BIOTROPIA The Southeast Asian Journal of Tropical Biology Vol. 31 No. 2, 2024: 157 - 168 DOI: 10.11598/btb.2024.31.2.1872 ISSN: 0215-6334 | e-ISSN: 1907-770X 157 DIVERSITY OF Citrus spp. FROM NORTH PENAJAM PASER REGENCY (IKN), EAST KALIMANTAN, BASED ON MORPHOLOGICAL CHARACTERS Linda Oktavianingsih1*, Noviana Crisdayanti2, Muhammad Fauzi Arif3, And Medi Hendra4 1Laboratory of Animal Anatomy and Microtechnique, Department of Biology, Faculty of Mathematics and Natural Sciences, Mulawarman University, Samarinda 75119, Indonesia. 2Undergraduate Student, Department of Biology, Faculty of Mathematics and Natural Sciences, Mulawarman University, Samarinda 75119, Indonesia. 3Laboratory of Biology, Department of Biology, Faculty of Mathematics and Natural Sciences, Mulawarman University, Samarinda 75119, Indonesia. 4Laboratory of Plant Anatomy and Systematics, Department of Biology, Faculty of Mathematics and Natural Sciences, Mulawarman University, Samarinda 75119, Indonesia ARTICLE HIGLIGHTS • This research is the first study to provide data on the diversity of Citrus spp. from North Penajam Paser Regency (IKN) area, East Kalimantan for further development and breeding program. • This research offers new information on the distribution of Citrus spp. from the North Penajam Paser Regency (IKN) area, East Kalimantan. • This research provides information on the phenetic relationships among Citrus spp. from the North Penajam Paser Regency (IKN) area, East Kalimantan, and the morphological characteristics that play the most significant role in their clustering. ABSTRACT Citrus spp. is a locally grown fruit in Indonesia with high diversity and widespread distribution in the country. However, the diversity and similarities among Citrus spp. from North Penajam Paser Regency, East Kalimantan, have not been previously studied. Morphological characteristics were used in this study to provide data on the diversity and similarities of Citrus spp. from North Penajam Paser. The data provided served as a primary resource for further development and breeding. Additionally, observation, sampling, morphological data characterization, and scoring were carried out before the data was analyzed. This study identified 35 accessions of Citrus spp., including Citrus × limon (L.) Osbeck "Jeruk Lemon", Citrus reticulata Blanco "Jeruk Siam", Citrus × aurantiifolia (Christm.) Swingle "Jeruk Nipis", Citrus × aurantium L. "Jeruk Manis", Citrus maxima (Burm.) Merr. "Jeruk Bali", Citrus × microcarpa Bunge "Jeruk Kalamansi", and Citrus × aurantium L cv. Sweet Orange Group "Jeruk Sunkist", distributed across Penajam, Sepaku, Babulu, and Waru subdistricts. The dendrogram divided the species into two large clusters. Cluster A was based on characteristics, such as tree habitus, cordate leaf apex, entire leaf margin, pink mesocarp color, and the absence of seeds. Meanwhile, Cluster B was characterized by shrub habitus, acute and rounded leaf apex, crenate leaf margin, white to orange mesocarp color, and numerous seeds in the fruit. The diversity indicated the high potential value of Citrus spp. from North Penajam Paser for further development through a plant breeding program. Keywords: Citrus spp, diversity, Kalimantan, morphological characters Article Information Received 14 February 2023 Revised 17 November 2023 Accepted 15 March 2024 *Corresponding author, e-mail: oktavianingsihlinda@yahoo.com, lindaoktavianingsih@fmipa.unmul.ac.id Research Article INTRODUCTION Indonesia is a mega biodiversity country, with an estimated 30,000 to 35,000 plant species dispersed across the landscapes (von Rintelen et al. 2017). The rich diversity is primarily attributed to several factors, including the equatorial proximity resulting in abundant rainfall, geological topographic variations, unique biogeographical conditions, diverse ecological conditions, and various climatic influences (Lohman et al. 2011). The distribution of numerous islands also leads to distinctive variations in the characteristics of species (Schouppe et al. 2017; Dart et al. 2012; Herlihy & Eckert 2005). This includes plants on Kalimantan Island, which is the third largest island in the world with enormous biodiversity. https://doi.org/10.11598/btb.2024.31.2.1872 mailto:oktavianingsihlinda@yahoo.com BIOTROPIA Vol. 31 No. 2, 2024 158 Studying plant diversity and classification is critical in facilitating the development of economically valuable plant varieties or cultivars (Swarup et al. 2020). This field of study provides essential genetic information for the resources that form the foundation of plant breeding programs (Shen et al. 2018). Furthermore, examining plant diversity and similarity limits the analysis of morphological characteristics, including organ shape, size, color, and surface (Balduzzi et al. 2017). Morphological characterization entails observing external features to assess and establish relationships among individuals, with each character assigned a value that can serve as a marker for assessing these relationships (Jensen 2009). Plants exhibit preferential traits that have evolved in response to various environmental conditions. Therefore, the study of morphological variation is essential as it shows the relationship with these environmental factors. Plants also possess an adaptive mechanism known as phenotypic plasticity, enhancing fitness and resilience in changing environmental conditions (Chevin et al. 2010; Valladares et al. 2014; Ayala et al. 2020). Citrus  spp. spread across Malay Archipelago, which includes Indonesia (Liu et al. 2012; Langgut 2017; Sofiyanti et al. 2022). These species are a valuable source of germplasm due to remarkable genetic diversity (Luro et al. 2011; Yu et al. 2018; Goh et al. 2022). Citrus spp. are also significant in local communities, serving as a vital source for various essential commodities. These include food, medicinal applications, and cosmetic formulations (Marti et al. 2009; Vijaylakshmi & Radha 2015; Samarina et al. 2021; Galovicová et al. 2022; Sapkota et al. 2022; Musara et al. 2023). Among the manifold applications, the fruits and leaves are frequently studied for potential cosmetics use, aromatic properties, and as indispensable ingredients in culinary and beverage preparations (Palazzolo et al. 2013; Khettal et al. 2016; Dosoky & Setzer 2018; Namani et al. 2018; Taufiq et al. 2019; Raghavan & Gurunathan 2021). Citrus spp. has been considerably cultivated across various countries, with a remarkable presence in South East Asia. For instance, several studies found numerous distinct  species in Indonesia, with many being hybrid products (Penjor et al. 2014; Susandarini et al. 2016; Yulian et al. 2021; Sofiyanti et al. 2022; Hardianto et al. 2023). "Jeruk Mandarin", which belongs to the  Citrus reticulata  Blanco, represents the most prevalent variety documented in Indonesia (Yulianti et al. 2020; Luro et al. 2023). "Jeruk Besar" refers to pomelos and is classified as  Citrus grandis  (L.) Osbeck (Ollitrault et al. 2020; Sofiyanti 2022), while "Jeruk Sitrun" is identified as  Citrus medica (L.) Burm.f. (Benedetto et al. 2021). Budiyati et al. (2016) carried out characterization to determine the diversity of Citrus spp. on Kalimantan Island, specifically in East Kalimantan, along with studies conducted in several other regions in the archipelago, such as Southeast Maluku, North Maluku, and East Java. Although the study in East Kalimantan was carried out generally in each region and was not specific for North Penajam Paser Regency, considerable variations were observed. North Penajam Paser Regency was designated as the national capital of the Republic of Indonesia (Ibu Kota Nusantara) following Law Number 3 of 2022 concerning the National Capital (UU IKN) and the Regulatory Database (JDIH BPK RI) on 15 February 2022. This caused a massive change in the region, including the reduction and loss of plant germplasms due to the large number of new arrivals and simultaneous development that damaged the environment. The impact of these changes dramatically affected the use of plants. Therefore, this study aimed to provide phenetic diversity and similarity data for Citrus spp. from North Penajam Paser (IKN) for the benefit of local communities. The results are expected to serve as fundamental data for future studies. MATERIALS AND METHODS Observation and Sampling This study was conducted from September 2021 to February 2022 in the North Penajam Paser Regency, East Kalimantan (Fig. 1). Citrus accessions were selected in this area and the observation site was selected based on potential location-based information provided by local communities. Sampling was carried out using the roaming method with a purposive sampling technique (Rugayah 2004). Leaves, flowering parts, fruits, and seeds that were observable were analyzed. All essential data were recorded using a camera to support the characterization. Analysis was carried out at the Laboratory of Plant Anatomy and Systematics, Faculty of Mathematics and Natural Sciences, Mulawarman University, Samarinda, East Kalimantan Province. Diversity of Citrus Spp. from North Penajam Paser Regency (IKN) - Oktavianingsih et al. 159 Figure 1 Location of collection of Citrus spp. accessions in North Penajam Paser Regency Characterization and Scoring Morphological characterization was conducted by analyzing the shape, length, texture, and color of the organs. A total of 50 critical characteristics were selected for characterization and converted into numerical values (Table 1) to assess diversity and similarity. The matrix score used in this analysis was based on Zufahmi and Nurlaila (2018) and IPGRI (1999). Each sample was assigned an accession number for identification purposes, and the identification process was carried out up to the species level. Table 1 Conversion of morphological characters Citrus spp. for scoring No Characters Scoring 1 Trunk ratio/rootstock diameter 0: 0 < 1 cm; 1: 1 = 1 cm; 2: > 1 cm 2 Scion trunk surface 0: Smooth; 1: Grooved 3 Tree shape 0: Ellipsoid; 1: Spheroid; 2: Obloid 4 Branch density 0: Sparse; 1: Medium; 2: Dense 5 Habitus 0: Shrub; 1: Tree 6 Stem color 0: Brown; 2: Green 7 Stem height 0: < 5 cm; 1: ≥ 5 cm 8 Stem shape 0: Perpendicular; 1: Perpendicular cylindrical 9 Stem surface 0: Thorny; 1: Flat 10 Spine length 0: ≤ 5 mm; 1: 6-15 mm; 2: 16-40 mm; 3: > 40 mm 11 Spine shape 0: Curved; 1: Straight 12 Shoot tip color 0: Green; 1: Purple; 2: Black; 3: Orange 13 Shoot tip surface 0: Glabrous; 1: Intermediate; 2: Pubescent 14 Leaf type 0: Sole; 1: Complex 15 Leaf apex 0: Acute; 1: Obtuse; 2: Rounded; 3: Cordate; 4: Sagittate 16 Leaf base 0: Acuminate; 1: Truncate 17 Leaf margin 0: Entire; 1: Crenate; 2: Sinuate 18 Leaf surface 0: Rough; 1: Smooth 19 Leaf shape 0: Ovate; 1: Elliptical 20 Leaf length 0: < 9 cm; 1: ≥ 9 cm 21 Leaf stalk 0: < 1.5 cm; 1: ≥ 1.5 cm BIOTROPIA Vol. 31 No. 2, 2024 160 No Characters Scoring 22 Leaf width 0: < 5 cm; 1: ≥ 5 cm 23 Green Intensity 0: Light; 1: Medium; 2: Dark 24 Leaf lamina 0: Elliptic; 1: Ovate; 2: Obovate; 3: Lanceolate; 4: Orbicular; 5: Obcordate; 6: Oblong 25 Petiole wings 0: Absent; 1: Present 26 Petiole width 0: Narrow: 1: Medium; 2: Broad 27 Petiole shape 0: Obcordate; 1: Obdeltate; 2: Obovate; 3: Linear 28 Fruit shape 0: Spheroid; 1: Ellipsoid; 2: Pyriform 3: Oblique; 4: Obloid; 5: Asymmetric 29 Fruit base shape 0: Necked; 1: Convex; 2: Truncate; 3: Concave; 4: Concave collared; 5: Collared with neck 30 Fruit apex shape 0: Acute; 1: Rounded; 2: Truncate; 3: Mammiform 31 Fruit diameter 0: < 33 cm; 2: ≥ 33 cm 32 Fruit skin (epicarp) 0: Green/yellow; 1: Red; 2: Dark yellow; 3: Orange; 4: Dark yellow; 5: Dark green; 6: Light green 33 Fruit surface texture 0: Rough; 1: Smooth; 2: Papillate; 3: Pitted 4: Bumpy; 5: Grooved 34 Fruit peel thickness 0: Dense; 1: Light 35 Mesocarp color 0: Pink; 1: Yellow; 2: White; 3: Green; 4: Orange; 5: Light orange 36 Fruit taste 0: Sweet; 1: Sour 37 Vesicle length 0: Short; 1: Long 38 Nature of oil glands 0: Very weak; 1: Conspicuous; 2: Strongly conspicuous 39 Crown color 0: White; 1: Yellowish; 2: Violet 40 Stamen length 0: < 1 m; 1: ≥ 1 cm 41 Number of stamens 0: < 4 each petal; 1: 4 each petal; 2: > 4 each petal 42 Amount of pollen 0: Low; 1: Normal; 2: High 43 Flower stalk length 0: < 1 cm; 1: > 1 cm 44 Flower type 0: Hermaphrodite; 1: Male; 2: Female 45 Number of seeds 0: Low; 1: High 46 Seed shape 0: Fusiform; 1: Clavate; 2: Cuneiform; 3: Ovoid; 4: Semi deltoid; 5: Spheroid; 6: Semi-spheroid 47 Seed surface 0: Smooth; 1: Wrinkled; 2: Hairy 48 Seed color 0: White; 1: Beige; 2: Yellowish; 3: Green; 4: Brown 49 Cotyledon color 0: White; 1: Bright yellow; 2: Bright green; 3: White and green; 4: Green; 5: Dark green; 6: Violet 50 Presence/absence of Seeds 0: Absent; 1: Present Source: IPGRI (1999); Zufahmi and Nurlaila (2018). Data Analysis Morphological characters were analyzed qualitatively and quantitatively. The qualitative analysis entailed using descriptive methods, where specimens were directly observed and matched with references (IPGRI 1999; Irsyam & Chikmawati 2015; Zufahmi & Nurlaila 2018), while quantitative analysis was carried out by measuring specific dimensions with a ruler, respectively. The process began with character selection as Operational Taxonomy Units (OTUs), with each chosen character assigned a standardized score (Table 1). All 50 selected characters were used for observation and converted into Diversity of Citrus Spp. from North Penajam Paser Regency (IKN) - Oktavianingsih et al. 161 standardized values. Subsequently, the data were entered into MVSP 3.1 (MultiVariate Statistical Package) software, which was used to generate the dendrogram and similarity index. The dendrogram was constructed using the Unweighted Pair Group with Arithmetic Mean (UPGMA) method with the Euclidean coefficient. RESULTS AND DISCUSSIONS Diversity In this study, Citrus spp. was found in Penajam, Sepaku, Babulu, and Waru at various altitudes, ranging from 1 masl to 40 masl (meters above sea level). There were 35 identified accessions classified into seven species of Citrus, namely Citrus × aurantium L. “Jeruk Manis”, Citrus × aurantiifolia (Christm.) Swingle “Jeruk Nipis”, Citrus × microcarpa Bunge “Jeruk Kalamansi”, Citrus reticulata Blanco “Jeruk Siam”, Citrus maxima (Burm.) Merr. “Jeruk Bali”, Citrus × limon (L.) Osbeck “Jeruk Lemon”, and Citrus × aurantium L. cv. Sweet Orange Group (Table 2). Morphological Characteristics The characterization of 35 Citrus spp. accessions was conducted using 50 morphological characters based on the IPGRI Descriptor for Citrus characterization guidebook (1999), Zulfahmi (2018), and Irsyam and Chikmawati (2015). These characters revealed variations in leaves, fruits, and seeds. However, due to the unavailability of flowers in some accessions, the characterization primarily focused on fruit and leaf characters. This approach was based on a study conducted by Susandarini et al. (2013), which also emphasized the morphological character analysis of leaf and fruit characteristics for the same reason. Leaf (Folium) Morphological variations in leaves included variations in leaf shape and color. Leaf shapes varied from ovate, obovate, lanceolate, and orbicular, to elliptic (Fig. 2). Leaf colors ranged from light green to dark green. Citrus × limon (L.) Osbeck and Citrus × microcarpa Bunge both exhibited ovate leaf shapes. Citrus × aurantiifolia (Christm.) Swingle had elliptic leaves, while Citrus × aurantium L. and Citrus reticulata Blanco featured lanceolate leaves. Citrus maxima (Burm.) Merr. had orbicular leaves and Citrus × aurantium L. cv. Sweet Orange Group displayed obovate leaves. Leaf color ranged from light to medium and dark. Specifically, Citrus × limon (L.) Osbeck leaves were green to dark green with a rough surface, acute leaf apex, and crenate margin. Citrus × aurantiifolia (Christm.) Swingle leaves were medium green with a rough surface, obtuse apex, acuminate base, and crenate margin. Citrus × microcarpa Bunge leaves were dark green with a smooth surface, obtuse apex, truncate base, and entire margin. Citrus × aurantium L. cv. Sweet Orange Group leaves were medium green with a smooth surface, acute apex, truncate base, and crenate margin. Citrus × aurantium leaves exhibited a medium to dark green color with a smooth surface, acute apex, acuminate base, and crenate margin. Citrus reticulata Blanco leaves were medium to dark green, with an acute apex, truncate base, smooth surface, and crenate margin. Lastly, Citrus maxima (Burm.) Merr. leaves were dark green with a cordate apex, truncate base, smooth surface, and entire margin. A distinguishing feature of Citrus maxima (Burm.) Merr. from other Citrus species was the presence of petiole wings. Citrus × limon (L.) Osbeck and Citrus reticulata Blanco lacked wings on the leaf stalks, while some Citrus species had leaf stalks with wings and others did not. Figure 2 depicts the morphological characteristics of Citrus leaves. Budiarto et al. (2017) emphasized the presence of petiole wings as a significant morphological character that distinguished Citrus species. The results showed variations in leaf lamina shapes, which differed from previous studies. Sofiyanti et al. (2022) reported that the leaf lamina shapes of Citrus spp. from Riau varied between elliptic, oblong, obovate, ovate, and oblanceolate shapes. The discrepancies in the identified accessions were primarily due to differences in the study populations. Previous studies also identified a greater number of Citrus spp. compared to this study. Species, such as Citrus × amblycarpa (Hassk) Ochse “Jeruk Saring”, Citrus × aurantium L. “Jeruk Hantu”, Citrus hystrix DC. “Jeruk Purut”, Citrus hystrix DC. “Jeruk Sundai”, Citrus longilimon Tanaka, and Citrus medica L. “Jeruk Pagar”, were not found in this study. This indicated that the variation in Citrus species from Riau was more significant than the species from North Penajam Paser, East Kalimantan. BIOTROPIA Vol. 31 No. 2, 2024 162 Table 2 List of Citrus spp, accession found in North Penajam Paser, East Kalimantan Accession number Species name Local name Sampling location Utility Altitude (masl) A-1 Citrus × limon (L.) Osbeck Jeruk Lemon Waru Foodstuffs, beverages, traditional medical applications 11 A-2 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Waru Foodstuffs and beverages 15 A-3 Citrus × limon (L.) Osbeck Jeruk Lemon Waru Beverages 16 A-4 Citrus × microcarpa Bunge. Jeruk Kalamansi Waru Foodstuffs and beverages 16 A-5 Citrus reticulata Blanco Jeruk Siam Waru Foodstuffs and beverages 19 A-6 Citrus × aurantiifolia (Christm.) Swingle Jeruk Nipis Waru Foodstuffs, seasoning ingredients 14 A-7 Citrus × microcarpa Bunge. Jeruk Kalamansi Waru Foodstuffs, seasoning ingredients 5 A-8 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Babulu Foodstuffs 40 A-9 Citrus reticulata Blanco Jeruk Siam Penajam Foodstuff 9 A-10 Citrus × aurantiifolia (Christm.) Swingle Jeruk Nipis Penajam Seasoning ingredients 1 A-11 Citrus maxima (Burm.) Merr. Jeruk Bali Waru Foodstuffs 12 A-12 Citrus × limon (L.) Osbeck Jeruk Lemon Babulu Beverages 38 A-13 Citrus × aurantium L. Jeruk Manis Waru Foodstuffs and beverages 28 A-14 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Waru Foodstuffs and beverages 28 A-15 Citrus × limon (L.) Osbeck Jeruk Lemon Penajam Beverages 4 A-16 Citrus × aurantium L. Jeruk Manis Penajam Foodstuffs 29 A-17 Citrus × microcarpa Bunge. Jeruk Kalamansi Penajam Seasoning ingredients 32 A-18 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Penajam Foodstuffs 14 A-19 Citrus × aurantiifolia (Christm.) Swingle Jeruk Nipis Penajam Foodstuffs, seasoning ingredients 4 A-20 Citrus × aurantium L. Jeruk Manis Penajam Foodstuffs 8 A-21 Citrus × aurantiifolia (Christm.) Swingle Jeruk Nipis Penajam Foodstuffs, seasoning ingredients 10 A-22 Citrus × aurantium L. Jeruk Manis Sepaku Foodstuffs 11 A-23 Citrus × microcarpa Bunge. Jeruk Kalamansi Sepaku Foodstuffs, seasoning ingredients 16 A-24 Citrus × limon (L.) Osbeck Jeruk Lemon Sepaku Beverages, traditional medical applications 15 A-25 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Sepaku Foodstuffs 6 A-26 Citrus × aurantium L. Jeruk Manis Sepaku Foodstuffs, beverages 24 A-27 Citrus maxima (Burm.) Merr. Jeruk Bali Sepaku Foodstuffs 12 A-28 Citrus × aurantiifolia (Christm.) Swingle Jeruk Nipis Sepaku Foodstuffs, seasoning ingredients 3 A-29 Citrus × limon (L.) Osbeck. Jeruk Lemon Babulu Foodstuffs, beverages 7 Diversity of Citrus Spp. from North Penajam Paser Regency (IKN) - Oktavianingsih et al. 163 Accession number Species name Local name Sampling location Utility Altitude (masl) A-30 Citrus × aurantium L. Jeruk Manis Babulu Foodstuffs 8 A-31 Citrus × microcarpa Bunge Jeruk Kalamansi Babulu Foodstuffs, seasoning ingredients 13 A-32 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Babulu Foodstuffs 13 A-33 Citrus × aurantium L. cv. Sweet Orange Group Jeruk Sunkist Babulu Foodstuffs 32 A-34 Citrus maxima (Burm.) Merr. Jeruk Bali Waru Foodstuffs 13 A-35 Citrus × aurantiifolia (Christm.) Swingle Jeruk Nipis Waru Foodstuffs, seasoning ingredients 14 According to Yulianti et al. (2020), there were 255 Citrus varieties, consisting of 43% mandarin- type, 4% tangerine-type, 0.7% sweet orange- type, 14.3% sour orange-type, 15% pummelo- type, and 23% rootstock, lime, and lemon. The large number of developed varieties makes the classification and identification process relatively complex. Furthermore, Citrus spp. can easily generate new hybrids through intraspecies or interspecies hybridization (Sofiyanti 2022). The species propagate via apomixis, allowing distribution over a wide range (Wang et al. 2017). Apomixis produces seeds that contain embryos from the somatic part of the nuclear cell (Kepiro & Roose 2010). A single seed is capable of developing from 2 to 30 embryos (Koltunow 1993; Wang et al. 2017). This ability is known as polyembryony, and breeders must be particularly aware due to the beneficial and detrimental effects (Wang et al. 2017). Fruit (Fructus) This study identified limited variations in fruit shape among Citrus spp. from North Penajam Paser. The observed fruit shapes ranged from spheroid to obloid, with the majority being obloid. Citrus × limon (L.) Osbeck exhibited a spheroidal shape with a convex base, acute apex, a diameter below 33 cm, a rough surface, and a dark green color. Citrus × aurantiifolia (Christm.) Swingle had an ellipsoid shape with a convex base, rounded apex, a diameter below 33 cm, a smooth surface, and a dark green color. Citrus × microcarpa Bunge was characterized by an obloid shape with a convex base, rounded apex, a diameter below 33 cm, a green or yellow color, and a smooth surface. Citrus × aurantium L. cv. Sweet Orange Group had an obloid shape with a convex base, rounded apex, a diameter below 33 cm, a dark-green color, and a rough surface. Citrus × aurantium L. featured an obloid shape with a convex base, rounded apex, a diameter below 33 cm, a dark-green color, and a rough surface. Citrus reticulata Blanco bore fruits with an obloid shape, truncated base, rounded apex, a diameter below 33 cm, a green or yellow color, and a smooth surface. Citrus maxima (Burm.) Merr. fruits were obloid in shape, had a concave base, rounded apex, a diameter above 33 cm, a green or yellow color, and a rough surface. This study showed that Citrus spp. of the same species generally exhibited consistent fruit shapes. This resullt contradicted the study conducted by Dorji and Yapwattanaphun (2011), which observed variations in fruit characteristics, even among the same species of Citrus. For instance, variations were found in fruit shape and skin surface texture for Citrus reticulata Blanco "Mandarin" across different sampling locations. Similar results were also reported in a study on Citrus maxima (Burm.) Merr. “Jeruk Bali” conducted by Susandarini et al. (2013), which successfully observed variations in fruit characteristics, including fruit shape, apex, base, peel texture, the presence of oil glands on the peel, mesocarp color, number and arrangement of fruit segments, fruit flesh color, texture, as well as the number of seeds. However, Citrus maxima (Burm.) Merr. exhibited different mesocarp colorations, including white and pink (Fig. 3). BIOTROPIA Vol. 31 No. 2, 2024 164 Figure 2 Leaf shapes Notes: Without wing petioles, i.e., Citrus × limon (L.) Osbeck “Jeruk Lemon” (a) A-1; (b) A-3; (c) A-15; (d) A-24; (e) A-29 and Citrus reticulata Blanco “Jeruk Siam” (f ) A-5.With wing petioles, i.e., Citrus maxima (Burm.) Merr. “Jeruk Bali” (g) A-34. Bar: 1 cm. Figure 3 Variations in mesocarp color on the pomelo plant Citrus maxima (Burm.) Merr Notes: (a) A-27 with white color; and (b) A-34 with pink color. Bar: 1 cm. Seed (Semen) Variations in the shape of Citrus seeds include ovoid, semi-deltoid, club-shaped, semi-spheroid, spheroid, and irregular forms (Fig. 4). The seed shape does not necessarily correlate with the species of Citrus. However, cuneiform seeds are typically associated with the pomelo, Citrus maxima (Burm.) Merr (Accession 11 and Accession 27). The semi- spheroid seed shape is commonly found in the Siamese orange, Citrus reticulata Blanco "Jeruk Siam" (Accession 5). Irregular seed shapes are observed in Citrus × limon (L.) Osbeck (Accession 15 and Accession 24), and Citrus × microcarpa Bunge (Accession 23). Cluster Analysis Characterization of 35 Citrus spp. accessions using 50 morphological characters produced a dendrogram divided into two main clusters (Fig. 5). Cluster A contained three accessions, while Cluster B contains 32 accessions. The clustering was determined by the similarity of the characters, and the level of similarity determined the relationship of each accession. The similarity value increased with the number of identical characters. Cluster A had identical characteristics in tree habitus, cordate leaf apex, entire leaf margin, pink mesocarp color, and seed absence. Cluster B had similar characteristics in shrub habitus, acute and rounded leaf apex, crenate leaf margin, white to orange mesocarp color, and numerous seeds in the fruit. Mesocarp The results showed variations in mesocarp color (Fig. 3), including pink, yellow, white, green, orange, and light orange. Citrus × limon (L.) Osbeck had a white mesocarp with a sour taste, prominent visible oil glands, and was seedless. Citrus × aurantiifolia (Christm.) Swingle exhibited a white mesocarp with a sour taste, prominent oil glands, and numerous seeds. Citrus × microcarpa Bunge exhibited a yellow mesocarp with a sour taste, faint oil glands, and was seedless. Citrus × aurantium L cv. Sweet Orange Group featured mesocarp in shades ranging from light orange to orange, a sweet taste, faint oil glands, and few seeds. Citrus × aurantium L. had orange mesocarp, a sweet taste, prominent oil glands, and few seeds. Citrus reticulata Blanco showed mesocarp in shades of yellow to orange, a sweet taste, faint oil glands, and numerous seeds. Citrus maxima (Burm.) Merr. had pink mesocarp with a sweet taste, faint oil glands, and was seedless. The mesocarp is located between the epicarp and the endocarp (Sadka et al. 2019), constituting the pericarp. The outer part of the mesocarp may contain oil vesicles and carotenoid dyes (Szczykutowicz et al. 2020), and the concentration of these components may vary depending on Citrus species, environmental conditions, and other factors. Certain species showed high tolerance to specific environmental conditions. Furthermore, the interior of the mesocarp consists of spongy, white parenchyma tissue (Mabberley 2004). Diversity of Citrus Spp. from North Penajam Paser Regency (IKN) - Oktavianingsih et al. 165 UPGMA Euclidean A-22 A-14 A-32 A-33 A-2 A-25 A-8 A-18 A-30 A-16 A-20 A-24 A-28 A-23 A-4 A-13 A-21 A-6 A-35 A-10 A-19 A-17 A-31 A-26 A-5 A-9 A-7 A-29 A-15 A-12 A-1 A-3 A-34 A-11 A-27 12 10 8 6 4 2 0 B2 B2.1 B2.2 B2.3 B1 B1.1 B1.2 A1 A2 Figure 4 The irregular shape of Citrus seeds Notes: (a) A-15; (b) A-23; (c) A-24; (d) A-5; (e) A-6. Bar: 5 cm. Each subcluster exhibited distinguishable characteristics that can be readily identified. Subcluster A was distinguished primarily by mesocarp color, and in this subcluster, there were variations in mesocarp color among the accessions. Specifically, subcluster A consisted of Citrus maxima (Burm.) Merr with varying mesocarp colors. Accession numbers A-11 and A-34 had pink mesocarp, while accession number A-27 had white mesocarp. Subcluster B comprised 32 accessions and was subdivided into two major groups based on several fruit-related characteristics, including fruit shape, fruit base shape, fruit apex shape, the nature of the fruit oil glands, and mesocarp and skin color. Subcluster B1 included Citrus × limon (L.) Osbeck which was further divided into two groups based on distinctive characteristics. These included the leaf apex color and the presence of seeds in the fruit. Subcluster B2 comprised Citrus × microcarpa Bunge, Citrus × aurantiifolia (Christm.) Swingle, Citrus reticulata Blanco, Citrus × aurantium L. “Jeruk Manis”, and Citrus × aurantium L cv. Sweet Figure 5 Dendrogram and clustering result of 35 Citrus spp. accessions from North Penajam Paser, East Kalimantan. Orange Group. The grouping in this cluster was primarily based on fruit shape, the nature of the oil glands, fruit apex shape, skin color, and mesocarp color. Generally, Subcluster B2 exhibited an obloid fruit shape, conspicuous oil glands, rounded fruit apex, and dense to thick mesocarp with varying colors. PCA (Principal Component Analysis) was performed to support the clustering result (Fig. 6). This analysis showed the pattern of grouping accessions and determined the influential characters in the clustering result. The dendrogram results showed that the key characters for grouping Citrus spp. accessions are related to seed characteristics. These characteristics confined the presence or absence of seeds, seed shape, seed surface characteristics, and the number and texture of the seed surface. In addition, the position of each accession on PCA axis was consistent with the groupings observed in the dendrogram. BIOTROPIA Vol. 31 No. 2, 2024 166 Figure 6 Principal Component Analysis of 50 characters and 35 Citrus spp. accessions from North Penajam Paser, East Kalimantan CONCLUSION Thirty five (35) accessions of Citrus spp. from North Penajam Paser Regency were successfully identified consisting of Citrus × limon (L.) Osbeck “Jeruk Lemon”, Citrus reticulata Blanco "Jeruk Siam", Citrus × aurantiifolia (Christm.) Swingle “Jeruk Nipis”, Citrus × aurantium L. “Jeruk Manis”, Citrus maxima (Burm.) Merr “Jeruk Bali”, Citrus × microcarpa Bunge "Jeruk Kalamansi", and Citrus × aurantium L cv. Sweet Orange Group “Jeruk Sunkist”. Citrus spp. found in Penajam, Sepaku, Babulu, and Waru at varying altitudes of 1-40 masl. The dendrogram generated from 50 characters in this study was divided into two large clusters. Cluster A was grouped based on the tree habitus, cordate leaf apex, entire leaf margin, pink mesocarp color, and seed absence. 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