P1-33BEL-000177 – Final Report FINAL REPORT NAMIBIA BBI SSFA April 2020 Development of Institutional Synergy to Enable Inventorying and Cultivation of Priority Species for Conservation in the Southern Africa Region: Medicinal Plants in the Apocynaceae Family The overall objective of this Project is to foster long-term cooperation between key and relevant Institutions in Southern Africa on addressing the unsustainable exploitation of medicinal plants in the Region. The initial focus is a group of plants in the Apocynaceae family that recently became popular in northern Namibia. The project endeavours to i) Conduct traditional and molecular-based taxonomic work to clarify the identity and phylogeny of the targeted species, ii) Contribute to an existing public DNA sequence database (e.g. NCBI), iii) Establish propagation initiatives to reduce pressure on the wild resource, and iv) Develop policy to accord the targeted species legal protection in Namibia and other range states. i) and ii) constitute the immediate Project objectives to which this CBD Bio-Bridge Initiative Small-Scale Funding contributes, through six key activities. All six activities were successfully implemented over the period March 2019 – March 2020, as detailed in the following report: 1. Outline in detail the activities implemented during the Project. Activity 1: Collection trip to northern Namibia – Herbarium voucher specimens of the targeted plants and leaf samples for molecular analysis will be collected from various sites in northern Namibia. Feedback: Herbarium plant specimens, leaf samples as well as soil samples were collected from 85 sites across nine Namibian regions in March – April 2019 (see Collection sites 2019 in “Report_BBI Namibia”). However, 2019 suffered a severe drought that affected specimen collection, especially in the arid northwest. Consequently, fieldwork was shortened by two weeks as the area west of Sesfontein/Opuwo was deemed too dry to be worth visiting. This year, 2020, has received good rains, presenting an opportunity to visit the area that was left out. Activity 2: Visit to Herbarium of Lubango (LUBA) – Namibian partners will visit LUBA to familiarise themselves with the facility and its staff, identify how it can enhance the current Project, discuss the logistics of the upcoming trips to the African Centre for DNA Barcoding (Activity 3) and laboratory training workshop in Windhoek (Activity 4), and determine the feasibility for a joint field collection in Angola in a subsequent phase of the project. Feedback: Three Namibian botanists visited LUBA and the host Institution ISCED for two days in September 2019. They familiarised themselves with the dhingila specimens held in the Herbarium and other taxa of interest, networked with staff, met the Director of ISCED, Directors of ISPT, and undertook a botanical excursion around Lubango. They were informed that LUBA was in the process of being transferred to the management of the ISPT. The Herbarium recommended a student from ISPT 1 P1-33BEL-000177 – Final Report and one from the University of Agostinho Neto in Luanda to attend the laboratory training workshop in Windhoek. The networking opportunity that the trip provided will go a long way in supporting continuation of the dhingila work and in contributing to other prospective projects of common interest. Angola is one of Namibia’s major biodiversity stakeholders; the two countries currently share two Trans-frontier Conservation Areas, SIONA and KAZA. News article: http://www.pcu.uct.ac.za/botanists-visit-lubango-angola-092019 Activity 3: Visit to African Centre for DNA Barcoding (ACDB) – Namibian and Angolan partners will visit the African Centre for DNA Barcoding in Johannesburg, South Africa, to familiarise themselves with the facility, identify how the sequencing services required from the Centre (Activity 5) can be accelerated, due to time constraints, and to strengthen linkages to support potential biodiversity projects in the visiting countries in accessing the facility in future. Feedback: Four Namibian botanists and two Angolan students undertook a one day visit to the ACDB in December 2019. The delegates were treated to presentations on various projects of the ACDB, a demonstration of the LifeScanner, introduction to the Lab-in-a-box, and a tour of the ACDB laboratory facility. Since Activity 4 preceded the visit the delegates took the samples to ACDB for sequencing. The visit provided insight into the Centre and a platform for communicating services needed or potential opportunities for collaboration. Activity 4: Molecular analysis laboratory training workshop (extraction and amplification) – A training workshop will be held at the University of Namibia to train Namibian and Angolan participants in laboratory methods for DNA extraction and amplification, using leaf samples collected in Activity 1. Emphasis will be placed on DNA barcoding, but other molecular analysis techniques will also be considered if deemed feasible. Feedback: A total of eight Namibian and two Angolan participants took part in the laboratory training workshop, which was held towards the end of 2019. They extracted and amplified DNA barcodes from 102 leaf samples of mostly dhingila species collected in the field, but some other Apocynaceae as well as other species of interest field-collected or taken from the National Herbarium or Botanic Garden were also included. The main training session took place from 25 November to 4 December 2019 and was blessed with opening remarks by the Head of the Biological Sciences Department, Prof. John Mufune. The main training was preceded by a trial run which processed the first 50 samples and tested three plant barcode markers, recommending matK for the main training session. News article: http://www.pcu.uct.ac.za/barcoding-dhingila-namibia Activity 5: Sequencing – Samples from Activity 4 will be sent to the African Centre for DNA Barcoding (ACDB) in South Africa for sequencing. Feedback: 48 samples were taken to the ACDB on 6 December 2019 for sequencing. Results were received on 6 March 2020. 2 P1-33BEL-000177 – Final Report Activity 6: Informatics – This activity will entail analysis of the molecular data matrix and possibly morphometric analysis. Feedback: Towards the end of the training workshop in December participants were introduced to sequence editing and construction of phylogenetic trees using sequences of the Namib Desert endemic Moringa ovalifolia. Upon receiving dhingila sequences from the ACDB, 10 of them were selected on the basis of geographic representativeness, and were then edited and submitted to the Barcode of Life Database (BOLD). The document “From Field to BOLD” in the folder “Report_BBI Namibia” outlines the workflow. Comparison of collected herbarium specimens with material in the National Herbarium of Namibia and blasting of chloroplast MatK DNA barcode sequences obtained from the training workshop against the Genbank (NCBI) sequence database have identified the targeted dhingila species as Marsdenia macrantha (Clotzsch) Schltr. (Apocynaceae) and Gymnema sylvestre (Retz.) R.Br. ex Schult (Marsdenia sylvestris (Retz.) P.I. Forst.) (Apocynaceae). Both species are used inseparably as “they taste the same, smell the same, and are both effective”, according to the words of a local dhingila trader who identified them as the “female dhingila” (M. macrantha) – based on the broader-shaped fruit and as “male dhingila” (G. sylvestre) – which has a slender fruit. It also appears that a vine Fockea species (Apocynaceae), a wild jasmine (Oleaceae), and possibly also the alien invasive rubber vine (Apocynaceae) are sometimes confused with the Marsdenia and Gymnema species, especially in the absence of fruit and flowers. While confamilials may present similar phytochemicals and curative properties there is clearly a need for awareness work to ensure that the local people can distinguish the “real dhingila” species. The literature indicates that M. macrantha naturally occurs in a number of African countries, while G. sylvestre is more widespread in tropical to subtropical Africa and beyond, with natural distribution extending as far east as Arabia, India, and China (Bruyns 2014). The leaves of G. sylvestre, marketed as Indian Ayurveda herbal tea, are used around the world mainly for slimming and to treat diabetes (see e.g. ScienceDirect and this site). While in Namibia the main focus is on the root, some of the ailments targeted are similar (see the document “Dhingila Products” in “Report_BBI Namibia”). There is emerging interest in Namibia to protect dhingila as an indigenous biological resource with potential health and commercial value as well as to ensure benefit-sharing from commercialisation of the traditional knowledge (TK) and any intellectual property associated with the plant, in line with the Nagoya Protocol and the Namibian Access to Biological and Genetic Resources and Associated Traditional Knowledge Act. However, claims of ownership of the G. sylvestre resource and TK could compete with those that could emerge from other range states such as India, where there is a documented long history of traditional use and an established trade. Evidence that there is variation significant enough to warrant delimitation of the Namibian and other regions’ G. sylvestre as separate taxa at any taxonomic level will help avert potential conflict. A degree of evolutionary divergence across such a wide geographic and climatic range is expected. Northwestern Namibia could possibly represent one of the driest,