Sooty Mould on Hibiscus rosa‐sinensis Caused by Leptoxyphium kurandae is Associated with Extrafloral Nectaries

In August 2013, sooty mould was observed on Chinese hibiscus (Hibiscus rosa‐sinensis) in a propagation nursery in Seoul, Korea. The sooty mould initially developed at the junction between the leaf blade and leaf petiole and then dispersed along the vein on the abaxial surface. The fungal growth pattern on the plants was quite different from general sooty moulds growing on honeydew secreted by insects on the plants. On the basis of the morphological characteristics and phylogenetic analysis using the internal transcribed spacer rDNA, this fungus was identified as Leptoxyphium kurandae. A pathogenicity test was carried out to fulfil Koch's postulates. Through field observation and a pathogenicity test, we found an association between the sooty mould and extrafloral nectaries. To our knowledge, this is the first report of sooty mould caused by L. kurandae on the extrafloral nectaries of H. rosa‐sinensis.     

Crotalaria spectabilis and Raphanus sativus as Previous Crops Show Promise for the Control of Bacterial Wilt of Tomato Without Reducing Bacterial Populations

Ralstonia solanacearum is responsible for bacterial wilt affecting many crops worldwide. The emergent population of R. solanacearum (phylotype IIB/4NPB) wilts previously resistant varieties and has rapidly spread throughout Martinique. No conventional method is known to control it. In this study, previous crops used as sanitizing crops were investigated as an environmentally safe alternative method of control. The ability of the emergent population of R. solanacearum to persist in planta and in the rhizosphere of Brassicaceae, Asteraceae and Fabaceae grown as previous crops was evaluated in controlled conditions, and the incidence of bacterial wilt was assessed in the following tomato crop. Results showed that all species carried R. solanacearum latently. Among Brassicaceae and Asteraceae, the highest density of R. solanacearum was found in planta and in the rhizosphere of Tagetes erecta. The density of the R. solanacearum population in the rhizosphere of Raphanus sativus cv. Karacter was significantly higher than that in Raphanus sativus cv. Melody. In Fabaceae, the density of R. solanacearum population in planta was statistically similar in all species. The density of the R. solanacearum population in the rhizosphere of Crotalaria juncea was significantly higher than that in Crotalaria spectabilis. This study showed for the first time that Crotalaria spectabilis and Raphanus sativus cv. Melody grown as previous crops improve the performance of the following tomato with similar effects on R. solanacearum populations in the soil as bare soil. The incidence of the disease in tomato decreased by 86% and 60%, after R. sativus cv. Melody and C. spectabilis, respectively, and the proportion of infected plants also decreased. These results suggest that C. spectabilis and R. sativus cv. Melody can be used as previous crops to help bacterial wilt control in ecological management strategies without drastic suppression of R. solanacearum population in stem tissues and in the rhizosphere.     

New Host Record of Myrothecium roridum Causing Leaf Spot on Abutilon megapotamicum from China

During September 2010, Abutilon megapotamicum plants with dark‐brown concentric spots on leaves were observed in a commercial glasshouse located in Beijing, China. This study was carried out to identify the causal agent of this disease based on Koch's postulates and morphological characteristics. Pathogenicity tests in the glasshouse showed that Myrothecium roridum Tode ex Fr. caused the leaf spot on A. megapotamicum plants, which were the same as those observed in naturally infected plants in the field. Moreover, to confirm the pathogen to species, the rDNA internal transcribed spacer (ITS) of isolate was PCR‐amplified using ITS1 and ITS4 primer pairs and sequenced. DNA analysis revealed a 100% species identity index for M. roridum. To the best of our knowledge, this is the first report of M. roridum on A. megapotamicum in China.     

What happens in the pith stays in the pith: tissue‐localized defense responses facilitate chemical niche differentiation between two spatially separated herbivores

Herbivore attack is known to elicit systemic defense responses that spread throughout the host plant and influence the performance of other herbivores. While these plant‐mediated indirect competitive interactions are well described, and the co‐existence of herbivores from different feeding guilds is common, the mechanisms of co‐existence are poorly understood. In both field and glasshouse experiments with a native tobacco, Nicotiana attenuata, we found no evidence of negative interactions when plants were simultaneously attacked by two spatially separated herbivores: a leaf chewer Manduca sexta and a stem borer Trichobaris mucorea. T. mucorea attack elicited jasmonic acid (JA) and jasmonoyl‐l ‐isoleucine bursts in the pith of attacked stems similar to those that occur in leaves when M. sexta attacks N. attenuata leaves. Pith chlorogenic acid (CGA) levels increased 1000‐fold to levels 6‐fold higher than leaf levels after T. mucorea attack; these increases in pith CGA levels, which did not occur in M. sexta‐attacked leaves, required JA signaling. With plants silenced in CGA biosynthesis (irHQT plants), CGA, as well as other caffeic acid conjugates, was demonstrated in both glasshouse and field experiments to function as a direct defense protecting piths against T. mucorea attack, but not against leaf chewers or sucking insects. T. mucorea attack does not systemically activate JA signaling in leaves, while M. sexta leaf‐attack transiently induces detectable but minor pith JA levels that are dwarfed by local responses. We conclude that tissue‐localized defense responses allow tissue‐specialized herbivores to share the same host and occupy different chemical defense niches in the same hostplant.     

Mixed Infection and Natural Spread of ‘Candidatus Phytoplasma asteris’ and Mungbean Yellow Mosaic India Virus Affecting Soya Bean Crop in India

Suspected phytoplasma and virus‐like symptoms of little leaf, yellow mosaic and witches’ broom were recorded on soya bean and two weed species (Digitaria sanguinalis and Parthenium hysterophorus), at experimental fields of Indian Agricultural Research Institute, New Delhi, India, in August–September 2013. The phytoplasma aetiology was confirmed in symptomatic soya bean and both the weed species by direct and nested PCR assays with phytoplasma‐specific universal primer pairs (P1/P6 and R16F2n/R16R2n). One major leafhopper species viz. Empoasca motti Pruthi feeding on symptomatic soya bean plants was also found phytoplasma positive in nested PCR assays. Sequencing BLASTn search analysis and phylogenetic analysis revealed that 16Sr DNA sequences of phytoplasma isolates of soya bean, weeds and leafhoppers had 99% sequence identity among themselves and were related to strains of ‘Candidatus Phytoplasma asteris’. PCR assays with Mungbean yellow mosaic India virus (MYMIV) coat‐protein‐specific primers yielded an amplicon of approximately 770 bp both from symptomatic soya bean and from whiteflies (Bemisia tabaci) feeding on soya bean, confirmed the presence of MYMIV in soya bean and whitefly. Hence, this study suggested the mixed infection of MYMIV and ‘Ca. P. asteris’ with soya bean yellow leaf and witches’ broom syndrome. The two weed species (D. sanguinalis and P. hysterophorus) were recorded as putative alternative hosts for ‘Ca. P. asteris’ soya bean Indian strain. However, the leafhopper E. motti was recorded as putative vector for the identified soya bean phytoplasma isolate, and the whitefly (B. tabaci) was identified as vector of MYMIV which belonged to Asia‐II‐1 genotype.     

Relationship Between Ralstonia solanacearum Diversity and Severity of Bacterial Wilt Disease in Tomato Fields in China

A field survey was conducted to determine the relationship between Ralstonia solanacearum diversity and severity of bacterial wilt disease in tomato plants grown in plastic greenhouses. Both vegetative and reproductive stages of the plants were surveyed, and the symptoms were empirically categorized into five scales: 0 (asymptomatic): 1st, 2nd, 3rd and 4th. The bacterial wilt pathogen was isolated from infected plants at each disease scale; pathogenic characteristics and population densities of the bacterial strains were assessed. Two hundred and eighty‐two isolates were identified as R. solanacearum, which were divided into three pathogenic types, virulent, avirulent and interim, using the attenuation index (AI) method and a plant inoculation bioassay. Ralstonia solanacearum was detected in all asymptomatic and symptomatic tomato plants, with population numbers, ranging from 10.5 to 86.7 × 105 cfu/g. However, asymptomatic plants harboured only avirulent or interim R. solanacearum, whereas tomato plants displaying 1st or 2nd disease degree contained interim and virulent strains. Additionally, 3rd and 4th degree plants harboured only virulent strains. The disease was more severe in vegetative‐stage plants (disease severity index (DSI) 0.20) with higher total numbers of interim and virulent R. solanacearum strains than those in reproductive‐stage plants (DSI 0.12). Three pathotypes of R. solanacearum coexisted in a competitive growth system in the tomato field, and their distribution closely correlated with the severity of tomato bacterial wilt.     

Pythium aphanidermatum Causing Pythium Rot on Lampranthus spectabilis in Korea

Pythium aphanidermatum was found to be associated with rotting of leaves and stems in Lampranthus spectabilis in the summers of 2013 and 2014. Infected plants were initially characterized by water‐soaked and discoloured tissue, which are soon covered with cottony aerial hyphae. Subsequently, infected tissues wilted, leaves and stems appeared desiccated, and infected plants died. Pathogenicity of a representative isolate was proved, fulfilling Koch's postulates. Sequence of the internal transcribed spacer region of ribosomal DNA from the isolate shared 100% identity with that of P. aphanidermatum. To our knowledge, this is the first report of P. aphanidermatum causing leaf and stem rot on L. spectabilis in Korea.     

Miniprimer PCR assay targeting multiple genes: a new rapid and reliable tool for genotyping Pantoea stewartii subsp. stewartii

Aim: Development of a ‘miniprimer’ PCR assay for genotyping Pantoea stewartii subsp. stewartii, the causal agent of the Stewart’s bacterial wilt on maize. Methods and Results: Four 10‐nucleotide (10‐nt) ‘miniprimer’ sets were designed and evaluated in the presence of Titanium Taq DNA polymerase. Under optimal reaction conditions, the miniprimer pair Uni‐BacF‐10/Uni‐BacR‐10 reproducibly generated identical banding patterns among 10 strains of P. stewartii subsp. stewartii, different patterns from strains of P. stewartii subsp. indologenes, other Panteoa species, Clavibacter michiganensis, Pectobacterium spp., Pseudomonas spp. and other bacterial species. The amplicons of Pantoea stewartii subsp. stewartii were cloned and sequenced to identify genes or DNA fragments that are targeted by the miniprimer PCR assay. Of the 14 ‘clone types’ identified, sequences of a 1·23‐kb fragment had a 99·8% similarity to part of the Pantoea stewartii zeaxanthin diglucoside biosynthetic operon ( AY166713 ). Other dominant cloned fragments included a 411‐bp amplicon that exhibited 99·8% similarity to the psaU gene (syn:ysaU; GQ249669 ), a type III protein‐secretion system complex of P. stewartii subsp. stewartii strain DC283, and a 548‐bp fragment showed 63% homology to the Asp/Glu racemase encoding gene in Erwinia tasmaniensis strain ET1/99. Conclusion: The miniprimer PCR assay reported here is highly discriminatory and reproducible in genotyping Pantoea stewartii subsp. stewartii. Significance and Impact of the study: This miniprimer PCR assay could be a new reliable and rapid tool for fingerprinting the Stewart’s wilt pathogen of maize.     

Occurrence of a New Orange Variant of Curtobacterium flaccumfaciens pv. flaccumfaciens,Causing Common Bean Wilt in Iran

A new orange variant of Curtobacterium flaccumfaciens pv. flaccumfaciens was isolated from seeds of common bean cv. Daneshkadeh and Dehghan stored in the seed banks in Khomein Bean Research Station, and field plants (cv. Local Khomein) in Arak, Iran. The pathogenicity of the isolates was confirmed on 5‐ to 7‐day‐old seedlings of cv. Daneshkadeh. Marginal necrosis and interveinal chlorosis on first trifoliate leaves were observed 10–15 days after inoculation. Amplification of 306 bp fragment of orange‐pigmented strains using CffFOR2‐ and CffREV4‐specific prime pair characterized them as C. flaccumfaciens pv. flaccumfaciens. Although the yellow‐pigmented variant of the causal agent was previously reported on cowpea, this is the first report of orange variant of C. flaccumfaciens pv. flaccumfaciens causing bacterial wilt on common bean in Iran.     

Variation in abundance of predicted resistance genes in the Brassica oleracea pangenome

Brassica oleracea is an important agricultural species encompassing many vegetable crops including cabbage, cauliflower, broccoli and kale; however, it can be susceptible to a variety of fungal diseases such as clubroot, blackleg, leaf spot and downy mildew. Resistance to these diseases is meditated by specific disease resistance genes analogs (RGAs) which are differently distributed across B. oleracea lines. The sequenced reference cultivar does not contain all B. oleracea genes due to gene presence/absence variation between individuals, which makes it necessary to search for RGA candidates in the B. oleracea pangenome. Here we present a comparative analysis of RGA candidates in the pangenome of B. oleracea. We show that the presence of RGA candidates differs between lines and suggests that in B. oleracea, SNPs and presence/absence variation drive RGA diversity using separate mechanisms. We identified 59 RGA candidates linked to Sclerotinia, clubroot, and Fusarium wilt resistance QTL, and these findings have implications for crop breeding in B. oleracea, which may also be applicable in other crops species.     

Identification and Characterization of Paraphoma chrysanthemicola Causing Leaf Spot Disease on Atractylodes japonica in China

Atractylodes japonica is a perennial herb in Compositae family, which is used for stomach disorders as a traditional Chinese medicine (Guo et al. 2006 ). In 2013, a leaf spot disease was first observed on plants of A. japonica in a production field of Fushun County, Liaoning Province, China. The disease had a speckled appearance initially. Lesions with grey–white centre and brown margin gradually developed and enlarged. Eventually, infections usually caused yellowing of the leaves and premature defoliation. The causal agent of infection on plants was identified as Paraphoma chrysanthemicola based on morphological and cultural characteristics, pathogenicity tests and phylogenetic analysis. To our knowledge, this is the first report of a leaf spot disease on A. japonica caused by P. chrysanthemicola in China.     

Influence of tomato spotted wilt virus on performance and behaviour of western flower thrips (Frankliniella occidentalis)

The general principles in pathogen transmission by insects involve a complex and specific interplay, in this case between thrips, tospovirus and their shared host plant, which has led to outbreaks of crop disease epidemics of economic and social importance. The possible processes and factors driving their co‐evolution were partly studied by rearing Frankliniella occidentalis [western flower thrips (WFT)] on either tomato spotted wilt virus (TSWV)–infected or uninfected Capsicum annum leaflets throughout their larval stages. Later, pupae were transferred individually on healthy leaf discs for further studies of the influence of TSWV on WFT development and behavioural patterns. The exposure of WFT to TSWV was found to improve performance with regard to longevity and survival, with mean longevity being significantly higher in TSWV‐exposed WFT compared to unexposed ones (F_(3,403) = 22.44, P < 0.0001). The observed improvement in survival was as a result of significant reduction in mortality for the WFT individuals exposed to TSWV (F_(3,383) = 849.94, P < 0.0001) compared to the unexposed. However, the results showed a significant reduction in mean daily fecundity overtime (F_10,10) = 246.66, P < 0.0001) and across the four treatments (F_(3,30) = 6.62, P = 0.001), as well as lifetime fecundity (F_(3,23) = 21.23, P < 0.0001) of the WFT exposed to TSWV compared to the unexposed reared on uninfected leaf discs. For preferential test, C. annum leaf discs infected with TSWV were more attractive to WFT as compared to healthy leaf discs (χ²_{(4, 34)} = 112.35, P < 0.0001). These results are envisaged to contribute to a clear understanding into the plant–vector–virus interaction, which is essential for accurate diagnosis and control of the TSWV epidemic, as well as the control of F. occidentalis as crop pest.     

Bioassay‐Guided Isolation of Antifungal Compounds from Disporopsis aspersa (Hua) Engl. ex Diels against Pseudoperonospora cubensis and Phytophthora infestans

Oomycetes are one type of the most highly destructive of the diseases that cause damage to some important crop plants, such as potato late blight, cucumber downy mildew, and grape downy mildew. As main approach of the ongoing search for new botanical fungicide from plant, the secondary metabolites of D. aspersa were investigated. Through efficient bioassay‐guided isolation, two new ( 1 and 2 ) and 12 known compounds ( 3  –  14 ) were isolated, and their structures were determined via extensive NMR, HR‐ESI‐MS, and IR. They were isolated from this genus for the first time except for compounds 11 and 12 . The biological properties of 1  –  14 were evaluated against Pseudoperonospora cubensis and Phytophthora infestans. Compounds 1  –  8 showed potent antifungal activity in vitro. Additionally, compound 3 has preferable control effect on cucumber downy mildew, showing dual effect of protection and treatment in vivo.     

Effects of leaf conditions and flight activity on the behaviour of Platypus quercivorus (Murayama) (Coleoptera: Platypodidae)

The important role of semiochemicals in the interactions between plants and insects has been extensively investigated. The volatiles produced by oak trees are thought to attract the ambrosia beetle Platypus quercivorus (Murayama), which causes wilt disease in Quercus trees, resulting in widespread damage. In the present study, we hypothesized that (a) P. quercivorus is attracted to the leaf volatiles emitted by host trees and (b) the response of P. quercivorus to leaf volatiles is affected by flight. An experiment was performed to survey the preferences of both sexes of P. quercivorus for the leaf volatiles of Quercus crispula Blume at various stages of leaf deterioration, represented by different number of days after cutting. Additionally, the effect of flight on both sexes was evaluated by testing the beetle flight on a flight mill. The results showed that P. quercivorus was attracted to the volatiles emitted from fresh leaves and was not attracted to those emitted from dry leaves. This suggests that leaf volatiles from healthy hosts are primary attractants for P. quercivorus. Further, males exhibited increased olfactory responses to leaf volatiles as their flight duration increased, whereas the opposite pattern was observed in females. These results suggest that the different ecological roles of the sexes contribute to differences in olfactory responses.     

Dynamic infection of Verticillium dahliae in upland cotton

• Verticillium wilt, an infection caused by the soilborne fungus Verticillium dahliae, is one of the most serious diseases in cotton. No effective control method against V. dahliae has been established, and the infection mechanism of V. dahliae in upland cotton remains unknown.
• GFP‐tagged V. dahliae isolates with different pathogenic abilities were used to analyse the colonisation and infection of V. dahliae in the roots and leaves of different upland cotton cultivars, the relationships among infection processes, the immune responses and the resistance ability of different cultivars against V. dahliae.
• Here, we report a new infection model for V. dahliae in upland cotton plants. V. dahliae can colonise and infect any organ of upland cotton plants and then spread to the entire plant from the infected organ through the surface and interior of the organ.
• Vascular tissue was found to not be the sole transmission route of V. dahliae in cotton plants. In addition, the rate of infection of a V. dahliae isolate with strong pathogenicity was notably faster than that of an isolate with weak pathogenicity. The resistance of upland cotton to Verticillium wilt was related to the degree of the immune response induced in plants infected with V. dahliae. These results provide a theoretical basis for studying the mechanism underlying the interaction between V. dahliae and upland cotton. These results provide a theoretical basis for studying the mechanism underlying the interaction between V. dahliae and upland cotton.

     

Global change‐type drought‐induced tree mortality: vapor pressure deficit is more important than temperature per se in causing decline in tree health

Drought‐induced tree mortality is occurring across all forested continents and is expected to increase worldwide during the coming century. Regional‐scale forest die‐off influences terrestrial albedo, carbon and water budgets, and land‐surface energy partitioning. Although increased temperatures during drought are widely identified as a critical contributor to exacerbated tree mortality associated with “global‐change‐type drought”, corresponding changes in vapor pressure deficit (D) have rarely been considered explicitly and have not been disaggregated from that of temperature per se. Here, we apply a detailed mechanistic soil–plant–atmosphere model to examine the impacts of drought, increased air temperature (+2°C or +5°C), and increased vapor pressure deficit (D; +1 kPa or +2.5 kPa), singly and in combination, on net primary productivity (NPP) and transpiration and forest responses, especially soil moisture content, leaf water potential, and stomatal conductance. We show that increased D exerts a larger detrimental effect on transpiration and NPP, than increased temperature alone, with or without the imposition of a 3‐month drought. Combined with drought, the effect of increased D on NPP was substantially larger than that of drought plus increased temperature. Thus, the number of days when NPP was zero across the 2‐year simulation was 13 or 14 days in the control and increased temperature scenarios, but increased to approximately 200 days when D was increased. Drought alone increased the number of days of zero NPP to 88, but drought plus increased temperature did not increase the number of days. In contrast, drought and increased D resulted in the number of days when NPP = 0 increasing to 235 (+1 kPa) or 304 days (+2.5 kPa). We conclude that correct identification of the causes of global change‐type mortality events requires explicit consideration of the influence of D as well as its interaction with drought and temperature.     

Diamondback moth egg susceptibility to rainfall: effects of host plant and oviposition behavior

Oviposition patterns of the diamondback moth (DBM), Plutella xylostella L. (Lepidoptera: Plutellidae), differ between common cabbage (Brassica oleracea L. var. capitata) and Chinese cabbage (Brassica rapa L. var. pekinensis) (Brassicaceae) host plants. This study shows that the moth prefers to oviposit on adaxial rather than abaxial leaf surfaces and petioles of both host plants. More eggs were laid in leaf veins than on leaf laminas of both host plants, especially in Chinese cabbage, where 94.6% of eggs were laid in veins. On Chinese cabbage, very few eggs were laid in clusters (≥2 eggs), whereas on common cabbage approximately 30% of eggs were laid in groups of 2 or more eggs. Removal of wax from common cabbage leaves dramatically increased the number of eggs laid singly on the leaf lamina of treated plants, suggesting that leaf waxes affect how eggs are distributed by ovipositing DBM. Eggs were most susceptible to removal by rainfall from the plant surface immediately (<1 h) after oviposition and when close to hatching (>72h old) whereas they were least susceptible 24 h after oviposition. Eggs laid on common cabbage plants were more susceptible to simulated rainfall than eggs laid on Chinese cabbage plants. On common cabbage plants, egg susceptibility to rainfall on different plant parts ranked adaxial leaf surfaces>petioles = abaxial leaf surfaces>stem, but there was no difference in egg susceptibility to rainfall on the various plant parts of Chinese cabbage. Furthermore, on common cabbage plants, eggs laid on both adaxial and abaxial leaf surfaces were afforded significant protection from the effects of rainfall by leaves higher in the plant canopy. On common cabbage plants, oviposition patterns reduce the potential impact of rainfall on eggs, possibly reducing the effect of this important abiotic mortality factor in the field.     

Effect of Zinc on the Development of Brown Spot in Rice

Brown spot, caused by the fungus Bipolaris oryzae, is one of the most destructive diseases of rice. This study investigated the effect of zinc rates on the development of brown spot in rice. Rice plants (cv. ‘Metica‐1′) were grown in hydroponic culture amended with Zn rates (applied as ZnSO₄^.7H₂O) of 0, 0.5, 1, 2 and 4 μm and inoculated with B. oryzae. The foliar concentration of Zn was determined. Leaf samples were assessed for disease severity, and then, area under brown spot progress curve (AUBSPC) was calculated. The relationship between Zn concentrations on leaf tissues and the rates of this micronutrient was best described by a positive linear regression model, while the relationship between the Zn rates and the AUBSPC was best described with a positive quadratic regression model. The correlation between Zn concentrations on leaf tissues and AUBSPC was positive and significant (r = 0.68, P < 0.05). The results from this study showed that high foliar concentration of Zn was associated with increasing rice susceptibility to brown spot.     

Identification of species and materia medica within Angelica L. (Umbelliferae) based on phylogeny inferred from DNA barcodes

DNA barcodes have been increasingly used in authentication of medicinal plants, while their wide application in materia medica is limited in their accuracy due to incomplete sampling of species and absence of identification for materia medica. In this study, 95 leaf accessions of 23 species (including one variety) and materia medica of three Pharmacopoeia‐recorded species of Angelica in China were collected to evaluate the effectiveness of four DNA barcodes (rbcL, matK, trnH‐psbA and ITS). Our results showed that ITS provided the best discriminatory power by resolving 17 species as monophyletic lineages without shared alleles and exhibited the largest barcoding gap among the four single barcodes. The phylogenetic analysis of ITS showed that Levisticum officinale and Angelica sinensis were sister taxa, which indicates that L. officinale should be considered as a species of Angelica. The combination of ITS + rbcL + matK + trnH‐psbA performed slight better discriminatory power than ITS, recovering 23 species without shared alleles and 19 species as monophyletic clades in ML tree. Authentication of materia medica using ITS revealed that the decoction pieces of A. sinensis and A. biserrata were partially adulterated with those of L. officinale, and the temperature around 80 °C processing A. dahurica decoction pieces obviously reduced the efficiency of PCR and sequencing. The examination of two cultivated varieties of A. dahurica from different localities indicated that the four DNA barcodes are inefficient for discriminating geographical authenticity of conspecific materia medica. This study provides an empirical paradigm in identification of medicinal plants and their materia medica using DNA barcodes.