8a, 9a, 10a and 11a).
 
 
 
Legume shattering, shape and apex  
 
The results in Table 2 show all studied Vigna spe-cies have a shattering (dehiscent) legume, except Vigna unguiculata, Vigna caracalla and Vigna radiata (No.2) collected from Egypt. The time of legume shattering is related to the maturity of the legumes; also all studies species were of the linear-terte legume shape, except V. unguiculata that is slightly curved and V. caracalla that is linearly flat. The apex of legume is straight beak in the subgenus Ceratotropis. There is declined beak in the subgenus Plectrotropis and subgenus Vigna and a hooked beak in the subgenus Sigmidotropis (Table 2, Plate 1 (1a, 2a, 3a, 4a, 5a) and Plate 2 (6a, 7a, 8a, 9a, 10a and 11a).
 
 
Colour
 
Legume colour at maturity varied from black, brown, brownish black, gray to creamy. It was black black in V. radiata, Vigna trilobata and Vigna pubescens collected from Ethiopia, while brown in V. radiata collected from Egypt and Vigna luteola collected from Ethiopia, brownish black in Vigna vexillata, gray in V. caracalla and creamy in Vigna pubescens collected from Saudi Arabia and V. unguiculata (Table 2 and Plate 1 (1a, 2a, 3a, 4a, 5a) and Plate 2 (6a, 7a, 8a, 9a, 10a and 11a). 
 
Surface texture
 
Surface texture of V. pubescens and the cultivated of V. radiata was pubescent, while in the V. radiata wild scabrous, glabrous in V. trilobata and V. caracalla, V. unguiculata and V. luteola (Table 2, Plate 1 (1a, 2a, 3a, 4a, 5a) and Plate 2 (6a, 7a, 8a, 9a, 10a and 11a).
 
Size
 
Three categories of legume size are recognized the smallest are those of wild species V. trilobata,: V. radiata and V. luteola; while the legume of V. radiata cultivated, V. pubescens and V. caracalla are of medium size. The remaining species have long legume (Table 2, Plate 1 (1a, 2a, 3a, 4a, 5a) and Plate 2 (6a, 7a, 8a, 9a, 10a and 11a). 
 
Number of seeds per legume
 
The number of seed per legume was recognized into two categories: the first with less than 10 seeds (V. radiate, V. trilobata, V. caracalla, V. pubescens and V. luteola) and the second with more than 10 seeds (Table 2).
 
Number of locules per legume
 
The number of locules per legume 6-8 in V. radiata and V. trilobata, 9-11, 9-12, 9-13 in V. luteola, V. radiata were collected from Egypt and V. caracalla respectively, while 8-10, 8-11 in V. pubescens, 22-28, 9-15, 16-20 in V. unguiculata respectively (Table 2).
 
Orientation
 
Two types of pod orientation were observed as erect and pendent. In V. radiata, collected from Egypt, and in V. unguiculata and V. caracalla the legume is pendent; while the legume is erect in V. radiata, V. trilobata, V. vexillata and Vluteola.
 
Seed set percentage
 
Seed set percentage was reported as two categories: the first less than 75% in wild V. pubescens while the second more than 75% in the remainder of the studied species (Table 2).
 
Stomata on the surface of mature legume
 
Most of the studied taxa possess barely sunken elliptical stomata with raised stomatal rims and long narrow aperture; the stomatal leveling ranged between superficial, at a level and semi-depressed. It is superficial only in V. vexillata, semidepresed in V. radiata V. trilobata and V. caracalla and at the level in the remainder of the studied taxa.
 
The shape
 
Shape of stomata is broadly to narrowly elliptical in V. vexillata and V. caracalla (Table 3; Plate 1, (4b and 5b); while elliptical shaped in the remainder of the studied taxa (Table 3; Plate 1 (1b, 2b, 3b) and Plate 2 (6b, 7b, 8b, 9b, 10b and 11b).
 
 
The size
 
There is an inter and intra-specific variation in the stomatal size. Stomata ranged in size  from  10-33 × 4-20 µm (length × width). However, the largest stomata are those of V. vexillata 30-33 µm long, and 17-20 µm wide. The smallest are those of V. luteola 10-14 µm long, 5-7 µm wide (Table 3).
 
The rim
 
Rims are usually broad and raised in all taxa. Most of the studied taxa possess stomata with one stomatal rim and peristomatal rim.
 
Cuticular and wax ornamentation
 
The reticulate cuticular ornamentation was recorded for V. luteola, section Vigna and V. unguiculata, V. pubescence collected from Saudia Arabia section Catiang while it is rugose in V. pubescens collected from Ethiopia. It is favularite in V. radiata and V. trilobata section Ceratotropis, while rugose in V. caracalla section Segmidotrops and ruminate in V. vexillata section Plectrotrops (Plate 1  (1b, 2b, 3b, 4b, 5b) and Plate 2 (6b, 7b, 8b, 9b, 10b and 11b; and Table 3).   
 
Hilum
 
Position
 
The hilum in the examined seeds was located in central position as shown in V. radiata, V. trilobata and V. caracalla Plates 3 (Figures 1, 2, 3, 4 and 5); whereas, the hilum was subcentral in the remainder of the studied taxa (Table 4; Figures 6, 7, 8, 9, 10 and 11 (Plates 4)). The hilum shape was oblong in V. radiata, elliptical in V. vexillata, V. caracalla and V. luteola, ovate in the other taxa; and the level is sunken in V. caracalla while raised in the remaining taxa.
 
The size
 
The size of hilum varied in all studied species and their accessions, where the size ranged from 2.0-3.0 mm ×1.0-1.4 mm to 0.7-1.0 × 0.5-0.7 mm (Length × Width). The longest size (2.0-3.0 mm) was found in V. unguiculata from Egypt (No. 8), (Figure 8), followed by 1.8 -2.0 mm in V. unguiculata from Ethiopia (No. 10), (Figure 10); the lowest one was  found  in  V. trilobata,  0.7 - 1.0 mm long, 0.5 - 0.7 mm wide (Figure 3).
 
The data in Table 4 show that there is a high difference in the size of the hilum between the taxa of the same species of the studied genus as in V. unguiculata (2.0-3.0 mm length, 1.6-1.7 mm length and 1.8-2 mm length), V. radiata (1.4-1.6 mm length and 1.2-1.4 mm length) and V. pubescens (1.1-1.3 mm length and 1.3-1.5 mm length); V. luteola (1.3-1.5 mm length) V. vexillata (1.2-1.6 mm length), and V. caracalla (1.4-1.6 mm length).
 
 
 
 
The rim aril and colours
 
The term aril is used for a fleshy to hard structure that develops from the funiculus or ovule after fertilization and invests part or all of a seed. Rim aril was absent in V. radiate and V. trilobata; whereas, it is well developed with superficial expansion and a cleft margin in V. vexillata and V. caracalla. However, in the remainder of the taxa, the aril is well developed with superficial expansion without cleft margin.  Diverse rim colours were observed varying from black, brown to yellowish brown (Figures 1a, 2a, 3a, 4a, 5a, 6a, 7a, 8a, 9a, 10a and 11a and Table 4).
 
Ornamentation   
 
The examinations of hilum surface by S.E.M showed different types of ornamentations were present in cultivated V. radiate. Among the ornamentation, there was rugose structure type, wax flakes depositions; and heavy globular waxy depositions of various size were also clearly seen. The hilum in wild V. radiata and V. trilobata was reticulate with some waxy flakes. In V. vexillata it is rugulose striate, while in V. caracalla, the cells of hilum are disposed in a uniserate compact filament and are fusiformed with the rim. However, in V. pubescens the hilum is reticulate colliculate. In V. unguiculata there is regulate pattern, and it contains a furrow-like array of longitudinal cells with light waxy deposition. The hilum of V. luteola is reticulate foveat (Figures 1b, 2b, 3b, 4b, 5b, 6b, 7b, 8b, 9b, 10b and 11b; Table 4).


 DISCUSSION

In this study, our observations of the macro and micromorphological features revealed the presence of considerable morphological variations between the different subgenus species. In subgenus Ceratotropis, the pods are pendent, non-shattered and brown in colour in cultivated V. radiate. Whereas, it is erect, shattered and black in colour in wild V. radiata and V. trilobata at maturity.  In   subgenu    Plectrotropis,  the  pod  is erect, shattered and brownish black in colour. In subgenus Sigmoidotropis, the pod is pendent and non-shattered; and grey in colour. In the subgenus Vigna, the pod is pendent and shattered or non-shattered/erect and shattered. In this respect these variations were previously observed by many authors such as Garba and pasqet (1998), Sangwan and Lodhi (1998), Peksen and Peksen (2013), and Popoola et al. (2015, 2017).  Many accessions within V. unguiculata show long and relatively large seeds with seed coat colour or patterns similar to the cultivated cowpea; although, some of their character's pod structure, pod position on the raceme and pod shattering are characteristics of the wild species. Also, diversity of V. vexillata is also well represented on the variability of the surface pubescence (that is, plant almost glabrescent, densely pubescent to bristly) as reported by Padulosi and Ng (1993).
 
The number of seeds per pod of the cultivated V. unguiculata (7-12 seeds) was relatively small compared with the cultivated (No. 8), (20-25 seeds) followed by the  wild  form  of  this  species  (15-18 seeds) from subgenus Vigna; and the other studied species which had a range of 6-10 seeds. However, the seeds per pod of the two accessions of V. radiata and trilobata (5-6 seeds) from subgenus Ceratotropis were relatively small compared with the other studied species of Vigna.  A similar variation in the pod size for different taxa of Vigna was reported by many investigators, and it could be attributed to the geographical distribution of these taxa (Barrett, 1990; Hymowitz, 1990; Pasquet, 1998; Pasquet and Vanderborght, 2000; Fery, 2002; Bisht et al., 2005). The usage of the scanning electron microscope (SEM) studies in examination of legume surface of 7 species of Vigna (11 accessions) revealed the importance of this technique as a taxonomic tool. The results in this investigation showed fairly heterogeneous stomatal surface patterns of the legume in the different species of Vigna; and also this data offer significant information to be used in classification similar to that studied by El-Hadidy (2004) for the genus Lotus, family Fabaceae.
 
The seed of V. radiata and V. trilobata section Ceratotropis  has  no  rim-aril,  while   the   other  species  studied have a superficial expansion of the rim aril. In comparison, V. vexillata section Plectrotropis and V. caracalla section Sigmidotropis have a superficial expansion with cleft margin, which is comparable to the dry tongue aril illustrated by Gunn (1981) for some Papilionoideae   taxa.   These   observations  are  also  in agreement with the studies of Fabiana et al. (2013) and Khedia et al. (2017), on some species of Vigna.
 
The examination of hilum ornamentation of some species, that were studied here for the first time by  SEM, have demonstrated the existence of diversity in various taxa; and revealed that the mature seed surface pattern is not identical; rather it showed the presence of a remarkable difference between the two collected accessions of each species such as in wild and cultivated forms collected from different countries. In contrast, the spermoderms of both collected taxa of V. pubescens are reticulate colliculate, regulate in V. vexillata and V. unguiculata, compact in V. caracalla, rugae in Vradiata wild while reticulate in V. radiata cultivated, and reticulate foveate in V. luteola; as also was illustrated by Kumar and Rangaswamy (1984), Fabiana et al. (2013) and Khedia et al. (2017).


 CONCLUSION

The macro and micro-morphological characteristics of legume and hilum were reported. This includes legume features such as shape, colour, size, surface texture, number of locules, number of legumes per peduncle, seed set percentage, and orientation. Moreover, by using scanning electron microscopy, additional detailed information was obtained such as stomatal shape, size, type, cuticular and wax ornamentation of the mature pod surface. In addition, hilum shape, size, position, and ornamentation were elucidated by using scanning electron microscopy. The results provide clear and important attributes of these species for the definition and identification of the taxa collected from different countries.


 CONFLICT OF INTERESTS

The authors have not declared any conflict of interests.


 ACKNOWLEDGEMENT

The authors express their gratitude to Prof. Dr. Abbas, A. El-Ghamery, Botany Department, Faculty of Science, Al-Azhar University for revision and valuable discussions in this manuscript.



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