Dna barcoding, mineral and phytochemical analysis of cape gooseberry (physalis spp.) accessions in Kenya

Abstract

Physalis genus belongs to the solanaceae family and is mostly a wild self-propagating orphan fruit plant. Physalis fruit contains nutritional and phytochemical compounds of importance to public health and is a potential ingredient for fortification of foods and beverages. In kenya, commercial cultivation of physalis has significantly increased for the last decade and the farmers are concerned about the reliability and identity of the planting material. Challenges facing the production of planting material include difficulty in precise identification of the physalis species due to lack of discriminatory morphological features. Dna barcoding is an effective tool in identification and discrimination of plant species. Therefore, the objectives of this study were to identify and discriminate the different physalis species in the wild and cultivated in kenya using dna barcoding as well as determine the nutritional and biochemical profile of selected physalis accessions. To identify the physalis species from selected counties in kenya, ribulose-1,5-bisphosphate carboxylase large (rbcl) gene and internal transcribed spacer 2 (its2) dna barcodes were used. Genomic dna was extracted from leaves and polymerase chain reaction (pcr) amplification was carried out using its2 and rbcl barcodes followed by sanger sequencing of the amplicons. Sequences were curated and aligned along their reference sequences based on multiple sequence comparison by log-expectation (muscle). The generated multiple sequence alignments were used to prepare phylogenetic trees for physalis species discrimination based on rbcl and its2 genes by bayesian inference based on mrbayes software. Genetic diversity, distance and polymorphism were also assessed for the physalis species through the use of dna sequence polymorphism (dnasp) software. Additionally, assessment of physalis species mineral content for nine representative accessions was performed on an atomic absorption spectrophotometer and phytochemical content determined by uv-visible spectrophotometry. Further, the radical scavenging activity of physalis accessions was determined for 2,2-diphenyl-2-picrylhydrazyl (dpph) and hydrogen peroxide radicals. Success rate of pcr amplification and sequencing of rbcl and its2 genes ranged from 82% to 88% and 65% to 67%, respectively. The phylogenetic tree generated for the rbcl sequences was unable to distinguish between physalis species. However, the phylogenetic tree generated for the its2 sequences successfully identified three species of physalis: physalis peruviana, physalis purpurea and physalis cordata. There was low nucleotide diversity and genetic distance of 0.01333 and 0.04, respectively, for the rbcl genes while high genetic diversity and distance was noted for the its2 sequences. The highest genetic diversity and distance between physalis species was noted between p. Peruviana and p. Cordata at 0.36923 and 0.703, respectively based on its2 sequences. The highest genetic nucleotide diversity and distance within species was noted for the p. Peruviana at 0.26324 and 0.46, respectively, for the its2 gene sequences. Tajima d values obtained indicated low frequency mutations among the physalis sequences based on rbcl and its2 barcodes. The mineral content of the physalis accessions was significantly different (p < 0.05) for calcium, copper, zinc, nickel and lithium between accessions. For the phytochemicals of physalis fruit accessions, tannic acid content was significantly different (p < 0.05), while the phenolic acid and flavonoid contents were not significantly different (p > 0.05). Physalis accessions dpph radical scavenging activity was significantly different (p < 0.05) while no significant difference (p > 0.05) was noted for hydrogen peroxide scavenging capacity. Therefore, this study identified kenyan physalis accessions based on its2 barcode region as p. Purpurea, p. Peruviana and p. Cordata. The findings also demonstrated that physalis accessions in kenya are rich in mineral and phytochemical contents as well as high antioxidant properties.