Timothy J. Motley

Complex origins of breadfruit (Artocarpus altilis, Moraceae): implications for human migrations in Oceania.

Breadfruit (Artocarpus altilis, Moraceae), a traditional starch crop in Oceania, has enjoyed legendary status ever since its role in the infamous mutiny aboard the H.M.S. Bounty in 1789, yet its origins remain unclear. Breadfruits closest relatives are A. camansi and A. mariannensis. DNA fingerprinting data (AFLP, amplified fragment length polymorphisms) from over 200 breadfruit cultivars, 30 A. camansi, and 24 A. mariannensis individuals were used to investigate the relationships among these species. Multivariate analyses and the identification of species-specific AFLP markers indicate at least two origins of breadfruit. Most Melanesian and Polynesian cultivars appear to have arisen over generations of vegetative propagation and selection from A. camansi. In contrast, most Micronesian breadfruit cultivars appear to be the result of hybridization between A. camansi-derived breadfruit and A. mariannensis. Because breadfruit depends on humans for dispersal, the data were compared to theories on the human colonization of Oceania. The results agree with the well-supported theory that humans settled Polynesia via Melanesia. Additionally, a long-distance migration from eastern Melanesia into Micronesia is supported.

Phylogenetics of the genus Scaevola (Goodeniaceae): implication for dispersal patterns across the Pacific Basin and colonization of the Hawaiian Islands

Scaevola, the only genus of Goodeniaceae that has extensively radiated outside of Australia, has dispersed throughout the Pacific Basin, with a few species reaching the tropical coastal areas of the Atlantic and Indian Oceans. Five Australian and most of the non-Australian species are placed in Scaevola section Scaevola based on their fleshy fruits, indeterminate inflorescences, and more arborescent habits. Analyses of ITS sequence data demonstrate that Scaevola is a monophyletic group if S. collaris is excluded and Diaspasis filifolia is included. The genus is Australian in origin, but there have been at least six separate dispersal events from Australia. Four of these dispersals each resulted in single extra-Australian species. The remaining two were followed by radiations that gave rise to large groups, each including one of the widespread strand species, S. taccada and S. plumieri. Remarkably, three of the six dispersals established species on the remote Hawaiian Archipelago, representing at present the largest number of colonizations by any flowering plant genus to these islands.

Molecular systematics of the Catesbaeeae-Chiococceae complex (Rubiaceae): Flower and fruit evolution and biogeographic implications

The classification of the Catesbaeeae and Chiococceae tribes, along with that of the entire Rubiaceae, has long been debated. The Catesbaeeae-Chiococceae complex (CCC) includes approximately 28 genera and 190 species primarily concentrated in the Greater Antilles (nearly 70% of the species), Central and South America, and in the western Pacific (three genera). Previous molecular studies, with broad sampling of the Rubiaceae, have shown the CCC to be a monophyletic group. The present study is a more detailed examination of the generic relationships within the CCC using two data sets, the nuclear ribosomal ITS regions and the trnL-F chloroplast intron and spacer. Maximum parsimony analyses lend further support to the previous hypotheses that the CCC is monophyletic and sister to Strumpfia maritima. However, within the complex several genera do not form monophyletic groups. Previous studies of the Rubiaceae suggest that the ancestral fruit type in the CCC is a multiseeded capsule. Indehiscent, fleshy fruits appear to have evolved three to four times within this lineage. Changes in floral morphologies within the complex tend to correspond to cladogenesis among and within genera. Finally, molecular analyses suggest one or possibly two long-distance dispersals from the Americas to the western Pacific.

Phylogeny of the herbaceous tribe Spermacoceae (Rubiaceae) based on plastid DNA data

Abstract In its current circumscription, the herbaceous tribe Spermacoceae s.l. (Rubiaceae, Rubioideae) unites the former tribes Spermacoceae s. str., Manettieae, and the Hedyotis–Oldenlandia group. Within Spermacoceae, and particularly within the Hedyotis–Oldenlandia group, the generic delimitations are problematic. Up until now, molecular studies have focused on specific taxonomic problems within the tribe. This study is the first to address phylogenetic relationships within Spermacoceae from a tribal perspective. Sequences of three plastid markers (atpB-rbcL, rps16, and trnL-trnF) were analyzed separately as well as combined using parsimony and Bayesian approaches. Our results support the expanded tribe Spermacoceae as monophyletic. The former tribe Spermacoceae s. str. forms a monophyletic clade nested within the Hedyotis–Oldenlandia group. Several genera formerly recognized within the Hedyotis–Oldenlandia group are supported as monophyletic (Amphiasma Bremek., Arcytophyllum Willd. ex Schult. & Schult. f., Dentella J. R. Forst. & G. Forst., Kadua Cham. & Schltdl., and Phylohydrax Puff), while others appear to be paraphyletic (e.g., Agathisanthemum Klotzsch), biphyletic (Kohautia Cham. & Schltdl.), or polyphyletic (Hedyotis L. and Oldenlandia L. sensu Bremekamp). Morphological investigations of the taxa are ongoing in order to find support for the many new clades and relationships detected. This study provides a phylogenetic hypothesis with broad sampling across the major lineages of Spermacoceae that can be used to guide future species-level and generic studies.

USING AMPLIFIED FRAGMENT LENGTH POLYMORPHISMS (AFLP) TO IDENTIFY BLACK COHOSH (ACTAEA RACEMOSA) 1

The rhizome ofActaea racemosa L., commonly called black cohosh, is a popular botanical dietary supplement used to treat female health concerns. The rhizomes used in black cohosh products are often collected from the wild. To ensure quality control, it is imperative that plants be correctly identified. This paper examines the use of the DNA fingerprinting technique, AFLP, as an analytical means of identifyingA. racemosa from three other closely related sympatric species. To this end, 262 AFLP markers were generated, and one unique fingerprint was identified forA. racemosa, whereas two, six, and eight unique fingerprints were identified for the closely related speciesA. pachypoda, A. cordifolia, andA. podocarpa, respectively. Two commercial black cohosh products were also subjected to AFLP analysis and shown to contain onlyA. racemosa. The results of this study suggest that AFLP analysis may offer a useful method for quality control in the botanical dietary supplements industry.ResumenDas Rhizom vonActaea racemosa L., allgemein als ‘black cohosh’ bezeichnet, ist eine beliebte pflanzliche Diätsergänzung, die für weibliche Gesundheitsprobleme benützt wird. Oft sind die in ‘black cohosh’-Produkten verwendeten Rhizome in freier Natur gesammelt. Um Qualitätskontrolle zu sichern, ist es zwingend, die Pflanzen richtig zu identifizieren. Diese Studie überprüft den Gebrauch der DNA-Fingerabdrucktechnik, AFLP, als analytisches Mittel der Identifizierung, umA. racemosa von drei anderen in ihrer Nähe beheimateten und nah verwandten Spezies zuunterscheiden. Zu diesem Zweck wurden 262 AFLP-Fingerabdrücke erzeugt. Für A. racemosa wurde ein einzigartiger Fingerabdruck identifiziert, während für die nah verwandten Spezies A. pachypoda zwei, A. cordifolia sechs, und A. podocarpa acht einzigartige Fingerabdrücke gefunden wurden. Zwei kommerzielle ‘black cohosh’ -Produkte wurden ebenfalls der AFLP-Analyse unterzogen, wobei nur A. racemosa nachgewiesen werden konnte. Die Resultate dieser Studie zeigen, daβ die AFLP-Technik eine nützliche Methode für die Qualitdtskontrolle in der pflanzlichen Diätsergänzungsindustrie bieten kann.

Systematics and Species Limits of Breadfruit (Artocarpus, Moraceae)

Abstract Breadfruit (Artocarpus, Moraceae) is an important staple in Oceania and throughout much of the tropics. Interpretations of species delimitations among breadfruit and its closest relatives have varied from recognition of one to several species. To better understand the systematics and ultimately the origins of breadfruit, we considered evidence from molecular data. Amplified fragment length polymorphism data for 261 individuals of breadfruit, its closest relatives, putative hybrids, and nine outgroup taxa were analyzed using neighbor joining and parsimony analyses. Three species, A. altilis (domesticated breadfruit), A. camansi, and A. mariannensis, are recognized and the existence of hybrids (A. altilis × A. mariannensis) verified. A revised treatment based on the molecular results, as well as morphological and geographical considerations, is presented.

Phytochemical fingerprinting to thwart black cohosh adulteration: a 15 Actaea species analysis.

INTRODUCTION The popular use of black cohosh products (Actaea racemosa L., syn. Cimicifuga racemosa L.) is growing as the demand for alternatives to estrogen therapy has increased. Critical to safe use is the assurance of unadulterated, high-quality products. Questions have been raised about the safety of black cohosh due to cases of liver toxicity in patients who reported taking it; subsequent evaluation found some products to be adulterated with other related herbal species. Correct plant species identification is a key first step for good manufacturing practices of safe black cohosh products. OBJECTIVES To develop analytical methods which distinguish black cohosh from other species (American and Asian) of Actaea increasingly found as adulterants in commercially available black cohosh products. MATERIAL AND METHODS Fifteen species of Actaea were collected from North America and Asia, and the phytochemical fingerprints of these samples were established using HPLC-PDA and LC-MS techniques. RESULTS The HPLC and LC-MS fingerprints for polyphenols and triterpene glycosides revealed distinct patterns that make black cohosh clearly distinguishable from most other species of Actaea. Two marker compounds, cimifugin and cimiracemoside F, were found to be important to distinguish black cohosh from most Asian species of Actaea. Formononetin was not found from either Asian or American species of Actaea. CONCLUSIONS Phytochemical fingerprinting is a practical, reliable method for authenticating black cohosh and distinguishing it from other species of Actaea increasingly found as adulterants in commercially available black cohosh products. This should facilitate the continued development of high-quality, unadulterated black cohosh products.

Cladogenesis and reticulation in the Hawaiian endemic mints (Lamiaceae)

The Hawaiian endemic mints, which comprise 58 species of dry‐fruited Haplostachys and fleshy‐fruited Phyllostegia and Stenogyne, represent a major island radiation that likely originated from polyploid hybrid ancestors in the temperate North American Stachys lineage. In contrast with considerable morphological and ecological diversity among taxa, sequence variation in the nrDNA 5S non‐transcribed spacer was found to be remarkably low, which when analyzed using standard parsimony resulted in a lack of phylogenetic resolution among accessions of insect‐pollinated Phyllostegia and bird‐pollinated Stenogyne. However, many within‐individual nucleotide polymorphisms were observed, and under the assumption that they could contain phylogenetic information, these ambiguities were recoded as new character states. Substantially more phylogenetic structure was obtained with these data, including the resolution of most Stenogyne species into a monophyletic group with an apparent recent origin on O’ahu (3.0 My) or the Maui Nui island complex (2.2 My). Subsequent diversification appears to have involved multiple inter‐island dispersal events. Intergeneric placements for a few morphotypes, seemingly misplaced within either Phyllostegia or Stenogyne, may indicate reticulation as one polymorphism‐generating force. For a finer scale exploration of hybridization, preliminary AFLP fragment data were examined among putative hybrids of Stenogyne microphylla and S. rugosa from Mauna Kea, Hawai’i, that had been identified based on morphology. Cladistic analysis (corroborated by multivariate correspondence analysis) showed the morphologically intermediate individuals to group in a strongly supported monophyletic clade with S. microphylla. Therefore, reticulation could be both historic and active in Stenogyne, and perhaps a force of general importance in the evolution of the Hawaiian mints. The relatively greater extent of lineage‐sorted polymorphisms in Stenogyne may indicate selective differentiation from other fleshy‐fruited taxa, perhaps through the agency of highly specialized bird pollinators that restricted gene flow with other Hawaiian mint morphotypes.

Chemical and Genetic Assessment of Variability in Commercial Radix Astragali (Astragalus spp.) by Ion Trap LC-MS and Nuclear Ribosomal DNA Barcoding Sequence Analyses

Radix Astragali (Huangqi) has been demonstrated to have a wide range of immunopotentiating effects and has been used as an adjuvant medicine during cancer therapy. Identity issues in the collection of Radix Astragali exist because many sympatric species of Astragalus occur in the northern regions of China. In order to assess the quality, purity, and uniformity of commercial Radix Astragali, 44 samples were purchased from herbal stores in Hong Kong and New York City. The main constituents, including four isoflavonoids and three saponins, were quantitatively determined by liquid chromatography mass spectrometry (LC-MS). There was significant sample-to-sample variability in the amounts of the saponins and isoflavonoids measured. Furthermore, DNA barcoding utilizing the variable nuclear ITS spacer regions of the 44 purchased Radix Astragali samples were sequenced, aligned and compared. Eight polymorphic point mutations were identified which separated the Radix Astragali samples into three groups. These results indicate that the chemical and genetic variability that exists among Radix Astragali medicinal products is still a consistency and quality issue for this herbal. Two-way ANOVA analysis showed significant effects on the contents of the seven tested compounds when both phylogenetic and geographic (i.e., point of purchase) factors were considered. Therefore, chemical profiles determined by LC-MS and DNA profiles in ITS spacer domains could serve as barcode markers for quality control of Radix Astragali.

POLYPHYLY IN GUETTARDA L. (RUBIACEAE, GUETTARDEAE) BASED ON NRDNA ITS SEQUENCE DATA1, 2

Abstract The genus Guettarda (Guettardeae–Rubiaceae) comprises approximately 150 species, ranging from eastern Africa through the islands of the Indian and Pacific Oceans to the Neotropics. Sequence data from the nuclear ribosomal DNA internal transcribed spacer (ITS) region were used to test the monophyly of Guettarda and its relationships to closely related genera within Guettardeae. The results indicate that Guettarda and two smaller genera, Antirhea and Stenostomum, are polyphyletic. Most Guettarda species fall into two distinct groups: a Neotropical lineage that also includes the widespread Indo-Pacific strand species G. speciosa (the type of the genus), and a New Caledonian lineage that, along with Antirhea and Timonius, comprises a dioecious Paleotropical clade. The Hawaiian endemic Bobea, traditionally considered close to Timonius and assumed to be of Old World origin, appears to be more closely related to Neotropical Guettarda species, suggesting that dioecy may have evolved twice within the tribe. The use of traditional gynoecium characters to delimit genera within Guettardeae is not congruent with the ITS phylogeny; other features, such as inflorescence architecture, sexual system, and palynology, appear to correlate more closely with the molecular phylogeny.