From Mountains to Molecules: Decoding the Hidden Diversity of Andrena Bees in the Himalayan Wilderness

From Mountains to Molecules: Decoding the Hidden Diversity of Andrena Bees in the Himalayan Wilderness | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article From Mountains to Molecules: Decoding the Hidden Diversity of Andrena Bees in the Himalayan Wilderness Tamjeeda Nisar, Sajad Ahmad Ganie, Manzoor Ahmad Paray, Shakeel Ahmad Mir, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7694933/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Four species of the genus Andrena : Andrena (Euandrena) bicolor , Andrena (Melandrena) cineraria , Andrena (Plastandrena) pilipes , and Andrena (Zonandrena) flavipes were reported for the first time from Jammu & Kashmir, India. Specimens were collected from Srinagar, Pulwama, and Budgam districts and analyzed using 23 morphological characters. In addition, 658 bp COI DNA barcodes were generated and deposited in GenBank, providing reliable molecular markers for species identification. Integration of morphological and molecular datasets revealed distinct diagnostic traits and intraspecific variations, indicating evolutionary divergence likely shaped by geographic isolation and ecological pressures in the Himalayan region. This study emphasizes the role of integrative taxonomy in accurate species delimitation and contributes to the understanding of pollinator diversity and conservation in the fragile Himalayan ecosystem. Entomology Andrena morpho-molecular characterization DNA barcoding pollinator diversity Himalayan ecosystem Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 INTRODUCTION Bees have fascinated humans for centuries, not only for their complex social organization but also for their vital role as pollinators. Unlike solitary insects that complete their life cycle independently, social bees live in colonies with distinct castes that cooperate for colony survival (Michener, 2007 ). Globally, bees are considered the most important pollinators, visiting the majority of flowering plants. It is estimated that nearly 75% of crops of global significance and about 90% of fruits, vegetables, and horticultural crops depend on insect pollination, with bees being the primary contributors (Klein et al., 2007; Ollerton, 2021 ). While honey bees ( Apis spp.) are widely recognized for their pollination efficiency due to traits such as floral constancy, colony strength, and manageability (Delaplane & Mayer, 2000 ; Abrol, 2012 ), non- Apis bees play an equally crucial role. These solitary and semi-social bees often outperform honey bees in specific conditions because of unique behaviors like buzz pollination, longer proboscis, faster foraging, and effective tripping mechanisms (Willmer et al., 2017; Rader et al., 2020 ). Their role has become more critical in recent decades with the decline of honey bee populations caused by habitat loss, monoculture farming, pesticide use, and emerging diseases (Goulson et al., 2015 ).Among solitary bees, the genus Andrena , commonly referred to as mining bees, is one of the largest bee genera, with more than 1,500 described species globally (Michener, 2007 ; Ascher & Pickering, 2023 ). They are especially important in temperate regions, where they serve as primary pollinators of both wild flora and cultivated crops (Wood et al., 2020 ). Recent taxonomic revisions have increased the number of Andrena species reported from India to 36, reflecting the country’s underestimated diversity (Wood & Gupta, 2023 ). These bees, typically ground-nesting, display a wide range of body sizes and colorations, including metallic and reddish forms, and are adapted to diverse ecological niches. Accurate identification of Andrena species remains challenging due to morphological similarities across taxa. To address this, molecular tools such as DNA barcoding have emerged as reliable methods (Bogusch et al., 2020 ). The mitochondrial cytochrome oxidase I (COI) gene has been widely adopted for species-level identification and phylogenetic analysis in bees (Hebert et al., 2003 ; Kek et al., 2017 ). DNA barcoding has proven particularly valuable for resolving cryptic taxa and validating species records (Moroń et al., 2019 ; Chen et al., 2022 ). In India, pioneering studies have used COI barcoding to document species like Andrena agilissima , thereby enhancing regional biodiversity databases (Chandra et al., 2017 ). Furthermore, recent methodological advances, including improved primer design and the use of degraded DNA from museum specimens, are expanding the scope of barcoding in bee systematics (Mitchell, 2022 ; Ashfaq et al., 2023).Thus, combining morphological and molecular approaches provides a powerful framework for understanding the diversity, taxonomy, and ecological roles of Andrena bees. This integration is particularly important in regions like India, where the diversity of solitary bees remains underexplored but ecologically significant. MATERIAL AND METHODS Study area: An extensive field survey was undertaken to document the diversity of Andrena species across three representative districts of the Kashmir Valley: Srinagar, Pulwama, and Budgam (Fig. 1). To ensure wide ecological coverage, three sites were strategically selected within each district. In Srinagar, surveys were carried out at Shalimar, Chatterhama, and Dhara; in Budgam, sampling was conducted at Chadoora, Badipora, and Wahabpora; while in Pulwama, the locations included Malangpora, Newa, and Rajpora. Collections were performed during daylight hours, targeting both cultivated fields and surrounding wild flora to maximize the likelihood of capturing the full range of foraging Andrena species. Particular attention was given to periods of peak bloom for major crops and naturally occurring bee forage plants, as these phenological windows represent the most active phases of bee visitation. Standard entomological techniques were employed to collect specimens, ensuring minimal damage for subsequent morphological and molecular identification.All collected material was processed and curated at the Research and Training Centre for Pollinators, Pollinizers, and Pollination Management (RTCPPPM), Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir (SKUAST-K), Shalimar. This facility provided the necessary infrastructure for specimen preservation, identification, and downstream molecular analyses, thereby serving as the focal point for the research program. Collection and Preservation method: Specimens of Andrena species were collected from different host plants such as apple, dandelion, poison hemlock, almond, and Persian speedwell across several districts of Jammu and Kashmir using a sweep net. The collected bees were immediately transferred to airtight glass killing jars charged with ethyl acetate. Ethyl acetate was chosen because it provides rapid euthanasia while maintaining the natural coloration of the insect and preventing hardening of body tissues, which is essential for subsequent taxonomic studies. After euthanasia, the specimens were carefully stretched, pinned, and mounted in insect collection boxes to ensure proper positioning and long-term preservation. To rehydrate dried specimens prior to examination, the pinned bees were placed in an airtight container with a moistened tissue or paper towel for 12–24 hours. This softening process made handling and dissection easier. To prevent fungal or mold growth during rehydration, a few drops of phenol or thymol were introduced into the container, ensuring safe and hygienic preservation of the specimens for detailed morphological and morpho-molecular analyses. Dissection and Mounting For genitalia preparation, relaxed specimens were first placed in a Petri dish containing either distilled water or ethanol to soften the body tissues. Under a stereo microscope, the abdomen was gently held with fine forceps, and the terminal abdominal segments (typically segments VII–VIII) were carefully detached by pulling backward with the aid of a minuten pin or micro-scissors.The excised abdomen was then immersed in a 10% potassium hydroxide (KOH) solution and heated at 60–70°C for 15–20 minutes. This maceration step dissolved soft tissues and cleared the muscles, thereby facilitating clear visualization of the genital structures. Following digestion, the material was rinsed several times with distilled water to remove residual KOH and to prevent further tissue degradation.The cleaned genitalia were mounted on a glass microscope slide using either glycerin (for temporary preparations) or Canada balsam (for permanent preparations). A cover slip was gently placed on top, ensuring the specimen was properly flattened without distortion, and the edges were sealed with clear nail polish or a synthetic mounting medium to avoid desiccation and fungal growth.Subsequently, morphological examinations were performed with a stereo zoom microscope (OLYMPUS SZX16). Standard morphometric terminology was employed, and the following parameters were recorded: body length; head length and width; thorax length and width; abdomen length and width; clypeus length and width; upper and lower interocular distances (UICD and LICD); maximum diameter of the antennal socket; forewing length and width; hind wing length and width; facial fovea (FOV) length and width; and lengths of individual antennal segments (AS). Ratios such as head length-to-width, FOV length-to-width, antennal segment 3-to-segment 1 (AS3/AS1), and UICD-to-LICD were also calculated. These measurements formed the basis for species-level identification and comparative taxonomic analyses. Authentication of Andrena spp. through DNA Barcoding Collected specimens were preserved either in a dry state or in 100% ethanol to ensure adequate preservation of DNA for molecular studies. Genomic DNA was extracted from the legs and thorax of adult bees using the DNeasy Blood & Tissue Kit (Qiagen, Cat. No. 69504), following the manufacturer’s protocol. The quality of the extracted DNA was assessed on agarose gel electrophoresis, with ethidium bromide used for staining, and electrophoresis carried out for one hour. DNA concentration and purity were subsequently determined using a Nanodrop spectrophotometer.Species-level identification was performed using the mitochondrial cytochrome oxidase subunit 1 (COI) gene marker. PCR amplification of the COI region was carried out using species-specific primers: forward primer (5′ ATTCAACCAATCATAAAGATATTGG 3′) and reverse primer (5′ TAAACTTCTGGATGCTCCAAAAAATCA 3′) as described by Gurpreet et al. (2016). Each PCR reaction was performed in a total volume of 10 µl, consisting of 1 µl DNA template, 1.5 µl forward primer, 1.5 µl reverse primer, 1.3 µl nuclease-free water, 0.6 µl dNTP mix, 0.1 µl Taq polymerase, 2 µl MgCl₂, and 2 µl Taq buffer.(Fig. 2) The amplification protocol included an initial denaturation at 97°C for 5 minutes, followed by 40 cycles of denaturation at 94°C for 1.5 minutes, annealing at 52°C for 1 minute, and extension at 72°C for 1 minute, with a final extension step at 72°C.Amplified PCR products were sequenced bidirectionally using both forward and reverse primers on an ABI 3730XL DNA Analyzer (Applied Biosystems, Gene Technology). The resulting chromatograms were assembled and edited using Geneious v11.0.3, while sequence alignment and analysis were carried out in BioEdit. All sequences generated in this study were submitted to GenBank (Table 1). Remaining DNA samples were archived at − 30°C in the Molecular Laboratory of Biotechnology, SKUAST-K, Srinagar, and are available for future research upon request. Table 1 Sequenced Taxa and GeneBank Accession Numbers: S.NO. Species Sex GeneBank Accession No. 1. Andrena cineraria Male OP646814 2. Andrena flavipes Female OP672272 3. Andrena pilipes Female OP646792 4. Andrena bicolor Female OP673542 Floral Resources: Information on the floral resources utilized by adult Andrenid bees was systematically recorded from the sampling locations across three districts of the Kashmir Valley, namely Srinagar, Pulwama, and Budgam, during the course of the study. At each site, observations were made to identify the plant species that served as foraging sources for the bees. These floral resources were then classified into two categories: family-wise and crop-wise. The taxonomic identification of plant families and species was carried out with the assistance of a professional taxonomist, while additional verification and cross-checking were done using reliable online botanical databases and literature. This approach not only ensured accurate documentation of the bee plant associations but also provided insights into the diversity of floral resources available to Andrena species across agricultural as well as wild landscapes of the valley. RESULTS Upon morphological examination of these species, these were identified as: Andrena cineraria, Andrena flavipes, Andrena pilipes and Andrena bicolor . This identification was authenticated through molecular means also. Andrena ( EuAndrena) bicolor Andrena picicornis –Kirby-1802 Andrena pilosula -Kirby -1802 Andrena gwynana –Kirby -1802 Andrena proxima –Smith -1847 Andrena aestiva –Smith -1849 Andrena consimilis -Smith -1849 Andrena laeviuscula – Schenck-1853 Material examined: 2♀, J&K, India: 2♀, Shalimar Srinagar, 34.148 0 N 74.883 0 E, 1588m, 19.IV.2021, Coll: Tamjeeda, Dandelion, Poison hemlock, Creeping thistle, Apple , Pear. Distribution: India, Scotland, Ireland, Western Palaeartic from Fennoscandia South to Iberia and Corsica, East of Siberia, North Africa, Israel and Iran. Diagnostic Characters Female ( ♀) Also called as Gwynne’s mining bee. Females appear larger as compared to males. Head is black coloured with white –black hair scattered all over the head. Thorax is usually black with golden white coloured hair scattered all over the thorax. Females with sparse pile of brownish hairs on the tergite that form weaker bands. Hind legs have dull orange colour.Mesosoma with orange yellow hairs, Trochanter of hind leg floculus brownish black, tibial scopa orange yellow, Metasomal disc with sparse punctuation. Measurements (Mean±SE in mm): Total body length: 15.16±1.29, Head length: 2.78±0.23, Head width: 3.49±0.29, Head: L/w: 0.79±0.06, Thorax length: 3.97±0.34, Thorax width: 3.23±0.28, Fore wing length: 7.86±7.87, Fore wing width: 2.76±0.23, Hind wing length: 7.28±0.61, Hind wing width: 1.90±0.17, Abdomen length: 5.62±0.48, Abdomen width: 3.88±0.32, Clypeus length: 1.34±0.11, Clypeus width: 2.29±0.19, UICD: 2.42±0.20, LICD: 2.41±0.20, UICD: LICD: 1.00±0.08, Scape length: 1.14±0.09, Pedicle length: 0.32±0.02, Flagellum length: 5.1±0.43, Antennal socket maximum diameter: 0.33±0.02, AS 3 : AS 1: 4.47±0.38, FOV length: 1.08±0.09, FOV width: 0.58±0.05, FOV: L/W: 1.86±0.15 . Andrena ( MelAndrena) cineraria Andrena cineraria ssp. Danuvia - E. Stockhert -1950 Andrena danuvia- E. Stoeckhert- 1950 Material examined: 4♂, J&K, India: 2♂: Newa Pulwama, 33.929 0 N 74.898 0 E, 1588m, 9.III.2022, Coll: Tamjeeda, Pear, Poison hemlock, Yellow star of Bethlehem, 2♂: Badipora Budgam, 33.917 0 N 74.796 0 E, 1610m, 26.III.2021, Coll: Tamjeeda, Dandelion, Almond, Persian speedwell. Distribution: India, Scotland, Ireland, Southern England, South of Iberia, East to Northern China. Diagnostic Characters: Male ( ♂ ): Also called as ashy brown mining bee. Males appear to be smaller than female. They have golden brown hair on the head. Thorax is black in colour with golden hair on the side of thorax. Abdomen black in colour with scattered golden hair all over tergum and sterum. Female have 12 segments in antennae, males have 13 segments in antennae. Genitalia broader than Andrena gravida . Measurements (Mean±SE in mm): Total body length: 11.87±1.00, Head length: 3.40±0.29, Head width: 4.24±0.37, Head: L/w: 0.80±0.06, Thorax length: 3.43±0.29, Thorax width: 2.89±0.24, Fore wing length: 7.74±7.74, Fore wing width: 2..57±0.21, Hind wing length: 2.52±0.21, Hind wing width: 2.52±0.21, Abdomen length: 4.76±0.40, Abdomen width: 2.85±0.24, Clypeus length: 1.55±0.13, Clypeus width: 3.12±0.27, UICD: 2.93±0.24, LICD: 2.97±0.25, UICD: LICD: 0.98 ±1.01, Scape length: 1.04±0.08, Pedicle length: 0.27±0.02, Flagellum length: 4.46±0.37, Antennal socket maximum diameter: 0.45±0.03, AS 3 : AS 1 : 4.28±0.36, FOV length: 1.25±0.10, FOV width: 0.75±0.07, FOV: L/W: 1.66±0.14. Andrena ( PlastAndrena) pilipes Andrena carbonaria - Linneaus- 1767 Material examined: 17♀: J&K, India: 4♀, Dhara, Srinagar, 4.I.2022, 34.169 0 N 74.919 0 E, 1776m, Coll: Tamjeeda, Pear, 5♀, Newa Pulwama, 33.929 0 N 74.898 0 E, 1588m, 9.III.2022, Coll: Tamjeeda, Dandelion, 2♀, Rajpora, Pulwama, 33.822 0 N 74.853 0 E, 1545m, Poison hemlock , Yellow star of Bethlehem, 6♀, Badipora Budgam, 33.917 0 N 74.796 0 E, 1610m, 4.V.2020, Coll: Tamjeeda, Creeping thistle. Distribution: India, Africa, Europe, Turkey and Northern Africa. Diagnostic Characters Female ( ♀) Honey bee sized black coloured Andrena species.Head is black in colour with light black coloured scattered hair, Thorax is black is colour with light golden coloured hair on the side of thorax, Abdomen is black in colour with light black scattered hair on tergum with orange coloured hair on sterum. Hind wing is larger as compared to fore wing. Golden coloured hair scattered over legs. It is a polyletic species. Measurements (Mean±SE in mm): Total body length: 4.73±0.40, Head length: 2.68±0.22, Head width: 3.20±0.28, Head: L/w: 0.83±0.07, Thorax length: 3.46±0.29, Thorax width: 3.23±0.28, Fore wing length: 4.99±4.99, Fore wing width: 1.90±0.17, Hind wing length: 6.56±0.55, Hind wing width: 1.79±0.15, Abdomen length: 3.59±0.30, Abdomen width: 2.81±0.24, Clypeus length: 1.13±0.09, Clypeus width: 1.80±0.15, UICD: 2.06±0.18, LICD: 1.82±0.15, UICD: LICD: 1.13±0.09, Scape length: 3.08±0.26, Pedicle length: 0.22±0.01, Flagellum length: 5.12±0.43, Antennal socket maximum diameter: 0.43±0.03, AS 3 : AS 1 : 1.66±0.14, FOV length: 1.02±0.09, FOV width: 0.86±0.08, FOV: L/W: 1.18±0.10. Andrena ( ZonAndrena) flavipes Andrena fulvicrus- Kirby- 1802 Andrena extricate – Smith -1802 Andrena contigua – Kirby -1802 Andrena flavipes var. cinerascens –Eversmann- 1852 Andrena interrupta - Schenck- 1869 Material examined: 19♀, J&K, India: 5♀, Shalimar Srinagar, 34.148 0 N 74.883 0 E, 1588m, 2.III.2021, Coll: Tamjeeda, 4♀, Chatterhama Srinagar, 34.184 0 N 74.868 0 E, 1592m, 9.III.2021, Coll: Tamjeeda, Apple and Dandelion, 2♀, Newa Pulwama 33.929 0 N 74.898 0 E, 1588m, 8.IV.2022, Coll: Tamjeeda, Pear, 3♀, Malangpora, Pulwama 33.893 0 N 74.982 0 E, 1626m, 8.IV.2022, Coll: Tamjeeda, Almond, 5♀, Chadoora, Budgam, 33.945 0 N 74.796 0 E, 1624m, 9.III.2022, Coll: Tamjeeda, Apple. Distribution: India, Turkey, Southern England and South Coast of Wales. Diagnostic Characters: Female ( ♀) Medium sized Andrena spps. Females have buffish white hair bands that fully occupy the apical depressions of tergite 1-4 with short black hairs between the bands, slight light brownish haired thorax.They have pygidial plate which is pointed apically. Golden coloured hairs on tibia and tarsus of fore and hind leg. Measurements (Mean±SE in mm): Total body length: 3.70±0.31, Head length: 2.16±0.19, Head width: 2.58±0.22, Head: L/W: 0.83±0.07, Thorax length: 2.73±0.23, Thorax width: 2.47±0.21, Fore wing length: 6.95±0.59, Fore wing width: 2.14±0.19, Hind wing length: 6.06±0.51, Hind wing width: 1.66±0.14, Abdomen length: 4.55±0.39, Abdomen width: 2.60±0.22, Clypeus length: 1.02±0.85, Clypeus width: 1.56±0.13, UICD: 1.74±0.14, LICD: 1.65±0.13, UICD: LICD: 1.05±0.08, Scape length: 2.87±0.24, Pedicle length: 0.22±0.01, Flagellum length: 5.43±0.46, Antennal socket maximum diameter: 0.29±0.02, AS 3 : AS 1 : 1.89±0.16, FOV length: 0.95±0.08, FOV width: 0.29±0.02, FOV: L/W: 3.27±0.27 . Molecular Characterization of Andrena spp : For molecular characterization, the mitochondrial Cytochrome Oxidase I (COX1) gene marker was employed. Polymerase Chain Reaction (PCR) was carried out to amplify the target region, and the resulting products were subjected to sequencing. The raw sequences obtained were carefully edited, trimmed, and aligned using BioEdit software to ensure accuracy and remove ambiguities.To study evolutionary relationships among the Andrena species, MEGA10 software was used for constructing phylogenetic trees and performing evolutionary analyses (Fig. 1). Both BioEdit and MEGA programs facilitated the alignment and comparison of the generated sequences with already available reference sequences.The curated nucleotide sequences were then submitted to the NCBI GenBank database , where accession numbers were assigned. These accession numbers now serve as permanent references for the isolated Andrena species. To verify species identity, the acquired sequences were analyzed using the BLASTN tool of NCBI, which compared them against global sequence repositories. This homology search confirmed the taxonomic position of the studied species. Finally, the nucleotide sequences of the COX1 gene for all the characterized Andrena species are presented below, providing a molecular-level reference for future comparative and taxonomic studies. 1.1: Sequencing of the PCR products of the COX-1marker regions of Andrena species: Following are the sequences obtained after amplification of COX-1 region of Andrena species: 1. Andrena bicolor: ( Accession number: OP673542) ATATGAGCAGGTATAATTGGTGCCTCCCTTAGATTCATCATTCGTATAGAACTAAGAAACCCAGGTAGTTGAATTAACAATGATCAATTTATAACTCAATGTTACATCTCATGCTTTTATTATAATTTTTTTCATAGTAATACCATTTATAATTGGAGGATTCGGAAACTGACTCACACCATTAATA CTAGGAGCACCTGATAGCTTTCCCTCGAATAAATAATATAAGATTTTGACTACTACCCCCATCAATTCTAATTATTTTAATAAGAATAGTTTTAAATTCAGGTTCTGGTACAGGATGAACAGTCTATCCCCCCCTTTCATCCTACGCCTTTCACCCATCATCATCAGTAGATTTAACAATTTTTT CTCTTCATATTGCAGGTATTTCGTCAATTATAGGAGCAATTAATTTTATTGTAACAATTTTGAATATAAAAAATATTTCACTAAATTATGATCAAATACCACTATTCCCATGATCAGTATTTATCACTACAAT. 2. Andrena cineraria: ( Accession number: OP646814 ) ATATTTCATCTTCGCTATATGAGCAGGAATAGTCGGTGCATCCCTAAGATTTATCATCCGAATAGAATTAAGAAACCCAGGTATATGAATCA ACAATGATCAATTATATAATTTAATCGTTACCTGGCACGCGTTTATTATAATTTTCTTCATAATAATGCCATTCATAATCGGAGGTTTCG GAAACTGACTCACACCGTTAATAATAGGAGCGCCAAATATGGCGTTTCCTCGTATAAATAATTTGAGATTTTGATCATTGCCCCCTTCAATTCTA ATAATTTTAATAAGAATAATCCTAAATTCGGGGTCTGGTACGGGATGAACAGTTTATCCACCACTGTCATCATACTCATTCCATC CATCATCATCAGTTGATTTAACAATTTTTTCACTTCACATTGCTGGTATTTCATCAATTATAGGGGCGATCAACTTTATTGTTACAATTCTTAATATAAA AAATATTTCATTTAATTATGATCAAATGCCACTGTTTTTTTGGTCTGTCTTGTTAACAGCATTTCTCTTATTAATCTCATTACAAGTTCTAGCTGGAGCAATTACAATATTACTATCAGATCGAAACTT. 3. Andrena flavipes: ( Accession number: OP672272 ) ATCTTCGCCATATGAGCGGGCATAATCGGAACCTCACTAAGATTTATTATCCGAATGGAATTAAGAAATCCAGGAAGCTGAATCAACAATGA TCAAATTTATAATTCAATTGTAACCTCACACGCTTTCATTATAATTTTCTTCATAGTTATACCATTCATAATCGGAGGTTTCGGAAACTGACTCA CACCGTTAATATTAGGAGCGCCCGACATGGCGTTCCCGCGAATAAACAACATAAGGTTTTGATTGTTACCACCCTCGATCTTAAT CATCTTAATAAGAATAGTTCTAAATTCGGGATCGGGAACAGGATGAATTTACCCACCACTATCCTACTCATTCCACCCATCCTCATCAGTAGATCTAACA ATCTTCTCACTTCACATTGCAGGTGTATCATCAATTATAGGTGCAATCAACTTTATCGTAACAATCCTAAATATAAAAAA TATCTCAATAAATTATGATCAACTACCACTATTCCCATGATCAGTATTTATTACCACAATCCTACTACTAATTTCCTTGCCAGTTTTAGCTGGTG CCATCACAATACTCCTATCAGATCGTAATTTAAACTCATCATTCTTCGATCCCATG. 4. Andrena pilipes: (Accession number: OP646792) AAATTTATAATACAATCGTAACCTCTCACGCCTTTATTATAATTTTCTTCATAGTTATGCCATTCATAATCGGAGGTTTCGGAAACTGACTC ACACCATTAATATTAGGAGCGCCCGACATGGCTTTCCCGCGAATAAAATAATATAAGATTTTGGCTACTCCCCCCTTCAATTCTAATTAT TTTAACAAAAATAATATTAAATTCGGGGTCCGGAACAGGTTGAACAATTTACCCTCCATTATCATCATATTCTTATCACCCATCATCCTC AACAGACCTAACAATTTTCTCACTGCACATTGCAGGAATTTCATCAATTATAGGAGCAATCAATTTTATTGTAACAATTCTTAACATAAAAAATA TTTCAATAAATTATGATCAACTACCCCTATTCCCATGATCAGTATTTATTACCACAATTCTTCTATTAATTTCTCTACCAGTTCTAGCTGGAGCTATT. 1. Exploration of Intra and interspecific species variation among collected Andrena species by Phylogenetic Analysis: The Neighbor-Joining (NJ) phylogenetic tree was constructed based on a 658 bp fragment of the COX1 gene obtained from four Andrena species sampled across Jammu and Kashmir. The resulting phylogeny (Fig. 3) demonstrated that the sequences were distinctly separated and grouped accurately with their respective conspecific taxa, indicating clear genetic divergence among the species. To infer the evolutionary relationships, the Maximum Likelihood (ML) method was employed under the General Time Reversible (GTR) model . The initial tree(s) required for the heuristic search were generated by applying the Neighbor-Joining algorithm to a matrix of pairwise genetic distances, which had been estimated using the Maximum Composite Likelihood (MCL) method . The robustness and reliability of the tree topology were assessed through a bootstrap analysis with 1000 replicates . Only those branches supported by more than 50% of bootstrap replicates were retained, while nodes with weaker support were collapsed. This ensured that the final bootstrap consensus tree reliably represented the evolutionary history of the analyzed taxa. Floral resources of Andrena species: Andrena species have been recorded from different floral resources of Kashmir. These floral resources have been categorized both family wise and crop wise as shown in Table 2. Distribution pattern of different species of Andrenid bees on different flora is shown in Table 2. Andrena flavipes was reported from Dandelion and Apple. However, Andrena bicolour was reported from Poison hemlock, Dandelion, Creeping thistle, Apple and Pear. Similarly, Andrena cineraria was found on Almond, Persian speedwell, Poison hemlock, Yellow star of Bethlehem and Pear and Andrena pilipes was reported from Almond, Dandelion, Poison hemlock, Yellow star of Bethlehem, Creeping thistle and Pear. Table 2: Distribution of different species of Andrenid bees on different flora: S.NO Species Crops 1 Andrena pilipes Almond ( Prunus amygdalus ), Dandelion (Taraxacum officinale) , Poison hemlock ( Conium maculatum ), Yellow star of Bethlehem ( Gagea lutea ), Creeping thistle ( Cirsium arvense ) and Pear ( Pyrus ). 2 Andrena bicolour Poison hemlock ( Conium maculatum) , Dandelion ( Taraxacum officinale ) , Creeping thistle (Cirsium arvense) , Apple ( Malus domestica ) and Pear ( Pyrus ). 3 Andrena flavipes Dandelion (Taraxacum officinale ) and Apple ( Malus domestica ). 4 Andrena cineraria Almond ( Prunus amygdalus ), Persian speedwell ( Veronica persica ), Poison hemlock (Conium maculatum) , Yellow star of Bethlehem ( Gagea lutea ) and Pear ( Pyrus ). DISCUSSION The present study provides a comprehensive morphological and molecular characterization of four important Andrena species: Andrena bicolor , A. cineraria , A. pilipes , and A. flavipes from the Kashmir Valley, India. These species represent significant pollinators contributing to both wild flora and cultivated crops in temperate ecosystems. The integration of classical morphological techniques with DNA barcoding has proven critical in accurately identifying species within the genus Andrena , which is notoriously challenging due to morphological similarity and cryptic diversity.Our morphological analysis revealed characteristic diagnostic features that aligned well with existing taxonomic descriptions, such as body size, coloration patterns, antennal segmentation, and genitalia structure. For example, A. bicolor was distinguishable by its larger body size and distinctive orange tibial scopa, while A. cineraria exhibited the typical ashy brown color and specific antennal ratios that differentiated it from congeners (Wood et al., 2020). Importantly, the observed measurements, such as UICD to LICD ratios and flagellum lengths, provide a quantitative basis for comparison with global datasets (Ascher & Pickering, 2023). Molecular identification using mitochondrial COI gene markers further corroborated the morphological diagnoses, overcoming limitations associated with subjective morphological assessment and potential misidentifications. The bidirectional sequencing and GenBank submission of COI sequences represent a valuable addition to the growing DNA barcode reference library for Indian solitary bees, as also highlighted by prior studies (Chandra et al., 2017; Moroń et al., 2019). The successful amplification and sequencing from both fresh and preserved specimens demonstrate the robustness of the molecular protocol employed (Mitchell, 2022; Ashfaq et al., 2023). The documented floral resources revealed a broad spectrum of host plants utilized by Andrena species, including apple ( Malus domestica ), pear ( Pyrus spp. ), poison hemlock ( Conium maculatum ), dandelion ( Taraxacum officinale ), almond ( Prunus dulcis ), and Persian speedwell ( Veronica persica ). These observations reinforce the ecological versatility of Andrena species and their vital role in supporting agricultural productivity and wild plant pollination in Kashmir’s agro ecosystems (Rader et al., 2020; Willmer et al., 2017). This is especially critical given the documented global decline of honey bees (Apis spp.) due to habitat loss, pesticide exposure, and emerging diseases (Goulson et al., 2015). The wide distribution of these Andrena species across Kashmir Valley’s districts further emphasizes their adaptability to diverse environmental conditions, with elevations ranging from 1545 to 1776 meters above sea level. This finding aligns with the global distribution patterns of these species across temperate regions (Michener, 2007; Wood & Gupta, 2023). However, challenges remain in the identification of Andrenid species, particularly when dealing with degraded specimens or closely related cryptic taxa. The success of DNA barcoding in this context highlights the need for further expansion of molecular reference databases, especially for underrepresented regions like India (Hebert et al., 2003; Kek et al., 2017). CONCLUSION In conclusion, the examination of Andrena pilipes , Andrena bicolour , Andrena flavipes and Andrena cineraria has offered significant insights into their diverse morphological adaptations and structural characteristics. Through meticulous description and molecular validation using DNA analysis, distinct features have been identified that enhance the accuracy of species recognition and classification. This comprehensive study not only highlights the ecological and scientific importance of these bees but also enriches our understanding of the evolutionary diversity within the Andrena genus. References Abrol DP (2012) Pollination biology: Biodiversity conservation and agricultural production, vol 100. Springer, pp 76–90. https://doi.org/10.1007/978-94-007-1942-2 Ascher JS, Pickering J (2023) Discover Life bee species guide and world checklist (Hymenoptera: Apoidea: Anthophila). 10.5281/zenodo.10019874 Bogusch P, Blahova E, Horak J (2020) Pollen specialists are more endangered than non-specialised bees even though they collect pollen on flowers of non-endangered plants. Arthropod-Plant Interact 14:759–769. 10.1007/s11829-020-09789-y Ballantyne G, Baldock KCR, Rendell L, Willmer PG (2017) Pollinator importance networks: Are non-bees significant? Ecol Entomol 42(5):1–11. 10.1038/s41598-017-08798 Chandra K, Wood TJ, Patiny S, Dutta A, Laha S, Smith B, Basu (2017) DNA barcoding of Andrena agilissima in India. J Entomol Res 41:123–130. 10.5852/ejt.2024.948.2637 Chen X, Agapow PM, Bininda-Emonds OR, Crandall KA, Gittleman JL, Mace GM, Marshall JC, Purvis A (2022) DNA barcoding reveals hidden diversity of bees in East Asia. Sci Rep 12:54–82 Delaplane KS, Mayer DF (2000) Crop pollination by bees. CABI Publishing Goulson D, Nicholls E, Botías C, Rotheray EL (2015) Bee declines driven by combined stress from parasites, pesticides, and lack of flowers. Science 347:125–595 Hebert PD, Cywinska A, Ball SL, DeWaard JR (2003) Biological identifications through DNA barcodes. Proceedings of the Royal Society B: Biological Sciences , 270:313–321 Kek SP, Bhat S, Paschapur AU, Subbanna ARNS, Stanley J, Gupta J, Mishra KK, Kumar (2017) Application of COI barcoding for bee species identification. Apidologie 48(3):377–387 Klein AM (2007) Importance of pollinators in changing landscapes for world crops. Proceedings of the Royal Society B: Biological Sciences, 274 :303–313 Liu C, Ashfaq M, Yin Y, Zhu Y, Wang Z, Cheng H, Hebert P (2023) Using DNA metabarcoding to assess insect diversity in citrus orchards. Peer J Entomol 11:153–338 Michener CD (2007) The bees of the world. Johns Hopkins University Mitchell A (2022) DNA barcoding with historical specimens: Unlocking museum collections. Mol Ecol Resour 22(7):2565–2578 Moroń D, León-Cortés JL, Bried JT, Roy DB (2019) Effectiveness of DNA barcoding for identifying wild bees. J Insect Conserv 23(5–6):829–840 Ollerton J (2021) Pollinators & pollination: Nature and society. Pelagic Publishing Rader R, Cunningham SA, Howlett BG, Inouye DW (2020) Non-bee insects as pollinators: Global patterns and implications. Proceedings of the National Academy of Sciences , 117: 144–151 Wood TJ, Bogusch P, Bláhová E, Horák J (2020) Diversity and ecology of Andrena bees: A global review. Insect Conserv Divers 13(6):507–523. https://doi.org/10.5852/ejt.2022.843.1947 Wood TJ, Gupta RK (2023) Revision of Indian Andrena bees. Eur J Taxonomy 913:1–58 Additional Declarations The authors declare no competing interests. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7694933","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":519458772,"identity":"d8f07bc3-aae6-411e-8eca-5fb6d17f057f","order_by":0,"name":"Tamjeeda Nisar","email":"data:image/png;base64,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","orcid":"https://orcid.org/0009-0003-0931-0849","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":true,"prefix":"","firstName":"Tamjeeda","middleName":"","lastName":"Nisar","suffix":""},{"id":519458773,"identity":"4f1b9d11-25f5-476f-92a7-1e8f296edecd","order_by":1,"name":"Sajad Ahmad Ganie","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Sajad","middleName":"Ahmad","lastName":"Ganie","suffix":""},{"id":519458774,"identity":"e4894fdf-9887-4028-8064-f6fffcf36199","order_by":2,"name":"Manzoor Ahmad Paray","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Manzoor","middleName":"Ahmad","lastName":"Paray","suffix":""},{"id":519458775,"identity":"6e825f24-73b1-4f26-afc0-d48d77ce9a1d","order_by":3,"name":"Shakeel Ahmad Mir","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Shakeel","middleName":"Ahmad","lastName":"Mir","suffix":""},{"id":519458776,"identity":"a83a7b56-5e95-4123-99e7-40a857f7a78d","order_by":4,"name":"Mohd Maqbool Mir","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Mohd","middleName":"Maqbool","lastName":"Mir","suffix":""},{"id":519458777,"identity":"a696de77-c064-45c2-9a5d-55a1e8b68a49","order_by":5,"name":"Amjad M Husaini","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Amjad","middleName":"M","lastName":"Husaini","suffix":""},{"id":519458778,"identity":"fbf07917-d0c0-474b-bf4e-a948c7488071","order_by":6,"name":"Sajad Majeed Zargar","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Sajad","middleName":"Majeed","lastName":"Zargar","suffix":""},{"id":519458779,"identity":"33d42562-9047-495b-8cad-d058a0e1066d","order_by":7,"name":"Parveena Bano","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Parveena","middleName":"","lastName":"Bano","suffix":""},{"id":519458780,"identity":"2950f15b-a56d-44b1-ad87-358cd1dbf21f","order_by":8,"name":"Saima Hamid","email":"","orcid":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","correspondingAuthor":false,"prefix":"","firstName":"Saima","middleName":"","lastName":"Hamid","suffix":""}],"badges":[],"createdAt":"2025-09-23 13:30:30","currentVersionCode":1,"declarations":{"humanSubjects":true,"vertebrateSubjects":false,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":true,"humanSubjectConsent":true,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-7694933/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7694933/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":92143394,"identity":"839eb886-670e-41b3-ae85-f8c382ccfc2f","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":15986464,"visible":true,"origin":"","legend":"","description":"","filename":"Papr.docx","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/1beeab115edf8deb05782eb4.docx"},{"id":92143341,"identity":"e4fa133e-f24c-49cb-87a3-5a037fa2e442","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"json","order_by":1,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":342,"visible":true,"origin":"","legend":"","description":"","filename":"rs7694933.json","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/6eca01ec07a6296b272a2976.json"},{"id":92145427,"identity":"8e445960-f5ac-4a20-ae1f-600240ac4f34","added_by":"auto","created_at":"2025-09-25 06:56:30","extension":"xml","order_by":2,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":88252,"visible":true,"origin":"","legend":"","description":"","filename":"rs76949330enriched.xml","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/74d69600333036f61ad10866.xml"},{"id":92143344,"identity":"a267dbb7-a212-451f-a19b-6362f6202a71","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"jpeg","order_by":3,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":1074,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/260c5ff46552480b28c76d36.jpeg"},{"id":92144563,"identity":"a5df2687-82a2-4d89-96ab-53fad885916a","added_by":"auto","created_at":"2025-09-25 06:48:30","extension":"png","order_by":4,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":553098,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage10.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/861dc13ad563c54f21139ba6.png"},{"id":92143348,"identity":"15b34f67-79b1-43d9-957c-f2395e8758b6","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"jpeg","order_by":5,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":658297,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage11.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/d426db78d6358f7cbef90a33.jpeg"},{"id":92144561,"identity":"92c09290-50f9-4daa-8c92-c7d379d2d2dd","added_by":"auto","created_at":"2025-09-25 06:48:30","extension":"jpeg","order_by":6,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":567001,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage12.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/59c92287563b2e0b6f6c2d7e.jpeg"},{"id":92143396,"identity":"e9d81a7a-5700-46a6-a212-1fc3bd34acbd","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":7,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":356851,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage13.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/0b8d53b8976a15fa4f553f6c.png"},{"id":92143366,"identity":"a512a60d-d432-4bbd-b740-0e81ed259e96","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":8,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":202069,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage14.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/6793bec878bfda5fd3897c75.png"},{"id":92143376,"identity":"b8956a52-55bb-4370-9e56-6954ce5b649a","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":9,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":404709,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage15.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/ab62c8dca6015f47bcbb73ef.png"},{"id":92144265,"identity":"f48fef7a-f78a-4cc2-993e-e7ae6b17562e","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":10,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":556569,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage16.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/5de8cb66a433b46540388684.png"},{"id":92143360,"identity":"1c7d1d86-a07f-4e59-be2a-a6f6312eda8b","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":11,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":328400,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage17.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/0517ab5c0e448e0fb062c5d0.png"},{"id":92143361,"identity":"d48873c5-3649-4225-9f83-3edba4b2b8a0","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":12,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":834244,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage18.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/acc6e51f22795422117e46dc.png"},{"id":92143393,"identity":"227f93ea-de3d-4372-b10b-6855ec52d6fd","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"jpeg","order_by":13,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":698779,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage19.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/3f3222cd13b343fb4d82697e.jpeg"},{"id":92145428,"identity":"f0e9773f-426c-4e81-bc79-7edf1b338fa9","added_by":"auto","created_at":"2025-09-25 06:56:30","extension":"png","order_by":14,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":270322,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/a6fbdc7ef8af81dcba0438e6.png"},{"id":92143363,"identity":"47466ab7-8e2a-4566-bf18-987cd1989b48","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"jpeg","order_by":15,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":385508,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage20.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/b1adde8e5236b0e9cd52be35.jpeg"},{"id":92144564,"identity":"0c09ca2c-94c5-4bdf-90df-315b0189caa5","added_by":"auto","created_at":"2025-09-25 06:48:30","extension":"png","order_by":16,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":278147,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage21.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/80d286821d887714063f8bc6.png"},{"id":92143356,"identity":"82a264a0-5837-46f5-bf4c-87100eae7190","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":17,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":325943,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage22.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/d4b2606dfa402579d48b3ada.png"},{"id":92143353,"identity":"8fc3ca9b-ffab-4766-ba4f-98fa2651a56c","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":18,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":331686,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage23.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/7f0375ef37cb3b1963740cb8.png"},{"id":92143358,"identity":"993081fc-902c-403b-952b-b813808a7254","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"jpeg","order_by":19,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":592032,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage24.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/ee0dd5805f41d6c0c135371e.jpeg"},{"id":92144277,"identity":"483e4376-43ef-41b9-ab82-201894939633","added_by":"auto","created_at":"2025-09-25 06:40:32","extension":"jpeg","order_by":20,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":11180,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage25.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/da9ac8e3625b97d739bbc6a5.jpeg"},{"id":92143362,"identity":"47b69480-2e99-4dd8-805e-9f4b53543fbe","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"jpeg","order_by":21,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":17235,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage26.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/1b3dd74bdf708b6a86c22156.jpeg"},{"id":92144262,"identity":"0269d2e3-2aca-467d-b086-c0456a68ef2d","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"jpeg","order_by":22,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":179240,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage27.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/4ea54658004947f76047a851.jpeg"},{"id":92144256,"identity":"395fbc63-5c12-41ac-9ff2-25c99f19dd1f","added_by":"auto","created_at":"2025-09-25 06:40:30","extension":"png","order_by":23,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":258655,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/f3e085eb65261174ad150307.png"},{"id":92144276,"identity":"ca7b6e2e-2f21-4e3e-9a05-6868e46976db","added_by":"auto","created_at":"2025-09-25 06:40:32","extension":"png","order_by":24,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":201614,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/b764e28c402a7441cf4938c0.png"},{"id":92143370,"identity":"1e608f0f-d474-4891-851b-bd1fceca2113","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"jpeg","order_by":25,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":846145,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage5.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/251c54fb3c9c4c87eb89244d.jpeg"},{"id":92144272,"identity":"e8c2cdd3-24be-44a5-8281-b6b5f1da681c","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":26,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":206405,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/1df2fb13ec8caff87cc4e1f7.png"},{"id":92143373,"identity":"60c1c094-a809-4215-aa94-bd9f8a9e2090","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":27,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":246629,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage7.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/32fdda2bbe60072a78089ec4.png"},{"id":92144566,"identity":"046eb6e1-90fb-477f-ad3b-a7f320189551","added_by":"auto","created_at":"2025-09-25 06:48:31","extension":"png","order_by":28,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":486679,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage8.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/76744e40b3d93fbf789e4398.png"},{"id":92144260,"identity":"4f894cfd-4e34-493c-a574-38aa2365152d","added_by":"auto","created_at":"2025-09-25 06:40:30","extension":"jpeg","order_by":29,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":451755,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage9.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/72fcd56d623fab133e46fd76.jpeg"},{"id":92144257,"identity":"c407e855-2ea4-4e9e-be80-0ad005fe11b7","added_by":"auto","created_at":"2025-09-25 06:40:30","extension":"jpeg","order_by":30,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":39572,"visible":true,"origin":"","legend":"","description":"","filename":"groupimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/012b52a35aaa5510e12ef6cd.jpeg"},{"id":92143404,"identity":"3a8bec72-bbf5-4679-944b-ca4c61508b54","added_by":"auto","created_at":"2025-09-25 06:32:32","extension":"png","order_by":31,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":935,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/b09f471a5fc1540c7ea70f67.png"},{"id":92143364,"identity":"f6254ab4-662a-44ed-9625-6de38db7f852","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":32,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":66022,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage10.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/979898024ee88f940b76a52a.png"},{"id":92143391,"identity":"e135c039-cc2c-4e2b-9f95-9f18ffebd93d","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":33,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":160347,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage11.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/5a4f2166e8b7aaf69e26b439.png"},{"id":92143399,"identity":"d66f07e0-a6c8-47cf-8044-1f182759a7f9","added_by":"auto","created_at":"2025-09-25 06:32:32","extension":"png","order_by":34,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":150090,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage12.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/3b58811c383e89e8128e5623.png"},{"id":92144271,"identity":"6f886325-021a-49ca-8408-4357258b9418","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":35,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":23336,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage13.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/0c71932451afd5b7dfdfe877.png"},{"id":92144568,"identity":"85b19658-a6e3-4999-84dc-5f5372bd791d","added_by":"auto","created_at":"2025-09-25 06:48:31","extension":"png","order_by":36,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":17857,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage14.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/fe34d64bb326b8eb867efc66.png"},{"id":92143368,"identity":"5a1f93c3-f412-4fd8-9ab5-18258117d167","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":37,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":42615,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage15.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/7229a42f6269fb339373eb01.png"},{"id":92144275,"identity":"aaa615f6-30b7-4151-a741-329dd8432885","added_by":"auto","created_at":"2025-09-25 06:40:32","extension":"png","order_by":38,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":58052,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage16.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/e24f36db45e02432f0148fa2.png"},{"id":92143372,"identity":"0a72f30d-fe64-4948-b1e9-1ad0c2f9b1cc","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":39,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":38958,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage17.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/1dfd2012d3ce8e65f10fb2e4.png"},{"id":92143378,"identity":"9838e535-2edb-4029-a412-4f5ef3ecec1c","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":40,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":92373,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage18.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/70e574cb525a1d195eac7c7b.png"},{"id":92143375,"identity":"1f7377b4-567d-4355-b2e4-8262d0b240bf","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":41,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":174045,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage19.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/4abb97faabd8837c8441550f.png"},{"id":92144565,"identity":"1ec9913d-beb9-4689-8d6e-a3da541b854d","added_by":"auto","created_at":"2025-09-25 06:48:31","extension":"png","order_by":42,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":35319,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/0488afc3ec12bffd7bea3bd7.png"},{"id":92143367,"identity":"59ba1b91-631f-4d0c-bb3a-1586bd52933d","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":43,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":112512,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage20.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/2b22f2fe9d1fab448bc7a949.png"},{"id":92143401,"identity":"8bb0b53e-83e4-4a5d-9bc2-2ce8b76fe850","added_by":"auto","created_at":"2025-09-25 06:32:32","extension":"png","order_by":44,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":42114,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage21.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/ffa7567c84973761d299b4b9.png"},{"id":92144263,"identity":"7e30ef1d-8359-45d4-8b8c-6b1d6a9c0197","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":45,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":31633,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage22.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/cae7f77257f064a5d7eb2390.png"},{"id":92143408,"identity":"eb78473c-e162-4a73-906e-cbff3976e397","added_by":"auto","created_at":"2025-09-25 06:32:32","extension":"png","order_by":46,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":29254,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage23.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/90458ca4421b2abbf3c531a0.png"},{"id":92144567,"identity":"59979a26-8f09-41bc-8a50-19c16c0f170f","added_by":"auto","created_at":"2025-09-25 06:48:31","extension":"png","order_by":47,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":140273,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage24.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/445c4c6b81d31cb7a76728a5.png"},{"id":92143380,"identity":"19ee7bf2-bb66-4357-a26b-df3259a79612","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":48,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":31421,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage25.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/36cf5bd1b172090f04bc70ca.png"},{"id":92144269,"identity":"b072e225-f6db-43d5-97b9-c3c3de19e166","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":49,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":44843,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage26.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/6e72619454ebbb879f4258f1.png"},{"id":92143403,"identity":"9cad7729-55b3-4cd1-ac61-a60963b999b2","added_by":"auto","created_at":"2025-09-25 06:32:32","extension":"png","order_by":50,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":90877,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage27.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/48311cfed60cef07db80cf1e.png"},{"id":92143388,"identity":"1905f4be-2107-40e7-9b46-414d85cb1202","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":51,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":33444,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/b3e6d19469d960cb781312e1.png"},{"id":92144273,"identity":"cf023735-3511-44b4-8648-c58e8cc66550","added_by":"auto","created_at":"2025-09-25 06:40:32","extension":"png","order_by":52,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":21898,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/0d7708313043ff522fc790ae.png"},{"id":92143387,"identity":"d1f4a89a-b279-4717-9c20-22066c7b85f4","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":53,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":207597,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/7aca338d0a673cf5c0c66948.png"},{"id":92143386,"identity":"8113039c-1778-4a34-9680-474e6dc9d27b","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":54,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":20327,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/c46065cc768e93b2de4c96be.png"},{"id":92144267,"identity":"8110f413-aea3-47f9-9738-c64b7c43be77","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":55,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":26923,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage7.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/888a9cc4cce83b21c0bbb219.png"},{"id":92144270,"identity":"bf023577-6698-4eaa-9641-5c3eb42e2682","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"png","order_by":56,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":53623,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage8.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/b73b132c66abcd4717541534.png"},{"id":92143382,"identity":"4a7809bc-c5b3-4088-9108-34ec0536e7db","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":57,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":156239,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage9.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/d7971b06043de15ba74bd4c9.png"},{"id":92143377,"identity":"82015833-8ecc-48e1-bd33-c0bace65deb4","added_by":"auto","created_at":"2025-09-25 06:32:31","extension":"png","order_by":58,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":16995,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinegroupimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/7076fc4e089f7f6dc514e407.png"},{"id":92143406,"identity":"8dabbbcd-beac-438c-89c7-7e4ead34ded5","added_by":"auto","created_at":"2025-09-25 06:32:32","extension":"xml","order_by":59,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":86782,"visible":true,"origin":"","legend":"","description":"","filename":"rs76949330structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/e4b57ad9f5e377bb93d0167c.xml"},{"id":92144268,"identity":"91ee61c3-a677-49f0-a21b-4f46ee85da7c","added_by":"auto","created_at":"2025-09-25 06:40:31","extension":"html","order_by":60,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":98740,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/f94a6b03fd112bc959415260.html"},{"id":92144248,"identity":"eae26c34-85b6-42be-baf1-4255cb38b559","added_by":"auto","created_at":"2025-09-25 06:40:30","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":520885,"visible":true,"origin":"","legend":"\u003cp\u003eMap illustrates the Kashmir Valley region, emphasizing three specific districts where \u003cem\u003eAndrena \u003c/em\u003espps were gathered.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/0956404fc39d04c154b9a300.png"},{"id":92143340,"identity":"812bbf4d-76f8-4004-a301-025b105c46d5","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":46834,"visible":true,"origin":"","legend":"\u003cp\u003eExtracted DNA (A) and PCR (B) of four different Andrenid spp.\u003c/p\u003e","description":"","filename":"2.2.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/43fade058aea6548287f3903.png"},{"id":92143343,"identity":"4450212e-1215-45f6-9148-ec32668dda35","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":63507,"visible":true,"origin":"","legend":"\u003cp\u003ePhylogenetic tree of different Andrenid spp.4 species identified using COXI.\u003c/p\u003e","description":"","filename":"3.3.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/7be6c5ef63d2ffc96daa0462.png"},{"id":92143346,"identity":"582b35ac-a2c9-4ee3-a3ec-aa40cb5d41b6","added_by":"auto","created_at":"2025-09-25 06:32:30","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":679713,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUnnumbered image in the Result section.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA:Bodylength,B.Head:i;Headlength,ii:Headwidth,iii:Clypeuslength,iv:Clypeus width,v:UICD,vi:LICD,vii:FOVlength,viii,FOVwidth,ix:Antennalsocketmaximum diameter. C. Thorax, D: Abdomen upper side E. Abdomen lower side, F. Fore wing, G; Hind wing, H; Antennal segment length: i; scape, ii; Pedicle, iii; Flagellum, I: Fore leg, J: Mid leg, K: Hind leg.\u003c/p\u003e","description":"","filename":"unnumber1.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/f8a7fe6924c311b483e6b914.png"},{"id":92144249,"identity":"4fefd348-ed56-4818-8214-91fb02835e32","added_by":"auto","created_at":"2025-09-25 06:40:30","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":747052,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUnnumbered image in the Result section.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA. Body length, B. Head: i; Head length, ii; Head width ,iii; Clypeus length, iv; Clypeus width, v; UICD ,vi; LICD, vii; FOV length, viii, FOV width, C:Thorax,D:Abdomen upper side, E. Abdomen lower side, F. Fore wing G.Hind wing ,H,Antennal segment length,i:scape,ii:pedicle,iii,Flagellum,I.Fore leg,J.Mid leg,K.Hind leg,L.Genitalia,M:S7,N:S8\u003c/p\u003e","description":"","filename":"unnumber2.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/129244afda0ad94f581bf738.png"},{"id":92144253,"identity":"25422771-d048-427a-9afc-199da54f2ecc","added_by":"auto","created_at":"2025-09-25 06:40:30","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":1125339,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUnnumbered image in the Result section.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA.Body length,B.Head,i:Head length, ii;Head width,iii;Clypeus length,iv;Clypeus width,v;UICD,vi;LICD,vii;FOV length,viii;FOV width,ix;Antennal segment maximum diameter,C.Thorax,D.Abdomen upper side,E.Abdomen lower side,F.Fore wingG.Hind wing ,H.Antennal segment length;I;Scape,ii;Pedicle,iii;Flagellum,I.Fore leg,J.Mid leg,K.Hind leg\u003c/p\u003e","description":"","filename":"unnumber3.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/b52c6f071452332b068c29b6.png"},{"id":92144559,"identity":"9d8ff39b-a494-4d79-9b45-3e8339e34103","added_by":"auto","created_at":"2025-09-25 06:48:30","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":742924,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUnnumbered image in the Result section.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA.Body length,B.Head ,i:Head length, ii: Head width, iii :Clypeus width,iv:Clypeus length,v:UICD,vi:LICD,vii:FOV length,viii,FOV Width , ix: Antennal socket maximum dia, C.Thorax,D:Abdomen upper side , E:Abdomen lower side, F:Fore wing, G, Hind wing ,H: Antennal segment length: i:scape,ii:pedicle,iii:Flagellum,I.Fore leg,J. Mid leg, K. Hind leg\u003c/p\u003e","description":"","filename":"unnumber4.png","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/83a982d95d9dda8049874cf9.png"},{"id":92145652,"identity":"b8f50d72-c55d-40b0-b437-92ed654e1b10","added_by":"auto","created_at":"2025-09-25 07:04:33","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":5268976,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7694933/v1/909f1e77-4e5a-49dd-b38c-18a355dbcdc7.pdf"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003e\u003cstrong\u003eFrom Mountains to Molecules: Decoding the Hidden Diversity of Andrena Bees in the Himalayan Wilderness\u003c/strong\u003e\u003c/p\u003e","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eBees have fascinated humans for centuries, not only for their complex social organization but also for their vital role as pollinators. Unlike solitary insects that complete their life cycle independently, social bees live in colonies with distinct castes that cooperate for colony survival (Michener, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Globally, bees are considered the most important pollinators, visiting the majority of flowering plants. It is estimated that nearly 75% of crops of global significance and about 90% of fruits, vegetables, and horticultural crops depend on insect pollination, with bees being the primary contributors (Klein et al., 2007; Ollerton, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eWhile honey bees (\u003cem\u003eApis\u003c/em\u003e spp.) are widely recognized for their pollination efficiency due to traits such as floral constancy, colony strength, and manageability (Delaplane \u0026amp; Mayer, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Abrol, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2012\u003c/span\u003e), non-\u003cem\u003eApis\u003c/em\u003e bees play an equally crucial role. These solitary and semi-social bees often outperform honey bees in specific conditions because of unique behaviors like buzz pollination, longer proboscis, faster foraging, and effective tripping mechanisms (Willmer et al., 2017; Rader et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Their role has become more critical in recent decades with the decline of honey bee populations caused by habitat loss, monoculture farming, pesticide use, and emerging diseases (Goulson et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2015\u003c/span\u003e).Among solitary bees, the genus \u003cem\u003eAndrena\u003c/em\u003e, commonly referred to as mining bees, is one of the largest bee genera, with more than 1,500 described species globally (Michener, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Ascher \u0026amp; Pickering, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). They are especially important in temperate regions, where they serve as primary pollinators of both wild flora and cultivated crops (Wood et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Recent taxonomic revisions have increased the number of \u003cem\u003eAndrena\u003c/em\u003e species reported from India to 36, reflecting the country\u0026rsquo;s underestimated diversity (Wood \u0026amp; Gupta, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). These bees, typically ground-nesting, display a wide range of body sizes and colorations, including metallic and reddish forms, and are adapted to diverse ecological niches. Accurate identification of \u003cem\u003eAndrena\u003c/em\u003e species remains challenging due to morphological similarities across taxa. To address this, molecular tools such as DNA barcoding have emerged as reliable methods (Bogusch et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). The mitochondrial cytochrome oxidase I (COI) gene has been widely adopted for species-level identification and phylogenetic analysis in bees (Hebert et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2003\u003c/span\u003e; Kek et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). DNA barcoding has proven particularly valuable for resolving cryptic taxa and validating species records (Moroń et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Chen et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). In India, pioneering studies have used COI barcoding to document species like \u003cem\u003eAndrena agilissima\u003c/em\u003e, thereby enhancing regional biodiversity databases (Chandra et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Furthermore, recent methodological advances, including improved primer design and the use of degraded DNA from museum specimens, are expanding the scope of barcoding in bee systematics (Mitchell, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Ashfaq et al., 2023).Thus, combining morphological and molecular approaches provides a powerful framework for understanding the diversity, taxonomy, and ecological roles of \u003cem\u003eAndrena\u003c/em\u003e bees. This integration is particularly important in regions like India, where the diversity of solitary bees remains underexplored but ecologically significant.\u003c/p\u003e"},{"header":"MATERIAL AND METHODS","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eStudy area:\u003c/h2\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eAn extensive field survey was undertaken to document the diversity of \u003cem\u003eAndrena\u003c/em\u003e species across three representative districts of the Kashmir Valley: Srinagar, Pulwama, and Budgam (Fig.\u0026nbsp;1). To ensure wide ecological coverage, three sites were strategically selected within each district. In Srinagar, surveys were carried out at Shalimar, Chatterhama, and Dhara; in Budgam, sampling was conducted at Chadoora, Badipora, and Wahabpora; while in Pulwama, the locations included Malangpora, Newa, and Rajpora. Collections were performed during daylight hours, targeting both cultivated fields and surrounding wild flora to maximize the likelihood of capturing the full range of foraging \u003cem\u003eAndrena\u003c/em\u003e species. Particular attention was given to periods of peak bloom for major crops and naturally occurring bee forage plants, as these phenological windows represent the most active phases of bee visitation. Standard entomological techniques were employed to collect specimens, ensuring minimal damage for subsequent morphological and molecular identification.All collected material was processed and curated at the Research and Training Centre for Pollinators, Pollinizers, and Pollination Management (RTCPPPM), Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir (SKUAST-K), Shalimar. This facility provided the necessary infrastructure for specimen preservation, identification, and downstream molecular analyses, thereby serving as the focal point for the research program.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eCollection and Preservation method:\u003c/h3\u003e\n\u003cp\u003eSpecimens of \u003cem\u003eAndrena\u003c/em\u003e species were collected from different host plants such as apple, dandelion, poison hemlock, almond, and Persian speedwell across several districts of Jammu and Kashmir using a sweep net. The collected bees were immediately transferred to airtight glass killing jars charged with ethyl acetate. Ethyl acetate was chosen because it provides rapid euthanasia while maintaining the natural coloration of the insect and preventing hardening of body tissues, which is essential for subsequent taxonomic studies. After euthanasia, the specimens were carefully stretched, pinned, and mounted in insect collection boxes to ensure proper positioning and long-term preservation. To rehydrate dried specimens prior to examination, the pinned bees were placed in an airtight container with a moistened tissue or paper towel for 12\u0026ndash;24 hours. This softening process made handling and dissection easier. To prevent fungal or mold growth during rehydration, a few drops of phenol or thymol were introduced into the container, ensuring safe and hygienic preservation of the specimens for detailed morphological and morpho-molecular analyses.\u003c/p\u003e\n\u003ch3\u003eDissection and Mounting\u003c/h3\u003e\n\u003cp\u003eFor genitalia preparation, relaxed specimens were first placed in a Petri dish containing either distilled water or ethanol to soften the body tissues. Under a stereo microscope, the abdomen was gently held with fine forceps, and the terminal abdominal segments (typically segments VII\u0026ndash;VIII) were carefully detached by pulling backward with the aid of a minuten pin or micro-scissors.The excised abdomen was then immersed in a 10% potassium hydroxide (KOH) solution and heated at 60\u0026ndash;70\u0026deg;C for 15\u0026ndash;20 minutes. This maceration step dissolved soft tissues and cleared the muscles, thereby facilitating clear visualization of the genital structures. Following digestion, the material was rinsed several times with distilled water to remove residual KOH and to prevent further tissue degradation.The cleaned genitalia were mounted on a glass microscope slide using either glycerin (for temporary preparations) or Canada balsam (for permanent preparations). A cover slip was gently placed on top, ensuring the specimen was properly flattened without distortion, and the edges were sealed with clear nail polish or a synthetic mounting medium to avoid desiccation and fungal growth.Subsequently, morphological examinations were performed with a stereo zoom microscope (OLYMPUS SZX16). Standard morphometric terminology was employed, and the following parameters were recorded: body length; head length and width; thorax length and width; abdomen length and width; clypeus length and width; upper and lower interocular distances (UICD and LICD); maximum diameter of the antennal socket; forewing length and width; hind wing length and width; facial fovea (FOV) length and width; and lengths of individual antennal segments (AS). Ratios such as head length-to-width, FOV length-to-width, antennal segment 3-to-segment 1 (AS3/AS1), and UICD-to-LICD were also calculated. These measurements formed the basis for species-level identification and comparative taxonomic analyses.\u003c/p\u003e\u003cp\u003e\u003cb\u003eAuthentication of\u003c/b\u003e \u003cb\u003eAndrena\u003c/b\u003e \u003cb\u003espp. through DNA Barcoding\u003c/b\u003e\u003c/p\u003e\u003cp\u003eCollected specimens were preserved either in a dry state or in 100% ethanol to ensure adequate preservation of DNA for molecular studies. Genomic DNA was extracted from the legs and thorax of adult bees using the DNeasy Blood \u0026amp; Tissue Kit (Qiagen, Cat. No. 69504), following the manufacturer\u0026rsquo;s protocol. The quality of the extracted DNA was assessed on agarose gel electrophoresis, with ethidium bromide used for staining, and electrophoresis carried out for one hour. DNA concentration and purity were subsequently determined using a Nanodrop spectrophotometer.Species-level identification was performed using the mitochondrial cytochrome oxidase subunit 1 (COI) gene marker. PCR amplification of the COI region was carried out using species-specific primers: forward primer (5\u0026prime; ATTCAACCAATCATAAAGATATTGG 3\u0026prime;) and reverse primer (5\u0026prime; TAAACTTCTGGATGCTCCAAAAAATCA 3\u0026prime;) as described by Gurpreet et al. (2016). Each PCR reaction was performed in a total volume of 10 \u0026micro;l, consisting of 1 \u0026micro;l DNA template, 1.5 \u0026micro;l forward primer, 1.5 \u0026micro;l reverse primer, 1.3 \u0026micro;l nuclease-free water, 0.6 \u0026micro;l dNTP mix, 0.1 \u0026micro;l Taq polymerase, 2 \u0026micro;l MgCl₂, and 2 \u0026micro;l Taq buffer.(Fig.\u0026nbsp;2) The amplification protocol included an initial denaturation at 97\u0026deg;C for 5 minutes, followed by 40 cycles of denaturation at 94\u0026deg;C for 1.5 minutes, annealing at 52\u0026deg;C for 1 minute, and extension at 72\u0026deg;C for 1 minute, with a final extension step at 72\u0026deg;C.Amplified PCR products were sequenced bidirectionally using both forward and reverse primers on an ABI 3730XL DNA Analyzer (Applied Biosystems, Gene Technology). The resulting chromatograms were assembled and edited using Geneious v11.0.3, while sequence alignment and analysis were carried out in BioEdit. All sequences generated in this study were submitted to GenBank (Table\u0026nbsp;1). Remaining DNA samples were archived at \u0026minus;\u0026thinsp;30\u0026deg;C in the Molecular Laboratory of Biotechnology, SKUAST-K, Srinagar, and are available for future research upon request.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eSequenced Taxa and GeneBank Accession Numbers:\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eS.NO.\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eSpecies\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eSex\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eGeneBank Accession No.\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1.\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e\u003cem\u003eAndrena cineraria\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eMale\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eOP646814\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2.\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e\u003cem\u003eAndrena flavipes\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eFemale\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eOP672272\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3.\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e\u003cem\u003eAndrena pilipes\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eFemale\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eOP646792\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e4.\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e\u003cem\u003eAndrena bicolor\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eFemale\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eOP673542\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\n\u003ch3\u003eFloral Resources:\u003c/h3\u003e\n\u003cp\u003eInformation on the floral resources utilized by adult \u003cem\u003eAndrenid\u003c/em\u003e bees was systematically recorded from the sampling locations across three districts of the Kashmir Valley, namely Srinagar, Pulwama, and Budgam, during the course of the study. At each site, observations were made to identify the plant species that served as foraging sources for the bees. These floral resources were then classified into two categories: family-wise and crop-wise. The taxonomic identification of plant families and species was carried out with the assistance of a professional taxonomist, while additional verification and cross-checking were done using reliable online botanical databases and literature. This approach not only ensured accurate documentation of the bee plant associations but also provided insights into the diversity of floral resources available to \u003cem\u003eAndrena\u003c/em\u003e species across agricultural as well as wild landscapes of the valley.\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003eUpon morphological examination of these species, these were identified as:\u003cem\u003e\u0026nbsp;Andrena cineraria, Andrena flavipes, Andrena pilipes\u0026nbsp;\u003c/em\u003eand\u003cem\u003e\u0026nbsp;Andrena bicolor\u003c/em\u003e\u003cem\u003e.\u003c/em\u003eThis identification was authenticated through molecular means also.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAndrena\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e(\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eEuAndrena) bicolor\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena picicornis\u003c/em\u003e \u0026ndash;Kirby-1802\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena pilosula\u003c/em\u003e-Kirby -1802\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena gwynana\u003c/em\u003e \u0026ndash;Kirby -1802\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena proxima\u003c/em\u003e \u0026ndash;Smith -1847\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena aestiva\u003c/em\u003e \u0026ndash;Smith -1849\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena consimilis\u003c/em\u003e-Smith -1849\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena laeviuscula \u0026ndash;\u003c/em\u003eSchenck-1853\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMaterial examined:\u0026nbsp;\u003c/strong\u003e2♀, J\u0026amp;K, India: 2♀, Shalimar Srinagar, 34.148\u003csup\u003e0\u003c/sup\u003eN 74.883\u003csup\u003e0\u003c/sup\u003eE, 1588m, 19.IV.2021, Coll: Tamjeeda, Dandelion, Poison hemlock, Creeping thistle, Apple , Pear.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDistribution:\u0026nbsp;\u003c/strong\u003eIndia, Scotland, Ireland, Western Palaeartic from Fennoscandia South to Iberia and Corsica, East of Siberia, North Africa, Israel and Iran.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDiagnostic Characters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFemale (\u003c/strong\u003e♀)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAlso called as Gwynne\u0026rsquo;s mining bee. Females appear larger as compared to males. Head is black coloured with white \u0026ndash;black hair scattered all over the head. Thorax is usually black with golden white coloured hair scattered all over the thorax. Females with sparse pile of brownish hairs on the tergite that form weaker bands. Hind legs have dull orange colour.Mesosoma with orange yellow hairs, Trochanter of hind leg floculus brownish black, tibial scopa orange yellow, Metasomal disc with sparse punctuation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMeasurements (Mean\u0026plusmn;SE in mm):\u0026nbsp;\u003c/strong\u003eTotal body length: 15.16\u0026plusmn;1.29, Head length: 2.78\u0026plusmn;0.23, Head width: 3.49\u0026plusmn;0.29, Head: L/w: 0.79\u0026plusmn;0.06, Thorax length: 3.97\u0026plusmn;0.34, Thorax width: 3.23\u0026plusmn;0.28, Fore wing length: 7.86\u0026plusmn;7.87, Fore wing width: 2.76\u0026plusmn;0.23, Hind wing length: 7.28\u0026plusmn;0.61, Hind wing width: 1.90\u0026plusmn;0.17, Abdomen length: 5.62\u0026plusmn;0.48, Abdomen width: 3.88\u0026plusmn;0.32, Clypeus length: 1.34\u0026plusmn;0.11, Clypeus width: 2.29\u0026plusmn;0.19, UICD: 2.42\u0026plusmn;0.20, LICD: 2.41\u0026plusmn;0.20, UICD: LICD: 1.00\u0026plusmn;0.08, Scape length: 1.14\u0026plusmn;0.09, Pedicle length: 0.32\u0026plusmn;0.02, Flagellum length: 5.1\u0026plusmn;0.43, Antennal socket maximum diameter: 0.33\u0026plusmn;0.02, AS\u003csub\u003e3\u003c/sub\u003e: AS\u003csub\u003e1:\u0026nbsp;\u003c/sub\u003e4.47\u0026plusmn;0.38, FOV length: 1.08\u0026plusmn;0.09, FOV width: 0.58\u0026plusmn;0.05, FOV: L/W: 1.86\u0026plusmn;0.15 .\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003e\u0026nbsp;Andrena\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e(\u003cem\u003eMelAndrena) cineraria\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena cineraria ssp. Danuvia\u003c/em\u003e- E. Stockhert -1950\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena danuvia-\u003c/em\u003e E. Stoeckhert- 1950\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMaterial examined:\u0026nbsp;\u003c/strong\u003e4♂, J\u0026amp;K, India: 2♂: Newa Pulwama, 33.929\u003csup\u003e0\u003c/sup\u003eN 74.898\u003csup\u003e0\u003c/sup\u003eE, 1588m, 9.III.2022, Coll: Tamjeeda, Pear, Poison hemlock, Yellow star of Bethlehem, 2♂: Badipora Budgam, 33.917\u003csup\u003e0\u003c/sup\u003eN 74.796\u003csup\u003e0\u003c/sup\u003eE, 1610m, 26.III.2021, Coll: Tamjeeda, Dandelion, Almond, Persian speedwell.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDistribution:\u0026nbsp;\u003c/strong\u003eIndia, Scotland, Ireland, Southern England, South of Iberia, East to Northern China.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDiagnostic Characters:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMale (\u003c/strong\u003e♂\u003cstrong\u003e):\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAlso called as ashy brown mining bee. Males appear to be smaller than female. They have golden brown hair on the head. Thorax is black in colour with golden hair on the side of thorax. Abdomen black in colour with scattered golden hair all over tergum and sterum. Female have 12 segments in antennae, males have 13 segments in antennae. Genitalia broader than \u003cem\u003eAndrena gravida\u003c/em\u003e .\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMeasurements (Mean\u0026plusmn;SE in mm):\u0026nbsp;\u003c/strong\u003eTotal body length: 11.87\u0026plusmn;1.00, Head length: 3.40\u0026plusmn;0.29, Head width: 4.24\u0026plusmn;0.37, Head: L/w: 0.80\u0026plusmn;0.06, Thorax length: 3.43\u0026plusmn;0.29, Thorax width: 2.89\u0026plusmn;0.24, Fore wing length: 7.74\u0026plusmn;7.74, Fore wing width: 2..57\u0026plusmn;0.21, Hind wing length: 2.52\u0026plusmn;0.21, Hind wing width: 2.52\u0026plusmn;0.21, Abdomen length: 4.76\u0026plusmn;0.40, Abdomen width: 2.85\u0026plusmn;0.24, Clypeus length: 1.55\u0026plusmn;0.13, Clypeus width: 3.12\u0026plusmn;0.27, UICD: 2.93\u0026plusmn;0.24, LICD: 2.97\u0026plusmn;0.25, UICD: LICD: 0.98 \u0026plusmn;1.01, Scape length: 1.04\u0026plusmn;0.08, Pedicle length: 0.27\u0026plusmn;0.02, Flagellum length: 4.46\u0026plusmn;0.37, Antennal socket maximum diameter: 0.45\u0026plusmn;0.03, AS\u003csub\u003e3\u003c/sub\u003e: AS\u003csub\u003e1\u003c/sub\u003e: 4.28\u0026plusmn;0.36, FOV length: 1.25\u0026plusmn;0.10, FOV width: 0.75\u0026plusmn;0.07, FOV: L/W: 1.66\u0026plusmn;0.14.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAndrena\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e(\u003cem\u003ePlastAndrena) pilipes\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena carbonaria\u003c/em\u003e- Linneaus- 1767\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMaterial examined:\u0026nbsp;\u003c/strong\u003e17♀: J\u0026amp;K, India: 4♀, Dhara, Srinagar, 4.I.2022, 34.169\u003csup\u003e0\u003c/sup\u003eN 74.919\u003csup\u003e0\u003c/sup\u003eE, 1776m, Coll: Tamjeeda, Pear, 5♀, Newa Pulwama, 33.929\u003csup\u003e0\u003c/sup\u003eN \u0026nbsp; 74.898\u003csup\u003e0\u003c/sup\u003eE, 1588m, 9.III.2022, Coll: Tamjeeda, Dandelion, 2♀, Rajpora, Pulwama, 33.822\u003csup\u003e0\u003c/sup\u003eN 74.853\u003csup\u003e0\u003c/sup\u003eE, 1545m, Poison hemlock , Yellow star of Bethlehem, 6♀, Badipora Budgam, 33.917\u003csup\u003e0\u003c/sup\u003eN 74.796\u003csup\u003e0\u003c/sup\u003eE, 1610m, 4.V.2020, Coll: Tamjeeda, Creeping thistle.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDistribution:\u0026nbsp;\u003c/strong\u003eIndia, Africa, Europe, Turkey and Northern Africa.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDiagnostic Characters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFemale (\u003c/strong\u003e♀)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eHoney bee sized black coloured \u003cem\u003eAndrena\u003c/em\u003e species.Head is black in colour with light black coloured scattered hair, Thorax is black is colour with light golden coloured hair on the side of thorax, Abdomen is black in colour with light black scattered hair on tergum with orange coloured hair on sterum. Hind wing is larger as compared to fore wing. Golden coloured hair scattered over legs. It is a polyletic species.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMeasurements (Mean\u0026plusmn;SE in mm):\u0026nbsp;\u003c/strong\u003eTotal body length: 4.73\u0026plusmn;0.40, Head length: 2.68\u0026plusmn;0.22, Head width: 3.20\u0026plusmn;0.28, Head: L/w: 0.83\u0026plusmn;0.07, Thorax length: 3.46\u0026plusmn;0.29, Thorax width: 3.23\u0026plusmn;0.28, Fore wing length: 4.99\u0026plusmn;4.99, Fore wing width: 1.90\u0026plusmn;0.17, Hind wing length: 6.56\u0026plusmn;0.55, Hind wing width: 1.79\u0026plusmn;0.15, Abdomen length: 3.59\u0026plusmn;0.30, Abdomen width: 2.81\u0026plusmn;0.24, Clypeus length: 1.13\u0026plusmn;0.09, Clypeus width: 1.80\u0026plusmn;0.15, UICD: 2.06\u0026plusmn;0.18, LICD: 1.82\u0026plusmn;0.15, UICD: LICD: 1.13\u0026plusmn;0.09, Scape length: 3.08\u0026plusmn;0.26, Pedicle length: 0.22\u0026plusmn;0.01, Flagellum length: 5.12\u0026plusmn;0.43, Antennal socket maximum diameter: 0.43\u0026plusmn;0.03, AS\u003csub\u003e3\u003c/sub\u003e: AS\u003csub\u003e1\u003c/sub\u003e: 1.66\u0026plusmn;0.14, FOV length: 1.02\u0026plusmn;0.09, FOV width: 0.86\u0026plusmn;0.08, FOV: L/W: 1.18\u0026plusmn;0.10.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003e\u0026nbsp;Andrena\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e(\u003cem\u003eZonAndrena) flavipes\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena fulvicrus-\u003c/em\u003eKirby-\u003cem\u003e1802\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena extricate \u0026ndash;\u003c/em\u003eSmith \u003cem\u003e-1802\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena contigua \u0026ndash;\u003c/em\u003eKirby\u003cem\u003e\u0026nbsp;-1802\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena flavipes var. cinerascens\u003c/em\u003e \u0026ndash;Eversmann- 1852\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena interrupta\u003c/em\u003e- Schenck- 1869\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMaterial examined:\u0026nbsp;\u003c/strong\u003e19♀, J\u0026amp;K, India: 5♀, Shalimar Srinagar, 34.148\u003csup\u003e0\u003c/sup\u003eN 74.883\u003csup\u003e0\u003c/sup\u003eE, 1588m, 2.III.2021, Coll: Tamjeeda, 4♀, Chatterhama Srinagar, 34.184\u003csup\u003e0\u003c/sup\u003eN 74.868\u003csup\u003e0\u003c/sup\u003eE, 1592m, 9.III.2021, Coll: Tamjeeda, Apple and Dandelion, 2♀, Newa Pulwama 33.929\u003csup\u003e0\u003c/sup\u003eN 74.898\u003csup\u003e0\u003c/sup\u003eE, 1588m, 8.IV.2022, Coll: Tamjeeda, Pear, 3♀, Malangpora, Pulwama 33.893\u003csup\u003e0\u003c/sup\u003eN 74.982\u003csup\u003e0\u003c/sup\u003eE, 1626m, 8.IV.2022, Coll: Tamjeeda, Almond, 5♀, Chadoora, Budgam, 33.945\u003csup\u003e0\u003c/sup\u003eN 74.796\u003csup\u003e0\u003c/sup\u003eE, 1624m, 9.III.2022, Coll: Tamjeeda, Apple.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDistribution:\u0026nbsp;\u003c/strong\u003eIndia, Turkey, Southern England and South Coast of Wales.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDiagnostic Characters:\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;Female (\u003c/strong\u003e♀)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMedium sized \u003cem\u003eAndrena\u003c/em\u003e spps. Females have buffish white hair bands that fully occupy the apical depressions of tergite 1-4 with short black hairs between the bands, slight light brownish haired thorax.They have pygidial plate which is pointed apically. Golden coloured hairs on tibia and tarsus of fore and hind leg.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMeasurements (Mean\u0026plusmn;SE in mm):\u0026nbsp;\u003c/strong\u003eTotal body length: 3.70\u0026plusmn;0.31, Head length: 2.16\u0026plusmn;0.19, Head width: 2.58\u0026plusmn;0.22, Head: L/W: 0.83\u0026plusmn;0.07, Thorax length: 2.73\u0026plusmn;0.23, Thorax width: 2.47\u0026plusmn;0.21, Fore wing length: 6.95\u0026plusmn;0.59, Fore wing width: 2.14\u0026plusmn;0.19, Hind wing length: 6.06\u0026plusmn;0.51, Hind wing width: 1.66\u0026plusmn;0.14, Abdomen length: 4.55\u0026plusmn;0.39, Abdomen width: 2.60\u0026plusmn;0.22, Clypeus length: 1.02\u0026plusmn;0.85, Clypeus width: 1.56\u0026plusmn;0.13, UICD: 1.74\u0026plusmn;0.14, LICD: 1.65\u0026plusmn;0.13, UICD: LICD: 1.05\u0026plusmn;0.08, Scape length: 2.87\u0026plusmn;0.24, Pedicle length: 0.22\u0026plusmn;0.01, Flagellum length: 5.43\u0026plusmn;0.46, Antennal socket maximum diameter: 0.29\u0026plusmn;0.02, AS\u003csub\u003e3\u003c/sub\u003e: AS\u003csub\u003e1\u003c/sub\u003e: \u0026nbsp;1.89\u0026plusmn;0.16, FOV length: 0.95\u0026plusmn;0.08, FOV width: 0.29\u0026plusmn;0.02, FOV: L/W: 3.27\u0026plusmn;0.27 .\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMolecular Characterization of\u0026nbsp;\u003c/strong\u003e\u003cem\u003eAndrena\u003c/em\u003e\u003cstrong\u003e\u0026nbsp;spp\u003c/strong\u003e:\u003c/p\u003e\n\u003cp\u003eFor molecular characterization, the mitochondrial \u003cstrong\u003eCytochrome Oxidase I (COX1)\u003c/strong\u003e gene marker was employed. Polymerase Chain Reaction (PCR) was carried out to amplify the target region, and the resulting products were subjected to sequencing. The raw sequences obtained were carefully edited, trimmed, and aligned using \u003cstrong\u003eBioEdit software\u0026nbsp;\u003c/strong\u003eto ensure accuracy and remove ambiguities.To study evolutionary relationships among the \u003cem\u003eAndrena\u003c/em\u003e species, \u003cstrong\u003eMEGA10 software\u003c/strong\u003e was used for constructing phylogenetic trees and performing evolutionary analyses (Fig. 1). Both BioEdit and MEGA programs facilitated the alignment and comparison of the generated sequences with already available reference sequences.The curated nucleotide sequences were then submitted to the \u003cstrong\u003eNCBI GenBank database\u003c/strong\u003e, where accession numbers were assigned. These accession numbers now serve as permanent references for the isolated \u003cem\u003eAndrena\u003c/em\u003e species. To verify species identity, the acquired sequences were analyzed using the \u003cstrong\u003eBLASTN tool\u003c/strong\u003e of NCBI, which compared them against global sequence repositories. This homology search confirmed the taxonomic position of the studied species. Finally, the nucleotide sequences of the COX1 gene for all the characterized \u003cem\u003eAndrena\u003c/em\u003e species are presented below, providing a molecular-level reference for future comparative and taxonomic studies.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1.1: Sequencing of the PCR products of the COX-1marker regions of \u003cem\u003eAndrena\u003c/em\u003e species:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;Following are the sequences obtained after amplification of COX-1 region of \u003cem\u003eAndrena\u003c/em\u003e species:\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1.\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eAndrena bicolor:\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e(\u003cstrong\u003eAccession number:\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eOP673542)\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eATATGAGCAGGTATAATTGGTGCCTCCCTTAGATTCATCATTCGTATAGAACTAAGAAACCCAGGTAGTTGAATTAACAATGATCAATTTATAACTCAATGTTACATCTCATGCTTTTATTATAATTTTTTTCATAGTAATACCATTTATAATTGGAGGATTCGGAAACTGACTCACACCATTAATA\u003cbr/\u003eCTAGGAGCACCTGATAGCTTTCCCTCGAATAAATAATATAAGATTTTGACTACTACCCCCATCAATTCTAATTATTTTAATAAGAATAGTTTTAAATTCAGGTTCTGGTACAGGATGAACAGTCTATCCCCCCCTTTCATCCTACGCCTTTCACCCATCATCATCAGTAGATTTAACAATTTTTT\u003cbr/\u003eCTCTTCATATTGCAGGTATTTCGTCAATTATAGGAGCAATTAATTTTATTGTAACAATTTTGAATATAAAAAATATTTCACTAAATTATGATCAAATACCACTATTCCCATGATCAGTATTTATCACTACAAT.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2. \u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eAndrena cineraria:\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e(\u003cstrong\u003eAccession number:\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eOP646814 )\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eATATTTCATCTTCGCTATATGAGCAGGAATAGTCGGTGCATCCCTAAGATTTATCATCCGAATAGAATTAAGAAACCCAGGTATATGAATCA\u003cbr/\u003eACAATGATCAATTATATAATTTAATCGTTACCTGGCACGCGTTTATTATAATTTTCTTCATAATAATGCCATTCATAATCGGAGGTTTCG\u003cbr/\u003eGAAACTGACTCACACCGTTAATAATAGGAGCGCCAAATATGGCGTTTCCTCGTATAAATAATTTGAGATTTTGATCATTGCCCCCTTCAATTCTA\u003cbr/\u003eATAATTTTAATAAGAATAATCCTAAATTCGGGGTCTGGTACGGGATGAACAGTTTATCCACCACTGTCATCATACTCATTCCATC\u003cbr/\u003eCATCATCATCAGTTGATTTAACAATTTTTTCACTTCACATTGCTGGTATTTCATCAATTATAGGGGCGATCAACTTTATTGTTACAATTCTTAATATAAA\u003cbr/\u003eAAATATTTCATTTAATTATGATCAAATGCCACTGTTTTTTTGGTCTGTCTTGTTAACAGCATTTCTCTTATTAATCTCATTACAAGTTCTAGCTGGAGCAATTACAATATTACTATCAGATCGAAACTT.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eAndrena flavipes:\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e(\u003cstrong\u003eAccession number:\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eOP672272 )\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eATCTTCGCCATATGAGCGGGCATAATCGGAACCTCACTAAGATTTATTATCCGAATGGAATTAAGAAATCCAGGAAGCTGAATCAACAATGA\u003cbr/\u003eTCAAATTTATAATTCAATTGTAACCTCACACGCTTTCATTATAATTTTCTTCATAGTTATACCATTCATAATCGGAGGTTTCGGAAACTGACTCA\u003cbr/\u003eCACCGTTAATATTAGGAGCGCCCGACATGGCGTTCCCGCGAATAAACAACATAAGGTTTTGATTGTTACCACCCTCGATCTTAAT\u003cbr/\u003eCATCTTAATAAGAATAGTTCTAAATTCGGGATCGGGAACAGGATGAATTTACCCACCACTATCCTACTCATTCCACCCATCCTCATCAGTAGATCTAACA\u003cbr/\u003eATCTTCTCACTTCACATTGCAGGTGTATCATCAATTATAGGTGCAATCAACTTTATCGTAACAATCCTAAATATAAAAAA\u003cbr/\u003eTATCTCAATAAATTATGATCAACTACCACTATTCCCATGATCAGTATTTATTACCACAATCCTACTACTAATTTCCTTGCCAGTTTTAGCTGGTG\u003cbr/\u003eCCATCACAATACTCCTATCAGATCGTAATTTAAACTCATCATTCTTCGATCCCATG.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e4.\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eAndrena pilipes:\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e(Accession number:\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eOP646792)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAAATTTATAATACAATCGTAACCTCTCACGCCTTTATTATAATTTTCTTCATAGTTATGCCATTCATAATCGGAGGTTTCGGAAACTGACTC\u003cbr/\u003eACACCATTAATATTAGGAGCGCCCGACATGGCTTTCCCGCGAATAAAATAATATAAGATTTTGGCTACTCCCCCCTTCAATTCTAATTAT\u003cbr/\u003eTTTAACAAAAATAATATTAAATTCGGGGTCCGGAACAGGTTGAACAATTTACCCTCCATTATCATCATATTCTTATCACCCATCATCCTC\u003cbr/\u003eAACAGACCTAACAATTTTCTCACTGCACATTGCAGGAATTTCATCAATTATAGGAGCAATCAATTTTATTGTAACAATTCTTAACATAAAAAATA\u003cbr/\u003eTTTCAATAAATTATGATCAACTACCCCTATTCCCATGATCAGTATTTATTACCACAATTCTTCTATTAATTTCTCTACCAGTTCTAGCTGGAGCTATT.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1. \u0026nbsp; Exploration of Intra and interspecific species variation among collected \u003cem\u003eAndrena\u003c/em\u003e species by Phylogenetic Analysis:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe \u003cstrong\u003eNeighbor-Joining (NJ) phylogenetic tree\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003ewas constructed based on a \u003cstrong\u003e658 bp fragment of the COX1 gene\u003c/strong\u003e obtained from four \u003cem\u003eAndrena\u003c/em\u003e species sampled across Jammu and Kashmir. The resulting phylogeny (Fig. 3) demonstrated that the sequences were distinctly separated and grouped accurately with their respective conspecific taxa, indicating clear genetic divergence among the species. To infer the evolutionary relationships, the \u003cstrong\u003eMaximum Likelihood (ML) method\u003c/strong\u003e was employed under the \u003cstrong\u003eGeneral Time Reversible (GTR) model\u003c/strong\u003e. The initial tree(s) required for the heuristic search were generated by applying the Neighbor-Joining algorithm to a matrix of pairwise genetic distances, which had been estimated using the \u003cstrong\u003eMaximum Composite Likelihood (MCL) method\u003c/strong\u003e. The robustness and reliability of the tree topology were assessed through a \u003cstrong\u003ebootstrap analysis with 1000 replicates\u003c/strong\u003e. Only those branches supported by more than 50% of bootstrap replicates were retained, while nodes with weaker support were collapsed. This ensured that the final bootstrap consensus tree reliably represented the evolutionary history of the analyzed taxa.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFloral resources of \u003cem\u003eAndrena\u003c/em\u003e species:\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAndrena\u003c/em\u003e species have been recorded from different floral resources of Kashmir. These floral resources have been categorized both family wise and crop wise as shown in Table 2. Distribution pattern of different species of Andrenid bees on different flora is shown in Table 2.\u003cem\u003e\u0026nbsp;Andrena flavipes\u0026nbsp;\u003c/em\u003ewas reported from Dandelion and Apple. However, \u003cem\u003eAndrena bicolour\u0026nbsp;\u003c/em\u003ewas reported from Poison hemlock, Dandelion, Creeping thistle, Apple and Pear. Similarly, \u003cem\u003eAndrena cineraria\u0026nbsp;\u003c/em\u003ewas found on Almond, Persian speedwell, Poison hemlock, Yellow star of Bethlehem and Pear and \u003cem\u003eAndrena pilipes\u003c/em\u003e was reported from Almond, Dandelion, Poison hemlock, Yellow star of Bethlehem, Creeping thistle and Pear.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2:\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;Distribution of different species of Andrenid bees on different flora:\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"765\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 7.44125%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eS.NO\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 18.5379%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eSpecies\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 74.0209%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eCrops\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 7.44125%;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 18.5379%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAndrena\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003epilipes\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 74.0209%;\"\u003e\n \u003cp\u003eAlmond (\u003cem\u003ePrunus amygdalus\u003c/em\u003e), Dandelion\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003e(Taraxacum officinale)\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003e,\u003c/em\u003e\u003c/strong\u003e Poison hemlock (\u003cem\u003eConium maculatum\u003c/em\u003e), Yellow star of Bethlehem (\u003cem\u003eGagea lutea\u003c/em\u003e), Creeping thistle (\u003cem\u003eCirsium arvense\u003c/em\u003e) and Pear (\u003cem\u003ePyrus\u003c/em\u003e).\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 7.44125%;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 18.5379%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAndrena\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003e\u0026nbsp;bicolour\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 74.0209%;\"\u003e\n \u003cp\u003ePoison hemlock (\u003cem\u003eConium maculatum)\u003c/em\u003e, Dandelion \u003cstrong\u003e(\u003cem\u003eTaraxacum officinale\u003c/em\u003e)\u003c/strong\u003e, Creeping thistle\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003cem\u003e(Cirsium arvense)\u003c/em\u003e, Apple (\u003cem\u003eMalus domestica\u003c/em\u003e) and Pear (\u003cem\u003ePyrus\u003c/em\u003e).\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 7.44125%;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 18.5379%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAndrena\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eflavipes\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 74.0209%;\"\u003e\n \u003cp\u003eDandelion \u003cstrong\u003e(Taraxacum officinale\u003c/strong\u003e) and Apple (\u003cem\u003eMalus domestica\u003c/em\u003e).\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 7.44125%;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 18.5379%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAndrena cineraria\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 74.0209%;\"\u003e\n \u003cp\u003eAlmond (\u003cem\u003ePrunus amygdalus\u003c/em\u003e), Persian speedwell (\u003cem\u003eVeronica persica\u003c/em\u003e), Poison hemlock \u003cem\u003e(Conium maculatum)\u003c/em\u003e,\u0026nbsp;Yellow star of Bethlehem (\u003cem\u003eGagea lutea\u003c/em\u003e) and Pear (\u003cem\u003ePyrus\u003c/em\u003e).\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eThe present study provides a comprehensive morphological and molecular characterization of four important \u003cem\u003eAndrena\u003c/em\u003e species: \u003cem\u003eAndrena bicolor\u003c/em\u003e, \u003cem\u003eA. cineraria\u003c/em\u003e, \u003cem\u003eA. pilipes\u003c/em\u003e, and \u003cem\u003eA. flavipes\u003c/em\u003e from the Kashmir Valley, India. These species represent significant pollinators contributing to both wild flora and cultivated crops in temperate ecosystems. The integration of classical morphological techniques with DNA barcoding has proven critical in accurately identifying species within the genus \u003cem\u003eAndrena\u003c/em\u003e, which is notoriously challenging due to morphological similarity and cryptic diversity.Our morphological analysis revealed characteristic diagnostic features that aligned well with existing taxonomic descriptions, such as body size, coloration patterns, antennal segmentation, and genitalia structure. For example, \u003cem\u003eA. bicolor\u003c/em\u003e was distinguishable by its larger body size and distinctive orange tibial scopa, while \u003cem\u003eA. cineraria\u003c/em\u003e exhibited the typical ashy brown color and specific antennal ratios that differentiated it from congeners (Wood et al., 2020). Importantly, the observed measurements, such as UICD to LICD ratios and flagellum lengths, provide a quantitative basis for comparison with global datasets (Ascher \u0026amp; Pickering, 2023).\u003c/p\u003e\n\u003cp\u003eMolecular identification using mitochondrial COI gene markers further corroborated the morphological diagnoses, overcoming limitations associated with subjective morphological assessment and potential misidentifications. The bidirectional sequencing and GenBank submission of COI sequences represent a valuable addition to the growing DNA barcode reference library for Indian solitary bees, as also highlighted by prior studies (Chandra et al., 2017; Moroń et al., 2019). The successful amplification and sequencing from both fresh and preserved specimens demonstrate the robustness of the molecular protocol employed (Mitchell, 2022; Ashfaq et al., 2023).\u003c/p\u003e\n\u003cp\u003eThe documented floral resources revealed a broad spectrum of host plants utilized by \u003cem\u003eAndrena\u003c/em\u003e species, including apple (\u003cem\u003eMalus domestica\u003c/em\u003e), pear (\u003cem\u003ePyrus spp.\u003c/em\u003e), poison hemlock (\u003cem\u003eConium maculatum\u003c/em\u003e), dandelion (\u003cem\u003eTaraxacum officinale\u003c/em\u003e), almond (\u003cem\u003ePrunus dulcis\u003c/em\u003e), and Persian speedwell (\u003cem\u003eVeronica persica\u003c/em\u003e). These observations reinforce the ecological versatility of \u003cem\u003eAndrena\u003c/em\u003e species and their vital role in supporting agricultural productivity and wild plant pollination in Kashmir’s agro ecosystems (Rader et al., 2020; Willmer et al., 2017). This is especially critical given the documented global decline of honey bees (Apis spp.) due to habitat loss, pesticide exposure, and emerging diseases (Goulson et al., 2015).\u003c/p\u003e\n\u003cp\u003eThe wide distribution of these \u003cem\u003eAndrena\u003c/em\u003e species across Kashmir Valley’s districts further emphasizes their adaptability to diverse environmental conditions, with elevations ranging from 1545 to 1776 meters above sea level. This finding aligns with the global distribution patterns of these species across temperate regions (Michener, 2007; Wood \u0026amp; Gupta, 2023).\u003c/p\u003e\n\u003cp\u003eHowever, challenges remain in the identification of Andrenid species, particularly when dealing with degraded specimens or closely related cryptic taxa. The success of DNA barcoding in this context highlights the need for further expansion of molecular reference databases, especially for underrepresented regions like India (Hebert et al., 2003; Kek et al., 2017).\u0026nbsp;\u003c/p\u003e\n\n"},{"header":"CONCLUSION","content":"\u003cp\u003eIn conclusion, the examination of \u003cem\u003eAndrena\u0026nbsp;\u003c/em\u003e\u003cem\u003epilipes\u003c/em\u003e,\u003cem\u003e\u0026nbsp;Andrena\u003c/em\u003e\u003cem\u003e\u0026nbsp;bicolour\u003c/em\u003e,\u003cem\u003e\u0026nbsp;Andrena\u0026nbsp;\u003c/em\u003e\u003cem\u003eflavipes\u003c/em\u003e and \u003cem\u003eAndrena cineraria\u003c/em\u003e has offered significant insights into their diverse morphological adaptations and structural characteristics. Through meticulous description and molecular validation using DNA analysis, distinct features have been identified that enhance the accuracy of species recognition and classification. This comprehensive study not only highlights the ecological and scientific importance of these bees but also enriches our understanding of the evolutionary diversity within the \u003cem\u003eAndrena\u003c/em\u003e genus.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAbrol DP (2012) Pollination biology: Biodiversity conservation and agricultural production, vol 100. Springer, pp 76\u0026ndash;90. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/978-94-007-1942-2\u003c/span\u003e\u003cspan address=\"10.1007/978-94-007-1942-2\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAscher JS, Pickering J (2023) Discover Life bee species guide and world checklist (Hymenoptera: Apoidea: Anthophila). \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.5281/zenodo.10019874\u003c/span\u003e\u003cspan address=\"10.5281/zenodo.10019874\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBogusch P, Blahova E, Horak J (2020) Pollen specialists are more endangered than non-specialised bees even though they collect pollen on flowers of non-endangered plants. Arthropod-Plant Interact 14:759\u0026ndash;769. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1007/s11829-020-09789-y\u003c/span\u003e\u003cspan address=\"10.1007/s11829-020-09789-y\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBallantyne G, Baldock KCR, Rendell L, Willmer PG (2017) Pollinator importance networks: Are non-bees significant? Ecol Entomol 42(5):1\u0026ndash;11. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1038/s41598-017-08798\u003c/span\u003e\u003cspan address=\"10.1038/s41598-017-08798\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eChandra K, Wood TJ, Patiny S, Dutta A, Laha S, Smith B, Basu (2017) DNA barcoding of \u003cem\u003eAndrena agilissima\u003c/em\u003e in India. J Entomol Res 41:123\u0026ndash;130. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.5852/ejt.2024.948.2637\u003c/span\u003e\u003cspan address=\"10.5852/ejt.2024.948.2637\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eChen X, Agapow PM, Bininda-Emonds OR, Crandall KA, Gittleman JL, Mace GM, Marshall JC, Purvis A (2022) DNA barcoding reveals hidden diversity of bees in East Asia. Sci Rep 12:54\u0026ndash;82\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDelaplane KS, Mayer DF (2000) Crop pollination by bees. CABI Publishing\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eGoulson D, Nicholls E, Bot\u0026iacute;as C, Rotheray EL (2015) Bee declines driven by combined stress from parasites, pesticides, and lack of flowers. Science 347:125\u0026ndash;595\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHebert PD, Cywinska A, Ball SL, DeWaard JR (2003) Biological identifications through DNA barcodes. \u003cem\u003eProceedings of the Royal Society B: Biological Sciences\u003c/em\u003e, 270:313\u0026ndash;321\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKek SP, Bhat S, Paschapur AU, Subbanna ARNS, Stanley J, Gupta J, Mishra KK, Kumar (2017) Application of COI barcoding for bee species identification. Apidologie 48(3):377\u0026ndash;387\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKlein AM (2007) Importance of pollinators in changing landscapes for world crops. \u003cem\u003eProceedings of the Royal Society B: Biological Sciences, 274\u003c/em\u003e:303\u0026ndash;313\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLiu C, Ashfaq M, Yin Y, Zhu Y, Wang Z, Cheng H, Hebert P (2023) Using DNA metabarcoding to assess insect diversity in citrus orchards. Peer J Entomol 11:153\u0026ndash;338\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMichener CD (2007) The bees of the world. Johns Hopkins University\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMitchell A (2022) DNA barcoding with historical specimens: Unlocking museum collections. Mol Ecol Resour 22(7):2565\u0026ndash;2578\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMoroń D, Le\u0026oacute;n-Cort\u0026eacute;s JL, Bried JT, Roy DB (2019) Effectiveness of DNA barcoding for identifying wild bees. J Insect Conserv 23(5\u0026ndash;6):829\u0026ndash;840\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eOllerton J (2021) Pollinators \u0026amp; pollination: Nature and society. Pelagic Publishing\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRader R, Cunningham SA, Howlett BG, Inouye DW (2020) Non-bee insects as pollinators: Global patterns and implications. \u003cem\u003eProceedings of the National Academy of Sciences\u003c/em\u003e, 117: 144\u0026ndash;151\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWood TJ, Bogusch P, Bl\u0026aacute;hov\u0026aacute; E, Hor\u0026aacute;k J (2020) Diversity and ecology of \u003cem\u003eAndrena\u003c/em\u003e bees: A global review. Insect Conserv Divers 13(6):507\u0026ndash;523. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.5852/ejt.2022.843.1947\u003c/span\u003e\u003cspan address=\"10.5852/ejt.2022.843.1947\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWood TJ, Gupta RK (2023) Revision of Indian \u003cem\u003eAndrena\u003c/em\u003e bees. Eur J Taxonomy 913:1\u0026ndash;58\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Andrena, morpho-molecular characterization, DNA barcoding, pollinator diversity, Himalayan ecosystem","lastPublishedDoi":"10.21203/rs.3.rs-7694933/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7694933/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eFour species of the genus \u003cem\u003eAndrena\u003c/em\u003e: \u003cem\u003eAndrena (Euandrena) bicolor\u003c/em\u003e, \u003cem\u003eAndrena (Melandrena) cineraria\u003c/em\u003e, \u003cem\u003eAndrena (Plastandrena) pilipes\u003c/em\u003e, and \u003cem\u003eAndrena (Zonandrena) flavipes\u003c/em\u003e were reported for the first time from Jammu \u0026amp; Kashmir, India. Specimens were collected from Srinagar, Pulwama, and Budgam districts and analyzed using 23 morphological characters. In addition, 658 bp COI DNA barcodes were generated and deposited in GenBank, providing reliable molecular markers for species identification. Integration of morphological and molecular datasets revealed distinct diagnostic traits and intraspecific variations, indicating evolutionary divergence likely shaped by geographic isolation and ecological pressures in the Himalayan region. This study emphasizes the role of integrative taxonomy in accurate species delimitation and contributes to the understanding of pollinator diversity and conservation in the fragile Himalayan ecosystem.\u003c/p\u003e","manuscriptTitle":"From Mountains to Molecules: Decoding the Hidden Diversity of Andrena Bees in the Himalayan Wilderness","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-25 06:32:25","doi":"10.21203/rs.3.rs-7694933/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"58a7875d-77ee-4aa0-861c-9c973234d7a9","owner":[],"postedDate":"September 25th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":55204617,"name":"Entomology"}],"tags":[],"updatedAt":"2025-09-25T06:32:25+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-25 06:32:25","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7694933","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7694933","identity":"rs-7694933","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}