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Evaluation of rapid molecular diagnostics for differentiating medicinal Kaempferia species from its adulterants.

ABSTRACT: Accurate detection of unique herbs is crucial for herbal medicine preparation. Zingiberaceae species, which are important in Ayurvedic medicine of India, are often misidentified in Northeast (NE) Indian herbal markets. Kaempferia galanga (Zingiberaceae) is one of the major components of popular Ayurvedic drugs used for rheumatic diseases (i.e., "Gandha Thailam" and "Rasnairandadi Kashayam"), contusions, fractures, and sprains. In NE India, herbal healers often misidentify plants from the Marantaceae family (e.g., Calathea bachemiana and Maranta leuconeura) as Kaempferia, which leads to adulteration of the medicinal herb. This misidentification of herbs occurs in NE India because Zingiberaceae plant barcoding information is inadequate. As a consequence, herbal medicine is not only therapeutically less effective but may also cause adverse reactions that range from mild to life-threatening. In this study, we used eight barcoding loci to develop "fingerprints" for four Kaempferia species and two species frequently mistaken for Kaempferia. The PCR and sequencing success of the loci matK, rbcL and trnH-psbA were found to be 100%; the combination of matK, rbcL, and trnH-psbA proved to be the ideal locus for discriminating the Kaempferia species from their adulterants because the combined loci showed greater variability than individual loci. This reliable tool was therefore developed in the current study for accurate identification of Kaempferia plants which can effectively resolve identification issues for herbal healers.

SUBMITTER: Basak S

PROVIDER: S-EPMC6704042 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Evaluation of rapid molecular diagnostics for differentiating medicinal <i>Kaempferia</i> species from its adulterants.

Basak Supriyo S Aadi Moolam Ramesh R Parida Ajay A Mitra Sudip S Rangan Latha L

Plant diversity 20190416 3

Accurate detection of unique herbs is crucial for herbal medicine preparation. Zingiberaceae species, which are important in Ayurvedic medicine of India, are often misidentified in Northeast (NE) Indian herbal markets. <i>Kaempferia galanga</i> (Zingiberaceae) is one of the major components of popular Ayurvedic drugs used for rheumatic diseases (i.e., "Gandha Thailam" and "Rasnairandadi Kashayam"), contusions, fractures, and sprains. In NE India, herbal healers often misidentify plants from the ...[more]

PMID: 31453420

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Project description:Smithia conferta Sm. is an annual herb widely used in Indian traditional medical practice and commonly known as "Lakshman booti" in Sanskrit. Morphological resemblance among the species of genus Smithia Aiton. leads to inaccurate identification and adulteration. This causes inconsistent therapeutic effects and also affects the quality of herbal medicine.This study aimed to generate potential barcode for authentication of S. conferta and its adulterants through DNA barcoding technique.Genomic DNA extracted from S. conferta and its adulterants was used as templates for polymerase chain reaction amplification of the barcoding regions. The amplicons were directed for sequencing, and species identification was conducted using BLASTn and unweighted pair-group method with arithmetic mean trees. In addition, the secondary structures of internal transcribed spacer (ITS) 2 region were predicted.The nucleotide sequence of ITS provides species-specific single nucleotide polymorphisms and sequence divergence (22%) than psbA-trnH (10.9%) and rbcL (3.1%) sequences. The ITS barcode indicates that S. conferta and Smithia sensitiva are closely related compared to other species.ITS is the most applicable barcode for molecular authentication of S. conferta, and further chloroplast barcodes should be tested for phylogenetic analysis of genus Smithia.The present investigation is the first effort of utilization of DNA barcode for molecular authentication of S. conferta and its adulterants. Also, this study expanded the application of the ITS2 sequence data in the authentication. The ITS has been proved as a potential and reliable candidate barcode for the authentication of S. conferta. Abbreviations used: BLASTn: Basic Local Alignment Search Tool for Nucleotide; MEGA: Molecular Evolutionary Genetic Analysis; EMBL: European Molecular Biology Laboratory; psbA-trnH: Photosystem II protein D1- stuctural RNA: His tRNA gene; rbcL: Ribulose 1,5 bi-phosphate carboxylase/oxygenase large subunit gene.

Project description:Kaempferia galanga and Kaempferia elegans, which belong to the genus Kaempferia family Zingiberaceae, are used as valuable herbal medicine and ornamental plants, respectively. The chloroplast genomes have been used for molecular markers, species identification and phylogenetic studies. In this study, the complete chloroplast genome sequences of K. galanga and K. elegans are reported. Results show that the complete chloroplast genome of K. galanga is 163,811 bp long, having a quadripartite structure with large single copy (LSC) of 88,405 bp and a small single copy (SSC) of 15,812 bp separated by inverted repeats (IRs) of 29,797 bp. Similarly, the complete chloroplast genome of K. elegans is 163,555 bp long, having a quadripartite structure in which IRs of 29,773 bp length separates 88,020 bp of LSC and 15,989 bp of SSC. A total of 111 genes in K. galanga and 113 genes in K. elegans comprised 79 protein-coding genes and 4 ribosomal RNA (rRNA) genes, as well as 28 and 30 transfer RNA (tRNA) genes in K. galanga and K. elegans, respectively. The gene order, GC content and orientation of the two Kaempferia chloroplast genomes exhibited high similarity. The location and distribution of simple sequence repeats (SSRs) and long repeat sequences were determined. Eight highly variable regions between the two Kaempferia species were identified and 643 mutation events, including 536 single-nucleotide polymorphisms (SNPs) and 107 insertion/deletions (indels), were accurately located. Sequence divergences of the whole chloroplast genomes were calculated among related Zingiberaceae species. The phylogenetic analysis based on SNPs among eleven species strongly supported that K. galanga and K. elegans formed a cluster within Zingiberaceae. This study identified the unique characteristics of the entire K. galanga and K. elegans chloroplast genomes that contribute to our understanding of the chloroplast DNA evolution within Zingiberaceae species. It provides valuable information for phylogenetic analysis and species identification within genus Kaempferia.

Project description:BackgroundScutellaria baicalensis Georgi has been extensively used as a medicinal herb in China for over 2000 years. They may be intentionally or inadvertently substituted or blended with comparable species in the local market, threatening clinical medication safety. Molecular markers are effective tools to prevent misidentification and eliminate doping and falsification among Scutellaria plants. This study screened four highly variable regions to identify Scutellaria and its adulterants. In addition, a phylogenetic analysis was performed using the complete cp genome combined with published Scutellaria species samples. Moreover, a comparative analysis of the cp genomes was conducted to investigate the cp genome evolution of S. baicalensis.ResultsThe complete cp genome of five species of Scutellaria was sequenced for the first time, and four previously published Scutellaria species were re-sequenced. They all exhibited a conserved quadripartite structure in their cp genomes, including two distinct regions, namely a small and large single copy region, respectively, and two inverted repeats encompassing the majority of ribosomal RNA genes. Furthermore, the nine species exhibited high conservation from aspects of the genome structure, codon usage, repeat sequences, and gene content. Four highly variable regions (matK-rps16, ndhC-trnV-UAC, psbE-petL, and rps16-trnQ-UUG) may function as potential molecular markers for differentiating S. baicalensis from its adulterants. Additionally, the monophyly of Scutellaria was ascertained and could be reclassified into two subgenera, subgenus Anaspis and subgenus Scutellaria, as evidenced by the phylogenetic analyses on sequences of cp genome and shared protein-coding sequences. According to the molecular clock analysis, it has been inferred that the divergence of Scutellaria occurred at approximately 4.0 Mya during the Pliocene Epoch.ConclusionOur study provides an invaluable theoretical basis for further Scutellaria species identification, phylogenetics, and evolution analysis.

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Evaluation of rapid molecular diagnostics for differentiating medicinal Kaempferia species from its adulterants.

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Evaluation of rapid molecular diagnostics for differentiating medicinal <i>Kaempferia</i> species from its adulterants.

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