Dedicated to inspiration

Andreas de Boer 21-09-1975 – 14-02-1998

Frederik Swart

13-03-1978 – 18-06-1999

Hans de Boer 20-02-1948 – 21-11-2009

Cover image: Courtesy of Ken Ishikawa, Creative Commons BY-SA 2.0

List of Papers

This dissertation is based on the following papers, which are referred to in the text by their Roman numerals.

I de Boer, H.J., Lamxay, V. (2009). Plants used during pregnancy, childbirth and postpartum healthcare in Lao PDR: A comparative study of the Brou, Saek and Kry ethnic groups. Journal of Ethnobi-ology and Ethnomedicine, 5: 25.

II Lamxay, V., de Boer, H.J., Björk, L. (2011). Traditions and plant use during pregnancy, childbirth and postpartum recovery by the Kry ethnic group in Lao PDR. Journal of Ethnobiology and Ethno-medicine, 7: 14.

III de Boer, H.J., Lamxay, V., Björk, L. (2011). Steam sauna and mother roasting in Lao PDR: Practices and Chemical constituents of essential oils of plant species used in postpartum recovery. BMC Complementary and Alternative Medicine, 11: 128.

IV de Boer, H.J., Lamxay, V., Björk, L. Transmission of Traditional Ecological Knowledge: Similarity of medicinal plant knowledge among the Brou, Saek and Kry in Lao PDR. Journal of Ethnophar-macology, [Submitted].

V de Boer, H.J., Vongsombath, C., Pålsson, K., Björk, L., Jaenson, G.T. (2010). Traditional repellents against haematophagous inverte-brates: A comparative study of plants used in 66 villages in Lao PDR. Journal of Medical Entomology 47: 400-414.

VI de Boer, H.J., Vongsombath, C., Käfer, J. (2011). A fly in the oint-ment: Evaluation of traditional use of plants to repel and kill blowfly larvae in fermented fish. PLoS ONE, 6(12): e29521.

VII de Boer, H.J., Schaefer, H., Thulin, M., Renner, S. Evolution and loss of long-fringed petals: A case study using a dated phylogeny of the snake gourds, Trichosanthes (Cucurbitaceae). BMC Evolutionary Biology, [Submitted]

VIII de Boer, H.J., Thulin, M. Synopsis of Trichosanthes (Cucurbita-ceae) based on recent molecular phylogenetic data. PhytoKeys, [Submitted]

IX Cooper, W.E., de Boer, H.J. (2011) A taxonomic revision of Tri-chosanthes L. (Cucurbitaceae) in Australia, including one new spe-cies from the Northern Territory. Austrobaileya, 8: 364–386.

Papers V and IX are reproduced with permission from the publishers.

Papers published during my PhD research not included in this dissertation. I Mati, E., de Boer, H.J. (2011) Trade and commercialization of herbal

medicine in the Qaysari Market, Kurdish Autonomous Region, Iraq. Journal of Ethnopharmacology, 133: 490-510

II Vongsombath, C., de Boer, H.J., Pålsson, K. (2011) Keeping leeches at bay: Field evaluation of plant-derived extracts against terrestrial blood-sucking leeches (Haemadipsidae) in Lao PDR. Acta Tropica, 119: 178-82

III Mati, E., de Boer, H.J. (2010) An assessment of contemporary knowledge on dyeing plant species and natural dye use in Kurdish Au-tonomous Region, Iraq. Economic Botany, 64: 149-160

IV de Boer, H.J. (2010) Local Awareness of Scarcity and Endangerment of Medicinal Plants in Roussenski Lom Natural Park in Northern Bul-garia. In: Pardo-de-Santayana, M., Puri, R.K. (Editors). Ethnobiology in the New Europe

V de Boer, H.J. (2009) African plants as antipathogen agents: Clinical and experimental evidence. In: Watson, R., Preedy, V. The Encyclope-dia of Herbal Medicine in Clinical Practice. CABI publishing, Walling-ford.

VI de Boer, H.J., Hagemann, U., Ericsson, J., Meyboom, R. (2007) Aller-gic reactions to Pelargonium-derived medicines. Drug Safety, 30: 677-680

VII Farah, M., Olsson, S., Bate, J., Lindquist, M., Edwards, R., Simmonds, M.S.J., Leon, C., de Boer, H.J., Thulin, M. (2006) Botanical Nomen-clature in Pharmacovigilance and a Recommendation for Standardisa-tion. Drug Safety, 29: 1023-1029

VIII de Boer, H.J., Thulin, M. (2006) Lectotypification of Callicocca ipe-cacuanha Brot. and neotypification of Cephaelis acuminata H.Karst., with reference to the drug Ipecac. Taxon, 54: 1080-1082

Contents

Introduction .................................................................................................. 11Lao PDR .................................................................................................. 11Ethnobotany ............................................................................................. 12Traditional medicine and women’s health ............................................... 13Traditional repellents ............................................................................... 15Phylogeny, dating and biogeography of Trichosanthes .......................... 15Aims ......................................................................................................... 18

Summary of Papers ...................................................................................... 20Paper I ...................................................................................................... 20Paper II .................................................................................................... 22Paper III ................................................................................................... 23Paper IV ................................................................................................... 25Paper V .................................................................................................... 26Paper VI ................................................................................................... 28Paper VII .................................................................................................. 30Paper VIII ................................................................................................ 33Paper IX ................................................................................................... 33

Conclusions .................................................................................................. 36

Research ethics ............................................................................................. 39

Svensk sammanfattning ................................................................................ 42

Acknowledgments ........................................................................................ 45

References .................................................................................................... 51

Figures

Figure 1. Sunda and Sahul, indicating the Wallace, Weber and Lydekker Lines. Wallacea is the area in between the Wallace and Lydekker Line. Courtesy of Maximilian Dörrbecker, Wikipedia, Creative Commons license. ................................................................................................. 17

Abbreviations

ABS Access and Benefit Sharing BEAST Bayesian Evolutionary Analysis Sampling Trees CBD Convention on Biological Diversity CITES Convention on International Trade in Endangered

Species of Wild Fauna and Flora DNA Deoxyribonucleic acid GBIF Global Biodiversity Information Facility GC-MS Gas Chromatography - Mass Spectrometry GTI Global Taxonomy Initiative Lao PDR Lao People’s Democratic Republic ML Maximum Likelihood MTA Material Transfer Agreement Mya Million years before present NBCA National Biodiversity Conservation Areas NPA National Protected Area NTFP Non Timber Forest Product NUoL National University of Laos, Lao PDR PCR Polymerase chain reaction RASP Reconstruct Ancestral State in Phylogenies RAxML Randomized Axelerated Maximum Likelihood S-DIVA Statistical Dispersal-Vicariance Analysis SIDA Swedish International Development Cooperation

Agency UU Uppsala University, Sweden

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Introduction

Lao PDR Lao PDR, land-locked in Southeast Asia, is bordered by China in the North, Myanmar in the Northwest, Thailand in the West, Cambodia in the South and Vietnam in the East. It is located in the lower Mekong Basin and covers an area of 236,800 km2. Four-fifths of the country is mountainous while the remaining one-fifth comprises mostly flood plains along the Mekong River. It has a tropical monsoon climate with two distinct seasons, the rainy season from the beginning of May to October and the dry season from November to April. Temperatures are highest in April and May (ranging from 30 – 39°C) and lowest in December and January (down to below 15°C). In the north and some mountainous areas elsewhere, mean temperatures can be as low as 10°C. Monsoon rains cause disturbance and flooding, and facilitate disease vector reproduction. Average annual rainfall ranges from 1300 to 3700 mm per year (Wikipedia 2012).

Lao PDR is a country of rich biological and cultural diversity. Lao PDR has one of the most pristine floras and faunas in South-East Asia, but major challenges facing nature in Laos are the internal pressures of economic growth, foreign investment projects, hydroelectric projects and logging. In 1993 the Lao government established 20 national biodiversity conservation areas (NBCAs). Lao economy is highly dependent on forest products due to the country’s low population density combined with abundant forest cover (Foppes & Ketphanh 2000). Collection of non-timber forest products (NTFPs) plays an important role in the livelihood systems of rural house-holds, typically accounting for 40–60 % of annual household income, and up to 80 % in areas of severe poverty (Douangsavanh et al. 2002). NTFPs are the safety net of many rural families, providing, through direct household consumption, a means of livelihood at times when everything else has failed. Through the sale of NTFPs the people obtain cash to buy rice and other ne-cessities (Douangsavanh et al. 2002).

The country’s landlocked location, mountainous terrain and inaccessibil-ity have benefited ethnic diversity. Various classifications of the ethnic groups exist. Classifications by ethnographers range from 42 to 132 groups (Spire 1907; Halpern 1961). The Lao official classifications range from a tripartite system, lowland, middle and highland, used up to the 1980s (Schliesinger 2003) to the current one of 49 groups (Lao National Front for

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Construction 2005). The population and linguistic pattern is the result of vast movements in the last 2000 years, mainly from China to the southern low-lands (Enfield 2005). Cultural and linguistic diversity in Lao PDR is reflect-ed in genetic data, and recent population genetic studies using mtDNA re-veal a highly complex haplotype structure (Bodner et al. 2011).

Ethnobotany Ethnobotany is the study of the traditional use of plants. Traditional use of plants is very broadly defined, and usually ethnobotany applies to the study of plants for food, construction, healthcare, instruments, fodder, fibers and dyes. The study of useful plants at Uppsala University dates back to the re-search of Linnaeus, who during his many travels throughout Scandinavia meticulously noted down his observations regarding their economic value. Ethnobiology studies the traditional use of plants and/or animals, and eth-noecology refers to the interactions between people and their environment. Linnaeus’ qualitative approach to data collection has typified this field of research, and many ethnobotanical studies focus on the documentation of species’ uses by different ethnic groups living in a more or less traditional way. The main merit of these qualitative studies is to document traditional plant use for future generations, as traditionally living people become amal-gamated into mainstream culture.

Traditional plant use can also be approached using quantitative techniques to measure cultural importance, consensus and similarity of plant use. Dif-ferent methods and indices have been used, and most rely on informant-use-reports, i.e. species s used in use-category u by informant i, where informant can be broadened to represent village, cluster or ethnic group if data is gath-ered through group or village interviews. Indices can be computed per spe-cies, use, use-category or informant, depending on the focus of the research (for reviews see Hoffman & Gallaher (2007) and Tardío & Pardo-de-Santayana (2008)). Indices incorporating additional data such as species recognition, use value, agreement, ecological knowledge and total knowledge can be used to test a variety of hypotheses beyond what inform-ant-use-reports allow (Reyes-García et al. 2003; Reyes-García, et al. 2006; McDade et al. 2007).

Species importance indices based on informant-use-reports enable us to quantify the relative importance of specific plant taxa to different human groups, and a variety of them has been proposed by different ethnobiolo-gists: Relative Frequency of Citation (RFC) (Trotter & Logan 1986), Rela-tive Importance Index (RI) (Pardo-de-Santayana 2003), Cultural Value Index (CV) (Reyes-García, et al. 2006), Cultural Importance Index (CI) (Tardío & Pardo-de-Santayana 2008), and Use-Value Index (UV) (Phillips & Gentry 1993).

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Instead of looking at informant agreement per species, various models have also been proposed that analyze informant agreement per use (e.g. use of plants to promote postpartum perineal healing, or plant-use combinations with putative uterotonic properties). Trotter & Logan (1986) in their pioneer paper on informant agreement, propose the informant agreement ratio (IAR). Heinrich et al. (1998) defined a distinct adaptation of the IAR, the informant consensus factor (FIC). As both the FIC and the IAR are usually calculated per ethnic group, Moerman (2007) proposed a cultural agreement ratio (CAR), a rethinking of the FIC at a meta-cultural level. The FIC and the IAR show con-sensus within culture, the CAR shows consensus across cultures, but a sub-sequent comparison of consensus between cultures requires a qualitative or statistical approach.

Traditional medicine and women’s health Medicinal plants have a significant role during pregnancy, birth and postpar-tum care in many rural areas of the world. Plants used in women’s health related conditions such as female fertility, amenorrhea, menorrhagia, birth control, pregnancy, birth (parturition), postpartum (puerperium) and lacta-tion, including infant care, have been documented for various ethnic groups (Singh et al. 1984; Bourdy & Walter 1992; Liulan et al. 2003; Jain et al. 2004; Ticktin & Dalle 2005; Zumsteg & Weckerle 2007). In Western tradi-tional medicine the label “old wives’ tales” has been applied to all knowledge of interest to women – fertility, birth, childcare – transmitted orally from one generation of women to the next, and the derogatory label reflects male devaluation and relegation to folklore of this exclusively fe-male realm of knowledge (Newman 1985). Research focusing on the use of these plants often focuses on the realm of knowledge of male traditional healers, and scholars have missed the wealth of knowledge that is held by women (Pfeiffer & Butz 2005).

Pregnancy, parturition and the puerperium each mark significant steps in matrescence (Mathews & Manderson 1981), and are not without risk to the mother and infant. Infant mortality (deaths per 1000 live births) and maternal mortality (maternal deaths per 100 000 live births) rates in many developing countries are respectively 10 and 100-fold those for Sweden (World Health Organization 2007; Population Division of the Department of Economic and Social Affairs of the United Nations Secretariat 2008).

The postpartum period is important in many Southeast Asian cultures, and is seen as a period of recovery and often entails a period of confinement ranging from 10 up to 45 days. In accordance with the terminology of hu-moral medicine pregnancy is seen as a hot state; with parturition heat is lost and the woman comes into a state of excess cold, and during the postpartum period care should be taken to restore the mother to equilibrium (Manderson

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2003). Confinement as a treatment involves staying inside and near heat, washing only with hot water, drinking hot drinks, eating hot food, and stay-ing away from draughts (Manderson 2003). Confinement as a term is fairly broad and can include steam bath and bathing, mother roasting, dietary pro-scriptions and consumption of medicinal plant decoctions.

Steam bath is common throughout Southeast Asia (for review see Man-derson (2003)). Steam saunas or baths often involve making a decoction out of medicinal plants. The participant may either take a bath directly as is common for the Yao in Laos, Thailand and Yunnan province, China (Long & R. Li 2004; S. Li et al. 2006), sit in a special room into which steam from the decoction is led (Manderson 2003), or sit in a tent-like construction with (Liulan et al. 2003), or on a seatless chair over (Zumsteg & Weckerle 2007), a pot containing the steaming decoction. By lifting the lid of the pot the steam is let out, containing essential oils and other volatile substances, for inhalation and absorption through the skin, and as the water cools down the mother uses the decoction to cleanse her body.

Mother roasting is culturally related to steam bath, but differs in that the convalescent individual lies on a bed placed above a brazier with charcoal embers on which aromatic plants are laid, thus enabling the essential oils to vaporize. Mother roasting is often done to ‘dry out’ and to stop the lochia, restore the uterus to pre-pregnancy condition and to alleviate postpartum abdominal pain (Manderson 2003). Noncompliance is regarded as having negative consequences to both maternal and child health (Pottier 1978). Mother roasting seems to be confined to Southeast Asia, where it is wide-spread and reported from Laos (Spire 1907; Halpern 1961), Malaysia (Manderson 1979), Myanmar (Nash 1965), the Philippines (Hart 1965), Thailand (Hanks 1966) and Vietnam (Coughlin 1965). Many of these cita-tions are from the middle of the last century, and the prevalence of mother roasting in Southeast Asia today is not very well-known. However Barennes et al. (2009) report that 97 % of mothers in their study of postpartum trad-tions among suburban Lao in Vientiane practice exposure to hot beds of embers. In our research with the Brou, Saek and Kry we found that all in-formants observed mother roasting (de Boer & Lamxay 2009), except the Kry Thae (Lamxay et al. 2011).

In the context of the introduction and modernization of primary healthcare systems in rural areas, and with training programs for traditional birth at-tendants focusing on the paradigms of Western medicine, traditional knowledge relating to women’s health has often been ignored (Jordan & Davis-Floyd 1993). Erosion and deterioration of traditional medical knowledge can be observed in many cultures and leads not only to a loss in biocultural diversity, but also in diversity of alternatives for primary healthcare (Maffi 2005). Documenting the use of plants by ethnic minorities is not only an important aspect of understanding and analyzing elements of

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traditional birth practices, but also a way to perpetuate knowledge at risk of being lost.

Traditional repellents Vector-borne diseases, such as malaria, dengue fever, and Japanese encepha-litis, are endemic in several Southeast Asian countries (Eldridge & Edman 2004). Exposure to the disease vectors in rural areas can be minimized by the use of insecticide-impregnated bed nets and arthropod repellents. The cost of synthetic chemicals for impregnation or use on human skin or clothes can be an important factor limiting their optimal use. Mosquito control pro-grams often distribute bed nets and chemicals for impregnation free of charge, and train people on how to impregnate and use the nets. Sometimes it is required that people subsequently pay for continued impregnation of the nets. In the countryside of Lao PDR, where 44% of the population lives be-low the World Bank poverty line (World Bank 2008), costs of U.S. $4 per net and U.S. $1 per impregnation are often unaffordable.

The value of plants traditionally used to repel blood-feeding arthropods has often been underestimated (Sukumar et al. 1991; Pålsson & Jaenson 1999; Moore & Lenglet 2004). However, certain plant species have relative-ly recently become important sources for valuable repellents, e.g., p-menthane-3,8-diol (PMD) from Corymbia citriodora and citronellal from both C. citriodora and Cymbopogon citratus (Debboun et al. 2007). Com-pared to synthetics many of these plant-based repellents have low toxicity, but low durability, with the notable exception of PMD that has long efficacy (Debboun et al. 2007). In addition, plants used to obtain extracts or oils con-taining natural repellents can often be easily cultivated at a low cost in most tropical areas; especially practical are herbaceous or shrub-like weedy spe-cies.

Traditional medicine utilization is officially supported by the Lao gov-ernment, and is considered part of the cultural heritage as well as useful in view of the problems of access to modern pharmaceuticals (Sydara et al. 2005). Traditional medicine is largely plant based and available at low cost. However, the majority of Lao people do not use plant-based traditional re-pellents against blood-feeding invertebrates, and there is no government-initiated research on traditional arthropod repellents (Vongsombath 2011).

Phylogeny, dating and biogeography of Trichosanthes The family Cucubitaceae consists of about 95 genera and 950-980 species (Schaefer & Renner 2011b), and includes some of the most important vege-table crops, such as melon (Cucumis melo), cucumber (C. sativus), water-

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melon (Citrullus lanatus), and squash and pumpkin (Cucurbit spp.), as well as a myriad of lesser-known useful species, e.g. luffas, calabash, winter mel-ons, and chayote (Schaefer & Renner 2011a). Trichosanthes is the largest genus in Cucurbitaceae, with over 90 species. It includes the snake gourd (Trichosanthes cucumerina), which is popular in South and Southeast Asian cuisine, and roughly 15 taxa that are important in traditional medicine (Perry & Metzger 1980). Gymnopetalum includes four species (de Wilde & Duyfjes 2006), some of which are used in traditional medicine (Perry & Metzger 1980).

Trichosanthes are mostly stout perennial climbers, 3 – 30 m long, with branched tendrils, mostly dioecious flowers, distinctly fringed petals, and often egg-sized brightly colored fruit. Palynological studies (Pruesapan & Van Der Ham 2005) have indicated that a diversity of pollen types exist in the genus with 3-(4-)porate and 3-(4-)colporate pollen with psilate, perforate, verrucate, reticulate, and regulate ornamentation. Dioecy, variation in vege-tative morphology, and incomplete herbarium collections complicate taxo-nomical studies, and have contributed to the description of nearly 300 taxa in Trichosanthes (de Wilde & Duyfjes 2010; IPNI 2011). No full taxonomic treatment of the genus exists, but in recent years revisions have been pub-lished for most of its distribution: India (Chakravarty 1959), China (Lu et al. 2011; Yueh & Cheng 1974; Yueh & Cheng 1980), Thailand (Duyfjes & Pruesapan 2004), Cambodia, Laos and Vietnam (Keraudren-Aymonin 1975), Malaysia, Indonesia, the Philippines and Papua New Guinea (Rugayah & de Wilde 1997; Rugayah & de Wilde 1999; de Wilde & Duyfjes 2004; de Wilde & Duyfjes 2010), Australia (Telford 1982; Cooper & de Boer 2011) and Japan (Ohba 1984).

Gymnopetalum strongly resembles certain Trichosanthes species, but the former lack thread-like fringes on the petals, and the overall shape of the folded petals in the mature bud is elongate, and longer than broad (short and rounded in Trichosanthes) (de Wilde & Duyfjes 2006). A revision of the genus was published by de Wilde & Dufyjes (2006), with minor nomencla-tural changes published later (de Wilde & Duyfjes 2008).

Infrageneric classifications of Trichosanthes have been proposed by vari-ous authors (Yueh & Cheng 1974; Jeffrey 1980b; Chen 1985; Huang et al. 1997; Rugayah & de Wilde 1999; de Wilde & Duyfjes 2004). The most re-cent classification of the genus by Rugayah & de Wilde (1999) classifies Trichosanthes into five sections, plus one section described in 2004 (de Wilde & Duyfjes 2004): sect. Trichosanthes, sect. Cucumeroides including subsect. Cucumeroides and subsect. Tetragonosperma, sect. Edulis, sect. Foliobracteola, sect. Involucraria including subsect. Involucraria and sub-sect. Pedatae, and sect. Asterosperma. However, both Rugayah & de Wilde (1999) and de Wilde & Duyfjes (2004; 2010) are reserved in their infrage-neric classifications, and mention a need for further investigation. The genus Gymnopetalum has been divided into two sections, Gymnopetalum contain-

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ing the type G. tubiflorum from southern India and Sri Lanka, and Tripo-danthera containing the southeast Asian and Malesian species (Cogniaux 1881). However this classification was not maintained by later authors (Jef-frey 1980a; Philcox 1997; de Wilde & Duyfjes 2006).

Trichosanthes has its center of origin in Southeast Asia, but ranges from India throughout Asia east to Taiwan, the Philippines and Japan, and south-east to New Guinea, Australia, Fiji and Vanuatu (de Wilde & Duyfjes 2010). Sections Trichosanthes, Cucumeroides, Foliobracteola, and Involucraria occur widely in mainland Asia, the Malay archipelago, and Australia. Sec-tion Asterosperma is restricted to Borneo (de Wilde & Duyfjes 2010), and section Edulis to New Guinea, the Solomon Islands, Vanuatu (de Wilde & Duyfjes 2010) and Australia (Cooper & Ford 2010). In Japan only sections Cucumeroides, Foliobracteola, and Involucraria are found (Ohba 1984). Gymnopetalum ranges from India through China and Southeast Asia into the Malay archipelago but does not cross Lydekker’s Line and is thus absent from the Sahul shelf (Lydekker 1896; de Wilde & Duyfjes 2010) (Figure 1).

Figure 1. Sunda and Sahul, indicating the Wallace, Weber and Lydekker Lines. Wallacea is the area in between the Wallace and Lydekker Line. Courtesy of Maxi-milian Dörrbecker, Wikipedia, Creative Commons license.

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Aims The research presented in this dissertation all relates to useful plant species from Laos. Three major subjects can be recognized.

Papers I, II, III, and IV all concern aspects of medicinal plant use by the Brou, Saek, and Kry ethnic groups living in Khammouan Province, with an emphasis on postpartum healthcare. Paper I aims to understand the diversity and importance of medicinal plants during pregnancy, parturition and post-partum recovery to the Brou, Saek and Kry ethnic groups. Paper II further explores postpartum medicinal plant use by the Kry, and includes detailed descriptions of Kry spatial taboos, pregnancy, parturition and confinement traditions. Paper III aims to identify the major essential oil constituents in the three most salient species used in mother roasting, steam sauna and steam bath, and correlates known bioactivity of these constituents with the exact traditional use in an attempt to elucidate the possible function and effi-cacy of the species and practices.

Papers V and VI; two excluded papers on plant repellents, one against leeches (Vongsombath et al. 2011), and one against mosquitoes (Vongsombath 2011); and an excluded manuscript on the effects of storage material and temperature on Hyptis suaveolens essential oil constituents (de Boer et al., in prep.); deal with traditional plant-based repellents against par-asitic arthropods and leeches. Paper V is the result of a national survey on traditional plant use to repel mosquitoes, leeches, ticks, mites, lice, and flies in Lao PDR. It aims to determine what species to select for subsequent repel-lence studies to find economical alternatives to synthetic repellents by estab-lishing what species are traditionally used as repellents (cf. Paper VI, Vong-sombath et al. (2011), Vongsombath (2011)). Paper VI aims to evaluate the repellent and larvicidal efficacy against blowfly larvae of the three most salient plant species used for the preservation of fermented fish.

Papers VII, VIII and IX report on molecular phylogenetics, molecular dating, biogeographical analyses, and morphological and nomenclature stud-ies of the snake gourd genus, Trichosanthes (Cucurbitaceae). Paper VII aims to test the monophyly of Trichosanthes and Gymnopetalum using a broad sampling of ~60% of their species and 4756 nucleotides of nuclear and plastid DNA. Cucurbitaceae fossils, species distribution ranges and the phy-logenetic data are used to obtain age estimates and to reconstruct the diversi-fication patterns and biogeographic history of the genera. Paper VIII ad-dresses the taxonomic implications that the phylogenetic analyses of the Trichosanthes and Gymnopetalum molecular data have for the taxonomy of both genera at subgenus, section and species level. Paper IX is a revision of the Australian species of Trichosanthes, including notes on habitat, distribu-tion maps and three identification keys. These species are of special interest as their presence there are the result of separate lineages colonizing Austral-asia (cf. Paper VII). The last treatment for Australia stressed that the taxon-

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omy was poorly known, critical comparison of Australian and Asian material was required, raised doubts about the uniqueness of T. holtzei, included an undescribed new taxon, Trichosanthes sp. A, and suggested that T. pen-taphylla might be conspecific with T. trifolia.

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Summary of Papers

In the following pages each paper will be summarized independently to give the reader a quick insight into its main objectives and results. To avoid re-dundancy these summaries do not elaborate on, or repeat too much of, the content of the papers. In addition reprints of all papers and/or manuscripts are appended to the printed version of this dissertation.

Paper I

Background This paper is the first manuscript I wrote intended for this dissertation, and was written during my time as a research fellow with Prof. Bruce Baldwin and Prof. Tom Carlson at the Jepson Herbarium, University of California at Berkeley. It was written at the herbarium, at the Willard Park playground and in our studio apartment on Otis St.

The study was conceived by Vichith Lamxay and myself, and was in-spired by Jeanine Pfeiffer and Ramona Butz’ paper on the importance of gender in relation to accessing cultural and ecological variation in ethnobi-ological research (Pfeiffer & Butz 2005). Their review pointed out that the role of women as repositories of traditional knowledge was neglected and overlooked, and that this was largely due to a male bias in researchers and informants. Data was collected during three field expeditions in the Nakai-Nam Theun reservoir catchment in Khammouan Province in December 2005, June 2006 and February 2007, and female interviewers conducted all interviews with female informants. Oudomphon Insisiengmay and Tou-lavanh Xayphone recorded the majority of the data, under supervision of Anneleen Kool.

The role Vichith Lamxay and I played in the data collection was to con-duct interviews with male informants simultaneously with the interviews with female informants, to ensure that the female interviews could proceed without interruption.

Study In many Southeast Asian cultures a culturally dictated period of postpartum confinement is observed. Plants play an important role in this recovery pro-

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cess, and are used during the postpartum period in local diet, traditional medicine, steam baths and during a practice dubbed mother roasting.

This research investigates the use of plants during pregnancy, parturition, postpartum recovery and infant healthcare among three ethnic groups, the Brou, Saek and Kry. Data were collected in 10 different villages through group and individual interviews with women by female interviewers. The results show that a total of 55 different plant species are used in women’s healthcare, of which over 90 % are used in postpartum recovery. Consensus Analysis rejected the hypothesis that the three ethnic groups belong to a sin-gle culture for postpartum plant use, and multidimensional scaling revealed non-overlapping clusters per ethnic group. The study presents the first data on plant use by the Brou, Saek and Kry, and one of the few studies on plants used in women’s healthcare among the ethnic groups of mainland Southeast Asia. It provides a wealth of novel insights into plant use during postpartum recovery rituals such as steam baths and mother roasting.

Relevance The study and its results are relevant for culturally appropriate integration of modern healthcare and traditional postpartum recovery practices among eth-nic minorities in Lao PDR. Large populations in rural Lao PDR live far from modern healthcare facilities, and rely on traditional practices and medicinal plants as their primary source of healthcare, and these practices are essential in their perception of postpartum recovery. These practices conflict with modern perceptions of antenatal care and postpartum recovery, and are mar-ginalized or discouraged by efforts to introduce modern healthcare as health care workers are concerned about dietary intake and hygiene (Barennes et al. 2009). The importance of these practices can be illustrated by data from villages on the outskirts of the capital Vientiane, where prevalence of tradi-tional postpartum practices was found to be: mother roasting (97 % of in-formants observed this after childbirth), herbal tea as sole beverage during postpartum confinement (95 %), and dietary proscriptions (90 %) (Barennes et al. 2009). This data shows that these practices are deeply ingrained in Lao society, and that modern perinatal care cannot replace them. An antagonist approach to introducing modern healthcare aimed at replacement of tradi-tional practices has not yielded the intended outcome. Instead it created a parallel system of healthcare, where communication is discouraged by each system’s negative perception of the other. Rural areas where modern healthcare is not widely available could serve as exemplary cases of cultural-ly sensitive integration of modern and traditional perinatal care benefitting local populations by reduced infant and maternal mortality, improved nutri-ent and dietary intake by newborns, infant, and convalescent mothers, and improved hygiene to reduce septicemia and other infections.

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Paper II

Background This paper builds on results found in the study for paper I, and was written during the winter of 2010-2011 by Vichith, Lars and myself. It was pub-lished in May 2011 just days before Vichith’s PhD defense.

The research was conceived after completion of Paper I, when we realized that variance in the data regarding the Kry ethnic group could not be at-tributed only to a lack of data collection, i.e. we had found that the Kry give birth outside the village in a specifically built hut, and use relatively few medicinal plants. We decided to make a special effort to interview the Kry on postpartum plant use and traditions. Vichith together with a number of female students from the Biology Bachelor’s and Master’s program at NUoL gathered in-depth data in the three Kry villages in the upper Nam Noi water-shed in Khammouan province during 2009 and 2010.

Study Plants play an important role in recovery during the postpartum period in diet and traditional medicine. However, little is known of the Kry, a small ethnic group whose language was recently described (Enfield & Diffloth 2009), concerning its traditions and use of plants during pregnancy, parturi-tion, postpartum recovery and infant healthcare. Recent previous studies reported on traditions and customs (Enfield 2009), and some of the plant used during postpartum recovery (de Boer & Lamxay 2009). This research aims to study those traditions and identify medicinal plant use. Data were collected in all three Kry villages in the upper Nam Noi watershed in Khammouane province, through group and individual interviews with wom-en by female interviewers. A total of 49 different plant species were record-ed for use in women’s healthcare. Plant use is culturally different from the neighboring Brou and Saek ethnic groups. Menstruation, delivery and post-partum recovery take place in separate, purpose-built huts and a complex system of spatial restrictions is observed. Traditions surrounding childbirth are diverse and have been strictly observed, but are undergoing a shift to-wards those from the neighboring ethnic groups, the Brou and Saek. Medici-nal plant use to facilitate childbirth, alleviate menstruation problems, assist recovery after miscarriage, mitigate postpartum hemorrhage, aid postpartum recovery, and for use in infant care, is more common than previously report-ed (49 species instead of 14).

Relevance The wealth of novel insights into plant use and preparation will help to un-derstand culturally important practices such as traditional delivery, spatial

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taboos, confinement and dietary restrictions, and their potential role or posi-tion in modern healthcare.

In addition, this study is the first to report in detail the parturition and postpartum practices of the Kry. The cultural importance of, and strong ad-herence to, parturition- and menstruation practices and spatial taboos, in combination with a small and impoverished population and high infant- and maternal mortality, make culturally appropriate introduction of modern ante-natal care difficult. Case-studies from another ethnic group with similar childbirth practices, the Katang, show that women are in favor of having birth at medical clinics as they are aware of the dangers of forest childbirth (Chithtalath & Earth 2001). It is not known whether the Katang have similar spatial taboos, and how clinic childbirth is integrated into their cultural be-liefs and traditions. The challenges of integrating partner-aided childbirth in a makeshift hut outside the village, followed by postpartum recovery in an often-ramshackle construction used by women during menstruation near the parental house, with a modern medical approach to parturition stipulating the importance of skilled birth attendants in an aseptic setting, are hard to over-come without the help of specialized midwives, medical anthropologists and gynecologists wishing to maintain a Kry cultural character.

Paper III

Background Right form the start of the research on Amomum (Zingiberaceae) by Vichith we aimed to undertake chemical studies on the secondary metabolite constit-uents of Amomum. Our first intention was to compare metabolite spectra between different species collected as non timber forest products for com-mercialization. Amomum is a significant exported natural resource in Lao PDR, and finding ways to differentiate seed capsules of material in trade would aid assessing the importance of different species in this trade. Another aspect was to identify those species with the highest essential oil content, and to select these for in-vitro micro-propagation. Unfortunately the taxo-nomic work on Amomum was more challenging than expected, as nearly each fieldtrip resulted in finding a new species, a collaborator on chemical analysis in Vietnam vanished into thin air, and a set of distilled oil samples together with the collaborator. Simultaneously we were gathering an ever-growing body of reports on the importance of Amomum in postpartum re-covery practices, and soon we realized that the metabolite spectra were more interesting to explain the role of the volatile constituents in steam bath, steam sauna and mother roasting. Plan B is often better than plan A. The Sarawak Biodiversity Centre carried out the GC-MS analysis, and we are very grateful for their help.

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Study Fundamental in traditional postpartum recovery in Lao PDR is the use of hotbeds, mother roasting, steam sauna and steam baths (de Boer & Lamxay 2009; Barennes et al. 2009). During these treatments medicinal plants play a crucial role, but little has been published about how the treatments are car-ried out precisely, which species are used, the medicinal properties of these species, and the medicinal efficacy of their chemical constituents (de Boer & Lamxay 2009; Lamxay et al. 2011). Sixty-five interviews, in 15 rural villag-es, with women of four different ethnic groups were conducted to survey confinement rituals, and postpartum plant use and salience. A total of 10 different species were used by three or more of the ethnic groups included in this study. All species were used in steam sauna and bath, but only three species were used in hotbed and mother roasting. Essential oils from the main species used were extracted using in-situ hydro-distillation and the main chemical constituents characterized using GC-MS. Essential oils of Amomum villosum, Amomum microcarpum and Blumea balsamifera were found to contain significant amounts of the following terpenes: β -pinene, camphor, bornyl acetate, borneol, linalool, D-limonene, fenchone, terpinen-4-ol and α-terpinene. Many of these terpenes have documented antimicrobial and analgesic properties, and some have also synergistic interactions with other terpenes. The mode of application in hotbed and mother roasting dif-fers from the documented mechanisms of action of these terpenes. Plants in these two practices are likely to serve mainly hygienic purposes, by segre-gating the mother from infection sources such as beds, mats, stools, cloth and towels. Steam sauna medicinal plant use through inhalation of essential oils vapors can possibly have medicinal efficacy, but is unlikely to alleviate the ailments commonly encountered during postpartum convalescence. Steam sauna medicinal plant use through dermal condensation of essential oils, and steam bath cleansing of the perineal area is possibly a pragmatic use of the reported medicinal plants, as terpene constituents have document-ed bioactive properties.

Relevance The importance of steam sauna, steam baths and mother roasting to postpar-tum recovery traditions in Lao PDR is indisputable, as over 90 % of people in suburban areas with convenient access to modern healthcare facilities observe these and other practices (Barennes et al. 2009). Modern healthcare initiatives are often very critical of the traditional practices, but is this skep-ticism justified? This study identifies the most important species, records their exact quantities and use within each of these traditions, and analyzes the essential oil constituents from the plant parts used. Although explicitly not a clinical study or attempt at such, it does correlate the documented bio-active properties of the terpenoid constituents with the reported and inter-

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preted functions of these during postpartum practices. The results show that the use of these specific species likely aids postpartum hygiene and health through the antimicrobial, analgesic and anti-inflammatory properties of their essential oil constituents.

Paper IV

Background The data behind this paper was possibly the main objective of studying me-dicinal plant use in among the Brou, Saek and Kry in the Nam Theun water-shed. Right from the start of the project we were interested in knowing the mechanisms and modes of transmission of traditional knowledge between ethnic groups living in the same area, i.e. with the same species available to all people. In practice, this turned out to be more challenging than expected as each new interview could yield new data, plants collected for identifica-tion would be sterile, or used species would only occur ‘in the forest’. To make the data manageable we decided to focus on the 115 species mentioned during the first three research trips, and exhaustively collect data on all pos-sible uses of these species in subsequent trips. In addition to this paper, this data is also used to create an illustrated popular scientific handbook on the Brou, Saek and Kry medicinal plants written in Lao intended for returning to the people who helped us in the field. This handbook is now ready to print.

Study Medicinal plant traditional knowledge is one of the most widely known val-ues of traditional ecological knowledge, as it provides primary healthcare, contributes to subsistence livelihoods, and for its potential value as a source of novel pharmaceuticals. People living in close contact with their surround-ings for many generations are hypothesized to have developed, through trial-and-error, in-depth knowledge of ecosystems, biodiversity, and their man-agement and utility. At the same time, people living a traditional lifestyle in tropical rainforests often suffer poor health, have short life expectancies, and lack effective cures to many ailments. This paradox between ancient tradi-tional medicine and a lack of effective cures raises the question whether medicinal plant knowledge results in an asymptotic climax, or a constantly evolving equilibrium of cures with proven efficacy and those under assess-ment. An in-depth study of 97 plant used in traditional medicine by the Brou, Saek and Kry ethnic groups in Lao PDR was made to test similarity in me-dicinal plant knowledge. Medicinal plants were used in 99 different ways in 510 species-use combinations. Medicinal uses could be generalized into 12 use categories with 747 species-category combinations. Sørensen (QS) and Jaccard (JI) similarity indices show Brou and Saek plant use appears to be

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most similar (QSBS 60.0; JIBS 75.1) followed by Kry and Saek (QSKS 51.6; JIKS 53.4), and then Kry and Brou (QSBK 46.9; JIBK 44.1). Intercultural simi-larities are quite low, considering that all three groups share the same geo-graphical and ecological area and have the same dependence on medicinal plants. Intercultural transmission is unimpeded but many cures are likely to be ineffective. Comparison of the similarities found here with similarities computed from other data show that these results are homologous with other sympatric ethnic groups, and much higher than those for allopatrically living groups. Medicinal plant knowledge does not reach a climax, but appears to continually evolve by trial-and-error, as effective cures to many ailments are unavailable.

Relevance The levels of similarity in medicinal plant use found in this study are possi-bly indicative of sympatric intercultural medicinal plant traditional knowledge similarities. Cures that appear to be effective might spread to other communities or ethnic groups, as intercultural transmission is unim-peded, but people still lack effective cures for the most common ailments, and keep innovating to find new cures. The resulting similarities between groups are not likely destined to attain a knowledge climax, but will remain a constantly changing patchwork of remedies that show efficacy, are of cul-tural significance, or currently under evaluation. These results are relevant as they tarnish the notions that traditional plant knowledge is ancient, the result of numerous generations of oral transmission and slow trial and error knowledge acquisition, and irreplaceable. Undoubtedly traditional medicinal plant knowledge develops within each culture and is an important part of a culture’s heritage, but medicinal plant use may differ from year to year and from generation to generation, as new remedies are sought to provide essen-tial healthcare.

Paper V

Background The research project by Chanda Vongsombath (Vongsombath 2011) aimed at evaluating the repellent activities of traditionally used plants against common hematophagous parasites, e.g. leeches, mosquitoes, and ticks. An essential foundation for this study was a clear understanding of what plant species are used in Lao PDR as repellents, and with that objective we con-ducted an extensive survey of repellent plant use throughout Lao PDR. The first three research trips Southern, Central, and Northern Lao PDR, were conducted by Chanda and myself together, and we exchanged theory and experiences on quantitative data collection, structured interviews, and plant

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voucher collections. During the trip in Northern Laos in the winter of 2007, I was without my check-in luggage, and spent it freezing in an undersized outfit bought in the market of Xieng Khouang. During the trip in Southern Laos in the fall of 2007 we were accompanied by my 4-months old son Flo-ris, and people were vastly more interested in our research that time.

The manuscript was drafted during my time as a research fellow at UC Berkeley, and the University of California’s library has been an invaluable source of reference literature for the literature investigation into the previ-ously documented repellent activities.

Study Hematophagous parasites such as leeches, ticks, mites, lice, bedbugs, mos-quitoes, and myiasis-producing fly larvae are common health problems in Lao PDR. Several arthropod-borne infections, e.g., malaria, dengue fever, and Japanese encephalitis, are endemic there. Effective vector control meth-ods including the use of pesticides, insecticide-treated bed nets, and synthet-ic and plant-based repellents are important means of control of such inverte-brates and the pathogens they may transmit. In this study, we documented traditional knowledge on plants that are used to repel or kill hematophagous arthropods, including mosquitoes, bedbugs, human lice, mites and ticks, fly larvae, and blood-sucking leeches. Structured interviews were carried out in 66 villages comprising 17 ethnic groups, covering a range of cultures, throughout Lao PDR. A total of 92 plant species was recorded to be used to obtain traditional repellents (including plants for pesticidal applications) in 123 different plant-ectoparasite combinations. The number and species of plants, and animal taxa repelled (or killed) per plant species differed per region, village, and ethnic group. Traditional use was confirmed in the scien-tific literature for 74 of these plant species, and for an additional 13 species using literature on closely related species. The use of botanical repellents and pesticides from many plant species is common and widespread in the Lao countryside. In the future, the identification of the active components in certain plants to develop more optimal, inexpensive repellents, insecticides, acaricides, or antileech compounds as alternatives to synthetic repellents or pesticides against blood-feeding insects, ticks, mites, and leeches, is war-ranted.

Relevance A key objective of these studies is to find inexpensive and accessible alterna-tives to synthetic repellents and pesticides for the impregnation of bed nets. During surveys local participants indicated that the cost of synthetic impreg-nation of bed nets was a major financial hurdle, even though a cost of US$ ~1 per impregnation to us may seem very minor. Knowing which common species are effective alternatives to costly synthetic repellents is likely to increase the willingness of people to impregnate, as well as the frequency of

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bed net impregnation, both of which could subsequently reduce the inci-dence of vector-borne diseases.

The ninety-two species of repellent plants reported and identified during this study are probably only a fraction of the total number of species used throughout the country. Knowing which plant species are effective as repel-lents, against which animal species, for which duration of time, and how to prepare and preserve extracts of these plants, is difficult for local people to do as most lack time and training for structured study and analysis of these properties. At the same time, hematophagous parasites are omnipresent and an important source of disease and mortality in the Lao countryside, and this knowledge could contribute significantly to health prospects. This survey and subsequent studies (de Boer et al. 2010; Vongsombath et al. 2011; Vongsombath 2011; de Boer et al. 2011) all aim to elucidate the diversity of used species and the efficacy of the most salient species (those that are men-tioned most frequently) through repellency studies. One of the most promis-ing species, the introduced Hyptis suaveolens (L.) Poit., was also the subject of an unfinished study on the preservation of essential oil extracts under different storage conditions.

Paper VI

Background This paper is based on a manuscript previously included in the dissertation of Chanda Vongsombath (Vongsombath 2011), and currently published in extensively revised form in PLoS ONE with a different author order and a new co-author. The design of the study was a collaborative effort by Thomas Jaenson, Chanda Vongsombath and me, whereas the execution was the sole responsibility of Chanda. The editorial comments on the manuscript after reviews implied a major effort to get the paper into shape and accepted. The contributions to the manuscript by Chanda and me so far had been very bal-anced, but this would clearly tip the balance towards me, and we decided together that I would be the first author, and that we would invite Dr. Jos Käfer from the Laboratoire de Biométrie et Biologie Evolutive at the Uni-versité de Lyon to assist with statistical analysis, the R-package and data presentation.

The research originates from the repellents survey (Paper V), where unan-ticipatedly we collected a lot of data on repellence of fly larvae from fer-mented fish. One of the interview questions concerned the use of plants to repel fly larvae from infested wounds, and responses related either to repel-ling fly larvae from infested wounds in cattle, or infestation in the traditional staple of fermented fish.

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Study In rural areas in Laos, fly larvae infestations are common in fermenting fish. Blowflies (Chrysomya megacephala, Diptera: Calliphoridae) are attracted to oviposit (and/or larviposit) onto fermenting fish which results in infestations with fly larvae. Our survey of traditional repellents found that knowledge of traditional use of plants to repel larvae during the production of fermented fish is common and widespread in Lao PDR. But how effective are the most salient species in repelling, and killing fly larvae in fermenting fish? To an-swer this we tested the three plant species most frequently reported to repel fly larvae for repellence and larvicidal activity, Tadehagi triquetrum (Faba-ceae), Uraria crinita (Fabaceae) and Bambusa multiplex (Poaceae). The results show that the repellent effect of fresh material of T. triquetrum and U. crinita, and the larvicidal effect of fresh B. multiplex, is significantly more effective than that of dried material of the same species, and the total effect (repellence and larvicidal effect combined) for each of the three spe-cies was significantly more effective for fresh than for dry material. The total effect of repelling and killing was not significantly different per species, 72%, 82%, and 77%, respectively. In conclusion we can state that the three most salient species are effective in repelling and killing fly larvae in the production of fermented fish. Their use may be essential to augment food safety during fermentation in traditional earthenware jars.

Relevance Proving that traditionally used plants are effective for the purpose they are traditionally used for may not seem like the most relevant research. Especial-ly when considering that nowadays many people in urban areas, and facili-ties for larger scale production, use plastic vats that can be properly sealed to avoid larval infestations. The three reviewers of the manuscript also men-tioned these points.

A closer look at the present situation in Lao PDR can clarify and justify the research. The majority of people in rural Lao PDR rely on earthenware jars for the fermentation of fish. In the future a shift from earthenware jars to plastic vats can be predicted in the countryside. This shift in all probability will be accompanied by a new resurgence in the use of earthenware jars, as middle class Lao citizens already today opt for a more traditional and better-flavored production of fermented fish. In other words, earthenware jars will remain a common and preferred method of fermented fish production and storage.

The repellence and larvicidal activity found in this study show that T. tri-quetrum and U. crinita are most effective in repelling fly larvae, and that B. multiplex is most effective in killing larvae. Data on other species used for repelling fly larvae is unavailable, but these three species have documented efficacy. Translating these results back into practice one should use either T.

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triquetrum or U. crinita to prevent blowflies from ovipositing in fermenting fish, or prevent fly larvae hatched at the mouth of the jar from moving to-wards the fish. B. multiplex can best be used to kill fly larvae when an infes-tation is discovered, but after first having attempted to remove the maggots.

Paper VII

Background Plant systematics and molecular biology have long been an interest of mine, and during the planning phase of the research proposal we identified differ-ent plant genera of importance to traditional medicine in Southeast Asia, and Lao PDR in particular, that were of interest for further systematic research. In the genus Trichosanthes (Cucurbitaceae) roughly 15 species are used in traditional medicine in Southeast Asia. The most important species is Trich-santhes kirilowii, which is included in the ancient Chinese Materia Medica (Mosig & Schramm 1955). It is reported to be used as expectorant, to treat hemorrhoids and dermal dryness (Chung 1961), be thirst-quenching (Liu 1952), have bechic, antipyretic, diuretic, lactagogue properties, and used as a treatment for ulcers, wounds, heat rash, chicken pox and other skin diseases (Chu 1968), be used as antiemetic, laxative cooling agent, depurative, siala-gogue, maturative, to treat sunstroke, sore throat, jaundice and large boils (Chung yao chih 1959), to treat coughs, tuberculosis, and hemoptysis (Mosig & Schramm 1955), a remedy for diabetes (Ishidoya 1925) and skin swellings (Roi 1955). Other species in the genus are used as substitutes for similar purposes, but also for other purposes such as treatment of infant diarrhea (T. tricuspidata), miscarriage recovery (T. wallichiana), malaria (T. wawraei), and postpartum complications (T. villosa) (Perry & Metzger 1980).

The genus Trichosanthes was by far the most interesting genus to study as it included a large number of medicinal species, was the subject of multiple finished and ongoing taxonomic revisions (Chakravarty 1959; Yueh & Cheng 1974; Keraudren-Aymonin 1975; Telford 1982; Ohba 1984; Rugayah & de Wilde 1997; Huang et al. 1997; Khunwasi 1998; Rugayah & de Wilde 1999; de Wilde & Duyfjes 2004; Duyfjes & Pruesapan 2004; Pruesapan & Van Der Ham 2005; de Wilde & Duyfjes 2006; de Wilde & Duyfjes 2010; Lu et al. 2011), and the status of its phylogenetic position within Cucurbita-ceae was only partly resolved (Jobst et al. 1998; Chung et al. 2003; Zhang et al. 2006).

Willem de Wilde and Brigitta Duyfjes helped with obtaining plant sam-ples for DNA extraction from their extensive collections from Indonesia and Thailand, as well as providing assistance on morphological and nomencla-tural questions. Two papers published in 2007 and 2008 (Kocyan et al. 2007; Schaefer et al. 2008) initiated a fruitful collaboration with Hanno Schaefer

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and Susanne Renner in which we combined samples and efforts to complete a full phylogeny of the genus.

Papers VII, and VIII and IX all relate to research on Trichosanthes. Paper VII presents the results of the phylogenetic study on the corolla fringes of Trichosanthes, and includes molecular dating and biogeographical analysis. Paper VIII deals with the nomenclatural implications of Paper VII and in-cludes a synopsis of all recognized species in the genus. Paper IX is a revi-sion of the genus in Australia, and builds on earlier work (Telford 1982; Cooper & Ford 2010), as well as unpublished data for a detailed phylogeny of the species occurring in Australia.

Study The Cucurbitaceae genus Trichosanthes comprises 90-100 species that occur from India to Japan, and southeast to Australia and Fiji. Most species have large white or pale yellow petals with conspicuously fringed margins, the fringes sometimes several cm long. Pollination is usually by hawkmoths. Previous molecular data for a small number of species suggested that a mon-ophyletic Trichosanthes might include the Asian genera Gymnopetalum (four species, lacking long petal fringes) and Hodgsonia (two species with petals fringed). In this study we tested these groups’ relationships using a sampling of 60% of their species and 4759 nucleotides of nuclear and plastid DNA. Time and direction of the geographic expansion of the Trichosanthes clade was inferred using molecular clock dating and statistical biogeographic reconstruction, and we also address the gain or loss of petal fringes. The results show that Trichosanthes is monophyletic as long as it includes Gym-nopetalum, which itself is polyphyletic. The closest relative of Trichosanthes appears to be the sponge gourds, Luffa, while Hodgsonia is more distantly related. Of nine morphology-based sections in Trichosanthes, three are sup-ported by the molecular results; and two new sections appear warranted. Molecular dating and biogeographic analyses suggest an Oligocene origin of Trichosanthes in Eurasia or East Asia, followed by diversification and spread throughout the Malesian biogeographic region and into the Australian continent. In conclusion, we can say that long-fringed corollas evolved inde-pendently in Hodgsonia and Trichosanthes, followed by two losses in the latter coincident with shifts to other pollinators but not with long-distance dispersal events. Together with the Caribbean Linnaeosicyos, the Madagas-can Ampelosicyos and the tropical African Telfairia, these cucurbit lineages represent an ideal system for more detailed studies of the evolution and func-tion of petal fringes in plant-pollinator mutualisms.

Relevance These papers are not phrased in development research relevance as easily as papers I-VI, but were not undertaken with that direct goal. Indirectly the research does have clear relevance for Swedish international development,

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as it builds Swedish research capacity in tropical plant systematics and mo-lecular phylogenetics, both important competences in knowledge exchange.

Plant systematics and plant identification are essential skills needed in many developing countries to document biological diversity and to monitor and control the access and transfer of plant genetic resources. Sweden aims to have a leading role in the development and implementation of the Con-vention on Biological Diversity (CBD) through active involvement in the Conferences of Parties (COP), and capacity-building to identify and voucher plant material is a crucial need for many developing countries in order to safeguard equitable access and benefit sharing of genetic resources. Global initiatives to democratize access to taxonomic information such as the Glob-al Taxonomy Initiative (GTI), the Global Biodiversity Information Facility (GBIF) and the extended online digital library of herbarium vouchers, Aluka all contribute to this access and benefit sharing.

From a scientific perspective this study is relevant in that it answers many questions regarding the relations and evolution of the genera Trichosanthes and Gymnopetalum. The phylogenetic analyses show that the species of the fringeless-corolla genus Gymnopetalum consist of two groups, both of which are nested within the larger fringed-corolla genus Trichosanthes.

The phylogeny indicates that corolla fringes have evolved a single time at the basis of the Trichosanthes-Gymnopetalum clade, and disappeared twice in the Gymnopetalum clades. Most taxa in the Trichosanthes-Gymnopetalum clade flower at night, but data on pollinators is sparse, especially for the Gymnopetalum species. It is likely that corolla fringes are an adaptation to nocturnal flowering, and the absence of fringes could indicate a further adap-tation to a specific pollinator group where fringes have no benefit, or to di-urnal flowering, as is likely the case for Gymnopetalum.

Molecular dating and biogeographical analysis of the phylogeny support a South or Southeast Asian evolution of the genus with subsequent radiation throughout Southern Asia, the Malay Archipelago and Australasia. In addi-tion it reveals that the Australian belong to five independent lineages, of which at least three are have colonized Australasia by Pliocene/Pleistocene long-distance dispersal from the Sunda shelf, or island-hopping from the Wallacea archipelago. The Sunda shelf consists of those areas of the Malay Archipelago that have been inter-connected with mainland Asia by land bridges at some point during the Pleistocene glacial oscillations, and extends eastwards in the archipelago towards Wallace’s Line. Wallacea is the part of the archipelago in between Wallace’s Line and the more easterly Lydekker’s Line. Lydekker’s Line separates these areas from the Sahul shelf, the area consisting of Australia, Tasmania, New Guinea and the Tanimbar Islands, which were also all connected at some point during the Pleistocene glacial oscillations.

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Paper VIII

Background The phylogenetic analyses of the Trichosanthes and Gymnopetalum molecu-lar data have implications for the taxonomy of both genera at subgenus, sec-tion and species level. These taxonomic changes are addressed in this paper.

Study The snake gourd genus, Trichosanthes L., is the largest genus in the Cucur-bitaceae family, with over 90 species. Recent molecular phylogenetic data of Trichosanthes and Gymnopetalum indicate that the species of the Gym-nopetalum need to be merged into Trichosanthes to maintain monophyly. In addition, some of the described sections are supported, others require to be recircumscribed, and in one case a new section needed to be described.

A revised infrageneric classification of Trichosanthes including Gym-nopetalum is proposed consisting of two subgenera, (I) subgen. Scotanthus comb. nov. and (II) subgen. Trichosanthes, eleven sections, (i) sect. Aster-ospermae, (ii) sect. Cucumeroides, (iii) sect. Edulis, (iv) sect. Foliobracteo-la, (v) sect. Gymnopetalum comb. et stat. nov., (vi) sect. Involucraria, (vii) sect. Pseudovariifera sect. nov., (viii) sect. Trichosanthes, (ix) sect. Tripo-danthera comb. nov., and (x) sect. Truncata, and (xi) sect. Villosae, stat. nov. A synopsis of all 91 Trichosanthes species recognized is presented, including T. orientalis comb. nov, T. tubiflora comb. nov., as well as T. scabra var. pectinata comb. nov and T. scabra var. penicaudii comb. nov.

Relevance Phylogenetic studies as part of PhD research often imply significant taxo-nomic changes, which are omitted or postponed beyond the dissertation. I have often thought that this was unfortunate, but have also been understand-ing as taxonomic changes and revisions are time-consuming and require different skills than molecular phylogenetics. The changes stemming from the Trichosanthes phylogeny are relatively minor, and have been grouped together in this paper with a synopsis of all currently recognized species, sections and subgenera.

Paper IX

Background The Australian species of Trichosanthes are of special interest as their pres-ence there is the result of five separate lineages colonizing Australasia. T. cucumerina is most closely related to species in India, T. pilosa to species from the Sunda shelf, and Borneo in particular, T. pentaphylla and T. mor-

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risii to T. papuana from the Tanimbar Island of Aru, and species in Wallacea and the Sunda Shelf, T. odontosperma to species in New Guinea, and T. subvelutina to species from both South and Southeast Asia.

The last treatment for Australia (Telford 1982) pointed out that the taxon-omy of the genus was poorly known, and that Australian and Asian material required additional critical comparison. In addition it raised doubts about the uniqueness of T. holtzei, including an undescribed new taxon, Trichosanthes sp. A, and suggested that T. pentaphylla might be conspecific with T. trifolia (L.) Merr. raised questions on morphological variation.

Trichosanthes sp. A was formally described and typified by Cooper & Ford (2010) as T. odontosperma, and I was consulted regarding the place-ment of the new species in the Trichosanthes phylogeny. It was then also decided to make a new revision of the species in Australia to resolve the remaining questions from Telford’s revision (1982), and to address a number of aberrant collections made in the meantime. It was also decided to base this revision on a molecular phylogeny of Trichosanthes with a broad sam-pling of accession from Australia.

The molecular study is still an ongoing project (to be completed by the summer of 2012), but the preliminary phylogenetic data has been used as a basis for this revision.

Study Trichosanthes is represented by six species in Australia: T. cucumerina, T. morrisii W.E.Cooper sp. nov., T. odontosperma, T. pentaphylla, T. pilosa and T. subvelutina.

Trichosanthes ovigera Blume has recently been synonymised with T. pi-losa (de Wilde & Duyfjes 2009), and T. holtzei is here included within this synonymy. All taxa are illustrated (with the exception of T. odontosperma, which was previously illustrated by Cooper & Ford (2010)), and distin-guished from other Australian species. Notes on habitat and distribution are included together with distribution maps. Three identification keys are pre-sented, two to the sections of Trichosanthes and one to the species of Tri-chosanthes in Australia.

Relevance Molecular phylogenetic studies rely heavily on the identifications of the sampled herbarium vouchers used to create the underlying sequence data. Surprising results in phylogenetic reconstructions always need to be checked for possible contaminations, laboratory mix-ups, or misidentified herbarium vouchers. The latter can be achieved by checking all determinations using revisions of the genus, collaborating with specialists, or by becoming a spe-cialist oneself. In this case the latest revision had become inadequate, and we decided that a new revision was the best way to study the taxonomy and phylogeny of Trichosanthes.

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The future molecular phylogeny will build on the identification of Aus-tralian and New Guinea material made as part of this revision.

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Conclusions

Plant systematics interfaces with other areas of research in myriad ways. The papers presented in this dissertation highlight a few of these interactions, some commonly known, e.g. molecular phylogenetics, biogeography, fossil-calibrated molecular dating, and other less intuitive, i.e. traditional plant use, medicinal plants for women’s health, evolution of traditional knowledge and insect repellents. The interfaces are of course innumerable, but the research presented here all required a solid foundation in systematics as all data is based on correct identification of botanical material. The studies on medicinal plants used for women’s healthcare show that plant use during pregnancy and parturition among the Brou, Saek and Kry is un-common, and informants report avoidance of (traditional-) medicine during pregnancy and parturition (Paper I). A review of medicinal plants for wom-en’s health in Southeast Asia (de Boer, unpublished) finds 165 species-use reports to aid parturition, and this contrast could indicate a specific cultural avoidance. It also shows that a wealth of plants (>70 species) are used for postpartum recovery in steam baths, steam sauna, dietary prescriptions, me-dicinal decoctions and infusions and mother roasting. The traditions of the Brou, Saek and Kry Phong are similar to those observed elsewhere in Lao PDR and Southeast Asia (Paper I). The traditions of the Kry Tae differ in having a complex system of spatial taboos. The mother gives birth outside the village in a specifically built birthing hut, and moves to her menstruation hut in the village only after the first week, and stays there for another period before returning to the parental house with her newborn (Paper II). Chemi-cal analysis of the three most commonly used species in steam sauna, steam bath and mother roasting shows that these species are rich in essential oils, many of which have known bioactivity. The mode of application in hotbed and mother roasting differs from the documented mechanisms of action, and plants are likely to serve mainly hygienic purposes in these practices, by segregating the mother from infection sources. Inhalation of essential oil vapors in steam sauna can possibly have medicinal efficacy, but is unlikely to alleviate the ailments commonly encountered during postpartum conva-lescence. Dermal condensation of essential oils during steam sauna, and cleansing of the perineal area with an infusion rich in essential oils during steam baths, are possibly efficacious uses of the reported medicinal plants (Paper III).

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The studies on medicinal plant use by the Brou, Saek and Kry ethnic groups show that medicinal plant use is dissimilar even between ethnic groups living alongside each other in the same area for many generations. We posit that their medicinal floras are constituted partly of species with empirically tested efficacy, partly of new remedies under evaluation, and partly of species used due to cultural significance. The empirically tested remedies are those that are most likely to be transmitted horizontally be-tween cultures, and reports on the use of the species can be found in scien-tific literature concerning Southeast Asia. We also posit that this dissimilari-ty is not caused by ancient traditional knowledge that predates their coexist-ence in the area, but by a constant innovation of traditional medicine to find cures for afflictions affecting these people. (Paper IV). The studies on traditional plant repellents against blood-feeding insects, ticks, mites, and leeches have shown that close to a hundred different plant species are used in rural Lao PDR. A key objective of these studies was to find inexpensive and accessible alternatives to synthetic repellents and pesti-cides for the impregnation of bed nets, but also as topical repellents, to be used on the skin or on clothes, against mosquitoes, ticks and leeches. The studies showed that a large potential for botanical alternatives exists (Paper V), that some species have good efficacy in laboratory trials (Paper VI), and good efficacy in field trials compared to synthetic repellents (Vongsombath et al. 2011; Vongsombath 2011). The molecular phylogeny of Trichosanthes (Cucurbitaceae) reveals that Tri-chosanthes forms a well-supported monophyletic clade only if Gym-nopetalum is included. Fossil-calibrated molecular dating of the phylogeny reveals an early Oligocene origin of the genus, with late Oligocene estimates for the stem nodes of the main clades, but with the remaining extant sections originating and diversifying during the Miocene. Biogeographical analysis shows a likely East or South Asian origin of Trichosanthes, with lineages diversifying and spreading throughout Australasia from the early Pliocene to the Pleistocene. Five lineages of Trichosanthes are found in Australasia, and at least three of these are the result of independent dispersal events from Asia (Paper VII). Taxonomical implications from the molecular phylogeny are placing the former Gymnopetalum taxa in Trichosanthes using either existing names or new combinations. To maintain the current infrageneric classification a new section is described, and a synopsis of all species is giv-en to clarify which sections include what species (Paper VIII). The revision of the Australian species of Trichosanthes addresses unresolved questions from an earlier revision, and studies ambiguous collections made in the mean time (Paper IX). In addition it serves as a basis for a further molecular phylogeny of the Australian taxa and their closest relatives.

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A research project with traditional plant use in Lao PDR as its focus can take different courses. This research diverged into three coherent directions, all of which were pursued with fervor and determination. This divergence is re-flected in my research outside this dissertation, and during the last 6 years I have pursued many other interesting leads, some successfully, others less so. In the future I intend to shift from broad to less broad, aim to increase quali-ty and impact, innovate to the edge and beyond, while at the same time keep-ing an open mind to student initiatives.

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Research ethics

In the years since the 1992 ratification of the Convention on Biological Di-versity (CBD), international research on plant genetic resources has become more regulated to protect the intellectual property rights of the stewards of traditional knowledge, and to ensure equitable access and benefit sharing of national germplasm resources.

CBD Article 8j in particular is relevant to ethnobotanical research: “Each contracting Party shall, as far as possible and as appropriate, subject to na-tional legislation, respect, preserve and maintain knowledge, innovations and practices of indigenous and local communities embodying traditional life-styles relevant for the conservation and sustainable use of biological diversi-ty and promote their wider application with the approval and involvement of the holders of such knowledge, innovations and practices and encourage the equitable sharing of the benefits arising from the utilization of such knowledge innovations and practices.”

The CBD, but also the rapid developments in the scientific fields of mo-lecular biology and genetics, served as a wake-up call to developed and de-veloping countries to the dangers of bioprospecting and biopiracy. DNA from herbarium vouchers could be extracted for the development of new crops, and anthropologists interested in traditional knowledge were suddenly backed by pharmaceutical conglomerates mass-screening medicinal plants for new pharmaceuticals. Prior to the CBD many countries already had strict regulations on research permission and material transfer to protect their bio-logical resources, and post CBD regulations still vary vastly from country to country, but awareness was raised and terms like Access and Benefit Sharing and biopiracy made it into the vocabulary of scientists.

The caveat, “subject to national legislation”, in the formulation of Article 8j makes it difficult for the “indigenous and local communities embodying traditional lifestyles” to claim any intellectual property rights over the knowledge if a country does not recognize their identity or autonomous rights. Countries like Indonesia and Lao PDR, that consist of many ethnic groups, define all people as their population, making it difficult to recognize culture-specific IPR and resulting benefits. However as a result of the 2010 Nagoya protocol on Access and Benefit Sharing, discussions are now un-derway to define mechanisms for compliance and noncompliance to Prior Informed Consent and Mutually Agreed Terms that will protect indigenous IPR.

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The research presented in this dissertation was part of a larger project includ-ing three PhD students; two from Lao PDR, Chanda Vongsombath (CV) and Vichith Lamxay (VL), funded by the Swedish International Development Cooperation Agency, Department for Research Cooperation (SIDA-SAREC) through its bilateral program with Laos, and one from Sweden, Hugo de Boer (HdB), funded by SIDA’s general research funding. The collaboration was formalized by a Memorandum of Understanding (MoU) between Uppsala University (UU) and the National University of Laos (NUoL), Letter of Intent (LoI) between involved partners, and a Material Transfer Agreement (MTA), including a chapter on Access and Benefit Sharing (ABS), also between UU and NUoL. The vice-chancellor of NUoL, under direction of the Ministry of Education of Lao PDR, granted research permis-sion for this project to CV, VL, and HdB.

All field research for Papers I-VI was discussed and planned by HdB, VL, and CV. Interviews and field trials for Papers I-VI, were carried out by, or under supervision of the Lao PhD students, CV and VL. Interviews for Papers I-IV were done by VL, or by Lao students from NUoL under super-vision of VL. Research permission to conduct research in the Nakai-Nam Theun NBCA was granted to VL by the Nakai-Nam Theun Watershed Man-agement and Protection Authority (WMPA) and the Nakai District Govern-ment Office. Research in the Nakai-Nam Theun NBCA area was carried out by a team led by VL, including a representative from the Nakai District Government Office, the Department of Agriculture and Foresty, the Nakai Public Health Office, the Nakai-Nam Theun WMPA, and Biology students from NUoL. Each village visit was initiated by a meeting with the village head and local party representative to introduce the project, show the re-search permission, and obtain Prior Informed Consent (PIC). Each village interview was introduced by VL and the village head to obtain Prior In-formed Consent (PIC), and all informants participated voluntarily. Plant material collected for herbarium vouchers was collected and pressed in the field at transferred to the herbarium of the Department of Biology, Faculty of Sciences, NUoL.

Interviews, hydro-distillation, field trials and laboratory experiments for Papers V-VI, were done by CV, or by Lao students from NUoL under su-pervision of CV. Prior to interviews for Paper V the District Government Office of each district would be contacted and visited to introduce the re-search, show the research permit, and obtain permission to conduct research in that district. Prior to interviews in villages in NBCAs the NBCA office would be visited to obtain research permission. In many cases District Gov-ernment Office or NBCA staff would be assigned to accompany the inter-views. Prior Informed Consent was obtained from each village head and each informant before conducting the interviews. Plant material collected for herbarium vouchers was collected and pressed in the field at transferred to the herbarium of the Department of Biology, Faculty of Sciences, NUoL.

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Plant material for DNA extraction and herbarium vouchers used for Pa-pers VII-IX, were obtained through loans from herbaria.

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Svensk sammanfattning

Traditionell användning av naturprodukter, speciellt växter som kan använ-das till läkemedel eller insektsrepellerande/insektsdödande medel, är mycket vanligt förekommande i Sydostasien. Denna avhandling baseras på omfat-tande undersökningar rörande traditionell användning av växter i Laos. Stu-dierna kan delas in i tre delprojekt: (i) Användning av medicinalväxter hos de etniska grupperna Brou, Saek och Kry; (ii) Användning av växter som repellenter och pesticider mot parasitära leddjur och blodiglar; samt (iii) Ormgurkssläktets (Trichosanthes,Cucurbitaceae), fylogeni och biogeografi; detta släkte hyser många medicinskt betydelsefulla arter.

Få människor på den laotiska landsbygden har tillgång till moderna läke-medel. Därför är traditionell användning av växter och andra naturprodukter här vanligt förekommande inom primärhälsovård och sjukvård. Relativt mycket är känt om hur medicinalväxter kan användas för behandling av olika sjukdomar. Men vilka växter som kan användas och hur de bör använ-das för att främja kvinnors hälsa, särskilt efter barnafödande, är relativt då-ligt undersökt.

Vektoröverförda sjukdomar som till exempel malaria, denguefeber och japansk encefalit är vanliga på den laotiska landsbygden. Syntetiska insekts-repellerande kemiska ämnen har, via biståndprojekt, under senare år börjat användas i ökande omfattning. Problem med resistens mot de få kemikalier som kan användas, är emellertid hela tiden ett reellt hot. Naturliga, växtpro-ducerade myggrepellerande ämnen, som människor ganska enkelt skulle kunna ta fram själva, vore ett bra alternativt, speciellt om de växtarter som har sådana ämnen kunde identifieras.

Ormgurkorna omfattar 90-100 arter och är det största släktet i gurkväxt-familjen. Många av arterna används traditionellt som läkeväxter. Eftersom de flesta arterna blommar på natten och är tvåbyggare, dvs. med han- och honblommor på skilda plantor, är släktet dåligt känt och herbariernas in-samlingar är ofta ofullständiga. En bättre kunskap om arternas inbördes re-lationer skulle kunna ge bättre vägledning om vilka grupper (klader) som hyser medicinalväxter. Dessa borde då mer ingående undersökas biokemiskt och farmakologiskt, men också med avseende på evolutionsbiologi och bio-geografi.

I projektet om medicinalväxter använde jag strukturerade intervjuer, studier av byarnas traditioner, botaniska vetenskapliga samlingar, samt ångdestillat-

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ion av växtmaterial för att erhålla essentiella oljor som analyserades kemiskt med gaskromatograf kopplad till masspektrometer (GC-MS-analyser). I by-arna Brou, Saek och Kry i Annamitebergen intervjuade vi invånarna om sin kultur, om eventuella seder i samband med barnafödandet, och om vilka medicinalväxter de använder eller har kunskap om. Under vandringar med informanter i byarnas omgivningar insamlade vi de växter som hade om-nämnts i intervjuerna. En del växter preparerades och artbestämdes och in-förlivades med samlingarna i Evolutionsmuseet’s Fytoteket i Uppsala, Royal Botanic Gardens i Edinburgh, Faculty of Sciences herbarium vid National University of Laos i Vientiane, och Nationaal Herbarium Nederland i Lei-den. Några arter inom släktet Amomum (Zingiberaceae), som är betydelse-fulla medicinalväxter och används i samband med barnafödande, valdes ut för kemiska analyser. Växtmaterial ångdestillerades för att få fram de essen-tiella oljorna, som analyserades med GC-MS.

I delprojektet om traditionella växtrepellenter använde jag som forsk-ningsmetoder strukturerade intervjuer, litteraturstudier, studium av botaniska samlingar, och laboratorieexperiment. Informanter i 66 byar i Laos intervju-ades om vilka växter de använder som repellenter. De rapporterade arterna insamlades och artbestämdes, varefter en litteraturstudie gjordes med avse-ende på publicerade data om de olika växtarternas eventuella leddjursdö-dande och/eller leddjursavvisande effekter. Några växter uppgavs ha effekt mot fluglarver, som infesterar fisk som fermenteras. Dessa växter undersök-tes mer noggrant med hänsyn till sina potentiella fluglarvsavvisande effekter.

I projektet om ormgurkorna användes följande forskningsmetoder: morfo-logiska studier av herbariematerial, molekylärbiologiskt laboratoriearbete, fossilkalibrerad åldersbestämning, samt fylogenetiska och biogeografiska analyser. Material för DNA-extraktion samlades in från 60 arter. Dessa om-fattar representanter för samtliga sektioner inom släktena Trichosanthes, Gymnopetalum och Hodgsonia. Olika markörer från kärn- och plastid-DNA sekvenserades och användes för den molekylär-fylogenetiska analysen. Fylogenin daterades med hjälp av dateringsprogram och användes vid den biogeografiska analysen.

Informanter i byarna Brou, Saek och Kry rapporterade att mer än 75 växtar-ter används för kvinnors hälsovård, speciellt vid ångbastu, ångbad, ’heta bädden’ (eng.: hot bed) och ’moderstekning’ (eng.: mother roasting); till dekokter och téer av läkeväxter; och till en speciell diet att intas efter barna-födandet. För nämnda folkgrupper och många andra folk i Laos är använd-ningen av läkeväxter i samband med barnafödandet och för moderns och barnets återhämtning under tiden därefter viktiga. Dessa traditioner bör in-förlivas med den vedertagna västerländskt inspirerade hälsovården, speciellt den som syftar till att minska mödra- och barnadödligheten. På så sätt skulle hälsovårdsreformerna kunna få ett snabbare och större genomslag.

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Dokumentationen av olika användningsområden för de vanligaste läke-växterna visar att likheten i bruk mellan de studerade etniska grupperna är relativt låg. Detta är anmärkningsvärt med tanke på att dessa folkgrupper under många generationer har levt i samma område och använt samma resur-ser från naturen. Anledningen är förmodligen att bristen på effektiva läke-medel har lett till en process med ständiga innovationer, där endast de mest effektiva botemedlen delats mellan folken.

Intervjuerna visade att närmare 100 olika växtarter används mot myggor, fluglarver, kvalster, fästingar, löss och blodiglar. Många av dessa växter innehåller kemiska ämnen som redan vetenskapligt har visats ha insektsdö-dande och/eller insektsrepellerande effekter. Det är sannolikt att kommande studier av dessa växter kan leda till billigare och effektivare alternativ till de dyra syntetiska kemiska repellenter och pesticider, som nu finns på mark-naden som skydd mot framförallt blodsugande insekter och fästingar.

Den fylogenetiska studien av ormgurkorna visade att Trichosanthes är pa-rafyletiskt, men att det tillsammans med Gymnopetalum bildar en monofyle-tisk grupp. Dateringen av Trichosanthes-fylogenin visar att släktet har sitt ursprung under tidig Oligocen, samt att många av de existerande sektionerna diversifierades under Miocen. Den biogeografiska analysen tyder på att släk-tet sannolikt har sitt ursprung i Syd- eller Sydostasien, och att minst fem kolonisationshändelser har skett i Australasien. De fylogenetiska resultaten innebär att Gymnopetalum-arterna bör inkorporeras i släktet Trichosanthes. Detta görs med hjälp av tillgängliga namn eller nya kombinationer. En sam-manställning av Trichosanthes med de nya kombinationerna och en revision av arterna i Australien presenteras i två publikationer.

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Acknowledgments

Many people have contributed to the work presented in this dissertation, and no doubt I will forget to mention and thank a few of those. Any people that I have forgotten should know that it was not intentional, but more likely be-cause nothing on the desk in front me elicited a cue to their name or person.

Making an acknowledgments section is a challenge. Placing the people into groups, and subsequently ranking them within the group is a feasible ap-proach. Paraphyly and reticulate relations does make it a bit challenging to place people, especially if one is stuck with the idea that each person should be mentioned only once. Everyone’s contribution is of course unique, and ranking those contributions is unfair and inaccurate.

Below everyone and all sources of funds are listed alphabetically, with a note on their role in this research: Abderrahim Ouarghidi (species complex, the Berber view on life, a solid foundation), Afsaneh Ahmadzadeh (social organizer, cooking recipes), Ag-neta Brandtberg Falkman (making the impossible happen, almost any publi-cation on my desk), Allison Perrigo (systematics PhD student, donkey occu-pier, the bane of Bente’s wrath), Alvaro Martinez Barrio (bioinformatician, Oukaimeden skier, Johnny Walker Green label), Anders Backlund (haggis chef, socialite, supportive and understanding), Anders Larsson (aka profes-sor Gurka, office hair-loss, dog biscuit addict, nearly as enthusiastic about phylogenies as Per E. and hybrids as Magnus P.), Andreas Wallberg (former systematics PhD student, real local, geek tips), Anja Rautenberg (former systematics PhD student, vintage dishwasher, Russian connection), Anna Maria Lundins stipendiefond, Anna-Karin Borg-Karlsson (examination committee member, GC-MS facilities for, Chanda’s PhD project), Anneleen Kool (systematics PhD student, intrepid explorer, terrible gardener, my wife, and mother of my dear children, just-in-time research, teaching fervor, time optimist, last remnant of the old guard), Annika Vinnersten (former system-atics PhD student, a warm-hearted welcome to the department, a predilection for Scotch whiskey), Åsa Krüger (former MSc student, patiently putting up with a couple of PhD students and their newborn baby, massive labwork), Åsa Kruys (UU systematist, meticulous, kind), Axel Dalberg Poulsen (Axel from the big garden with the waterfall in Oslo, a predilection for Swedish

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cakes, Vichith’s PhD project), Bengt Oxelman (Former department head, warm-hearted welcome to systematics in Uppsala, support for various appli-cations and activities, Scotch whiskey aficionado), Bente de Boer (daughter, showing me that there is more in life than research, I hope you enjoy reading this later!), Bernice Kay Zerger (taking care of Floris in Berkeley), Björn-Axel Beier (former systematics PhD student, thank you for your pipettes!), Bozo Frajman (former systematics PhD student, workaholic, Bozi), Brent Mishler (UC Berkeley moss systematist, institutional host during my ex-change at Berkeley), Brian Su (interpreter and research collaborator at KUN and during fieldwork with the Red-headed Yao in Jinping county), Brigitta Duyfjes (Leiden systematist, morphologist, always available for Trichosan-thes questions, source of most of my Trichosanthes samples), Bruce Baldwin (UC Berkeley systematist, department host during my exchange at Berkeley, interested, always working), Cajsa Anderson (former systematics PhD stu-dent, swearing in the Göteborg dialect, strong opinions, a crappy car), Cam-eron Williams (tree climber, positive, Rikke’s partner), Carolin Frank (par-enting inspiration, gourmet, displaced), Catarina Ekenäs (former systematics PhD student, ethnobotanist, in-between), Catarina Rydin (former UU sys-tematist, contagious enthusiasm for science), Chanda Vongsombath (PhD researcher partner, research on Papers V and VI, fieldwork, copious quanti-ties of Lao Lao, thank you for a great project together!), Chengjie Fu (sys-tematics postdoc, Chinese delectables aficionado), Christian Vogl (BDLN and Erasmus network, wine, hefeweizen, local Austrian food), Christina Wedén (former systematics PhD student, truffles, a house for our little red car, and loan of a car to pick up our Moroccan friends from Arlanda on an ill-fated adventure to Eka), Chuck Davis (Halloween, Hancock St, and my host at HUH), Cindy Looy (candid, treehut-dweller, my house-fix-fix), Colin Ridsdale (straightforward language, a love of Beer Lao, Rubiaceae dets), Cory Zigler (acquaintance to baseball and Budweiser, beer brewer, father-to-be), Curators of many different herbaria, Dave Ardell (academic inspiration, California eye-opener, parenting inspiration, cocktail inspiration, and play-ing a key role in me being here defending my dissertation today!), Dave Schmalz (father, reviewer, copy-editor, Backen electrician and builder), Da-vid Soffa (like-minded, hoarder, 2926 Otis St architect), Dick Andersson (former systematics PhD student, founder of a foster-plant-plan garden somewhere), Dien de Boer (mother, role model in many ways, Dirk’s baby sitter), Ding He (systematics PhD student, acclaimed board member, a.k.a. Chairman Ding), Dirk de Boer (son, showing me that there is more in life than research, hope you enjoy reading this later!), Dorothée Vallot (French embassy in Sweden scientific liaisons officer), Douglas Stone (former UU systematics postdoc, matured California beach-boy turned hard-working South African), Elisabeth Långström (former systematics PhD student, my self-appointed, much appreciated, PR manager), Elsbeth Scholtes (unrelent-ing help with student registration, always available for a chat), Emma Lundh

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(former MSc student, hard working, slightly overdue, and finally complet-ing), Erin Smith (GDF colleague, patient administrator and organized be-yond my capacity), EU-SYNTHESYS grant to study at GB-TAF Royal Bo-tanic Gardens Edinburgh, Evan Mati (former MSc student, megalomaniac, colloquial, connected), Floris de Boer (son, showing me that there is more in life than research, one of the few people who actually figured out how I spend my days at the Department), Frank Lindh (SLU Chemist, supportive and encouraging from our first meeting), Frida Eggens (former systematics PhD student, Swedish flat-bread baker, Backen pizza eater), Gary Martin (Global Diversity Foundation architect, passionate ethnobiologist and lectur-er, research colleague, a super introduction to many research partners and colleagues, and forever beat me taking longer to complete a PhD, but always a few countries behind), Gijs van der Schot (Not Sinterklaas), Gity Behrevan (support at Sida-SAREC), Gloria Gallardo (CEFO coordinator, CSD, the interdisciplinary approach, research network), Håkan Svensson (patiently scanning my Lao colleagues’ computers, both willing and reluctant, but al-ways), Hanno Schaefer (Cucurbitaceae systematist, great research collabora-tor, and bird-watching, coffee-drinking, vegan, all-season-cyclist, workahol-ic), Hans Ellegren (zealous, supportive, connected), Hans-Henrik Fuxelius (positive, crazy, Floris’ first dissertation), Heather McMillen (SEB student rep, unrecognized pioneer market ethnobotanist), Hedi Peterson (bioinfor-matician, True Estonian, Juniper twigs and mesi, Belorussian Olympic ath-lete, manager), Hege Vårdal (former systematics PhD student, one of the few Norwegians I understand, enthusiastic), Heidi Solstad (poppies, old profes-sors, Akureyri), Helge Ax:son Johnsons stiftelse, Henk Beentje (helpful, innovative, good company), Henrik Lantz (former systematics PhD student, busy, activity juggler), Henry Bass (knowledgeable, Belmont, cheese con-noisseur), Inga Hedberg (UU systematist, mother of ethnobotany at UU, thank you for welcoming Anneleen and me to the department in 2001), Jan Wieringa (Brahms helpdesk, Trichosanthes records, Christmas parties, dishwashers, trailer hitches, pond and pool swimmer), Jennifer Sowerwine (Butler Creek, Southeast Asian minorities, and many overlaps in research interests), Jenny Smedmark (SU systematist, pre-dissertation-party-parties, proud owner of a cat), Joel Altstein (Super Cambridge host, creative house renovator, and forgiving car lender), Johan Nylander (hard-working beyond healthy, perfectionist, avid evolutionary biology theorist), Johannes Lundberg (proving the existence of Gnarp, Japanese Trichosanthes samples, speleophilia), John Petterson (stipendmeister, papa, searching for novel vi-ruses to improve the world), Jos Käfer (impromptu research collaborator, statistics and R wiz, ethnobotanist with a sole focus on alcoholic drinks, Paronychia spotter, mountain goat, and emergency baby-sitter), Jos van der Maesen (WUR systematist, WUR supervisor for my research in Tanzania, identifications of Lao legumes), Karin Olsson (financial administrator, who probably saved my project from bankruptcy many times over), Karolina

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Larsson (former systematics PhD student, worm-woman, grateful vet con-nection), Katarina Andreasen (examination committee member, UC Berke-ley connection, intro card to Bridget’s kindness), Katinka Pålsson (feliphile, fieldwork with a rolling suitcase, bird-head soup), Kristina Articus (former systematics PhD student, the Finnish nation, super supportive), Lars Björk (allround farmacognosist, chemist, innovator and inventor, supervisor, re-search partner, potato, beer and bridge donations, thank you for being a great supervisor!), Lars Hiertas minnesfond, Lenny Kouwenberg (bacon and blue cheese, microbreweries, and cats), Liljewalchs resestipendium, Lily Altstein (lead-free mom, former Jesuit fraternity landlady), Madeleine Julin (former student, adventurist, henna artist, bold Sahara Saab expeditionist), Magnus Lidén (UU systematist, adventurist, researcher of plants in remote areas, significant source of inbox clutter), Magnus Popp (pre-dissertation-party-parties, convergent offspring evolution), Marc Sosef (WUR systematist, the ghost of John, jointly defeated by a tire), Marcin Kierczak (Parties at Back-en, Erken, Åland, visiting Zyrardow, skiing in Oukaimeden, teaching in Marrakech, the North Star Bay incident of 1968, the Tungaska blast of 1908, and many more), Marcus Lindborg (former student, adventurist, beer brewer, bold Sahara Saab explorer), Maria Backlund (former systematics PhD stu-dent, supportive, whiskey and Scottish culinary connoisseur), Maria Lindström (systematics PhD student, muay thai connection, positive), Maria Romeralo (UU systematist, transformed slime molds into tangible matter, Spanish clock), Mark Newman (E systematist, Amomum specialist, Vichith’s co-supervisor, Edinburgh host, and well-known Gaelic TV-botanist), Matil-da Segersäll (former MFS student, determined, patient), Mats Hjertson (bo-tanical collections, many loan requests, help transferring collection data to the museum’s clunky database), Mats Thulin (UU systematist, co-supervisor, taxonomy and nomenclature master, thank you for being a great supervi-sor!), Matt Guilliams (Berkeley systematist, Marah fossil, picnics at Willard Park), Mattias Forshage (former systematics PhD student, I have now passed the old-guard torch to Anneleen), Michael de Boer (little brother, interpreter and research collaborator at KUN and during fieldwork with the Red-headed Yao in Jinping county), Mohamed Elhaouzi (GDF Morocco, probably the kindest person in the world), Mohamed Farah (Uppsala Monitoring Centre connection, medicinal plants synonymy research), Nahid Heidari (laboratory technician, and so much more), Nan Laird (the kindest and most hospitable statistics professor I have ever met), Nat Bletter (SEB student rep, ethnobo-tanist and Madre chocolate flavormeister), Nick Enfield (Kry linguistics and culture, ethnoclassification, and coffee in Nijmegen), Omar Fiz Palacios (UU systematist, afflicted with parentitis, Spanish clock), Paul Kessler (bo-tanical collections, identification of material, Lao research gossip), Peer-reviewers (some 20 kind researchers that spent their precious time reviewing my papers), Pelle Lindström (introduced the term ‘man castle’ into my vo-cabulary, and always in for another beer), Per Alström (former UU systema-

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tist, great public outreach, Southeast Asian warblers), Per Erixon (former systematics PhD student, positively selected whiskey guru, thanks for using your Bayes block!), Per Kornhall (former systematics PhD student, antithe-ist, interested, supportive), Petra Korall (UU systematist, unwilling insomni-ac, combining excellent research with a large family), Raj Puri (BDLN and Erasmus partner, GDF colleague, Anthropac support), Regnells botaniska resestipendium, Renee Ankarfjärd (support at Sida-SAREC), Rick Stepp (GDF colleague, inspiration source), Rikke Reese Naesborg (former system-atics PhD student, barefoot lichenologist, happily married in California), Robert-Jan den Tex & Kathelijne Balfoort (ambitious, slightly anachronistic ornithologist, never figured out the location of the EBC parking lot, absorbed roughly 10% of my EBC research time drinking our department’s coffee, great company), Ron Meyboom (UMC connection, herbal pharmacovigi-lance, a justified wariness of herbal medicine, but a superstitious belief in the health benefits of alcohol beverages), Rota Razafimandimbison (Warm-hearted welcome to Malagasy life, another house-fix-fix), Samira Garboui (former systematics PhD student, wonderful family, help finding accommo-dation), Sandra Baldauf (UU systematist, head of Systematic Biology group, sudoku and popcorn, thank you for coming to Uppsala, and being a support-ive, sometimes frustrated, but successful group leader), Sanja Savic (former systematics PhD student, Uppsala systematist, workaholic, upplandsmål), Selena Ahmed (my twin sister, Kunming connection, former owner of a Jane Austen copy of Pride and Prejudice currently in my home), Sernanders stif-telse, Sida-SAREC grant SWE-2005-338, Sofie Joosse (fixing-fix, exchang-ing real estate, a chaotic holiday to Tenerife, frituurpan time-share, CEFO activities), Solveig Freudenthal (support at Sida-SAREC), Stefan Ås (pa-tiently scanning my Lao colleagues’ computers, tips on maintenance of wooden boats), Sten Olsson (Uppsala Monitoring Centre connection, medic-inal plants synonymy research), Stiftelsen Hierta-Retzius fond, Stina Bolinder (former systematics student, current systematics PhD student at SU, a.k.a. Fake-Stina, Grek-Stina and Second-Stina, definitely a bad conscience), Stina Weststrand (systematics PhD student, social organizer, little-known retailer of Upplands Flora), Sue Bass (knowledgeable, organized, Belmont, traveler), Sunniva Aagaard (former systematics PhD student, slightly more pessimistic than myself, karsk, Trøndelag hospitality), Susanne Renner (Cu-curbitaceae systematist, inspiring research collaborator), Sven och Dagmar Saléns stipendiefond, Swedish Research Links, Sylvain Razafimandimbison (examination committee member, Rubiaceae specialist, Malagasy connec-tion, thank you for arranging a baby sitter for Bente during the AETFAT meeting), Thijs Ettema & Carolien Dirks (House buying, Lavsjön picnics, Lekstuga moving), Thomas Jaenson (the medical entomologist, co-supervisor, concerned, but reassuring main supervisor of Chanda, thank you for being a great supervisor!), Thomas Norberg (SLU Chemistry professor, chemist, supportive and encouraging from our first meeting, honorary mem-

50

ber of Gysinge Talibanerna, a shared love-hate relationship with window renovation), Tom Carlson (UC Berkeley host, Butler Creek, the annual Salmon run BBQ in San Francisco), Torsten Eriksson (SU systematist, pre-dissertation-party-parties, proud owner of a cat), Uppsala University – Uni-versity of California at Berkeley exchange fellowship, Vichith Lamxay (former systematics PhD student, collaborator on ethnobotany in Laos, co-author on Paper I, II, III and IV, Lao sushi, weeks and weeks of fieldwork, socials at the Lao embassy, Lao-lao in Ban Buuk, thank you for a great pro-ject together!), Wallenbergsmedel till Rektors förfogande, Wendy Cooper (Trichosanthes collaborator, co-author on Paper IX, 281 emails received and 145 sent, source of 95% of Australian material), Wijnand Boonstra (fixing-fix, exchanging real estate, a chaotic holiday to Tenerife, frituurpan time-share, finished his PhD ‘essay’ a few years earlier), Willem de Wilde (Lei-den systematist, morphologist, always available for Trichosanthes questions, source of most of my Trichosanthes samples), William Robichaud (Nakai-Nam Theun WMPA, great guy and good company, thank you for your emergency help during the summer of 2008!), Yonas Isaak Tekle (former systematics PhD student, discussions on the toughness of chicken meat).

51

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