Analysis of genetic diversity in Turkish sesame (Sesamum indicum L.) populations using RAPD markers⋆

The genetic diversity of 38 cultivated populations of Sesamum indicum L. from four different regions of Turkey was estimated at the DNA level with the random amplified polymorphic DNA (RAPD) technique. Sixty-one bands were obtained for all populations 78% of which were polymorphic. Analysis of molecular variance (AMOVA) was used to investigate the genetic diversity of the populations which yielded highly significant differences among populations within regions (91.9% of the total genetic diversity). According to AMOVA and Shannon's index that were performed separately for each region, the highest value of genetic variation was observed among Northwest region populations (CV = 7.7; H₀ = 0.304) and lowest in the Southeast regions' populations (CV = 2.6; H₀ = 0.068). Nei and Li's similarity index was calculated and phylogenetic tree was established using the neighbor-joining algorithm. This phenetic analysis grouped 35 of 38 accessions in six groups leaving three highly diverse accessions outside. Wagner phylogenetic method was used to assess the phylogenetic relationships among the populations. In the majority-rule consensus tree, only 7 of the 32 forks showed above 60% occurrence. Using Principal Coordinate Analysis (PCO) of the RAPD data set, the groups were clearly separated along the first three axis. These results indicate that RAPD technique is useful for sesame systematics, and should be valuable for the maintenance of germplasm banks and the efficient choice of parents in breeding programs.     

Genetic diversity in bambara groundnut (Vigna subterranea (L.) Verdc) landraces assessed by Random Amplified Polymorphic DNA (RAPD) markers

Genetic diversity in 12 landraces of bambara groundnut (Vigna subterranea), an indigenous African legume, was evaluated using Random Amplified Polymorphic DNA (RAPD) markers. DNA from individuals of each landrace was also analysed to determine the level of heterogeneity within landraces. RAPDs revealed high levels of polymorphism among landraces. The percentage polymorphism ranged from 63.2% to 88.2% with an average of 73.1% for the 16 RAPD primers evaluated. The construction of genetic relationships using cluster analysis groups the 12 landraces in two clusters. RAPDs are useful for the genetic diversity studies in V. subterranea and can identify variation within landraces.     

Genetic diversity analysis in Boerhavia diffusa L. of different geographic locations in India using RAPD markers

Boerhavia diffusa is extensively used in herbal medicines as well as in the Ayurvedic system, because it contains a set of clinically important compounds. In the present study, the genetic variability in Boerhavia diffusa between accessions of different geographical origin within the Indian territory is assessed through random amplified polymorphic DNA (RAPD) markers. Twenty-eight accessions of Boerhavia were screened with eighteen primers of which nine were found to be the most informative. The degree of polymorphism was found to be high in accessions collected from different places of Uttar Pradesh (Set II) in comparison to other states of India (Set I). A relatively lower level of polymorphism was recorded in accessions collected from diverse locations around Lucknow (Set III). Accessions from neighbouring geographical regions exhibited more similarity than those from distant regions (as revealed by the set I analysis). Certain diagnostic markers may be correlated with morphological character(s) such as plant type. BDL appeared most distinct and divergent from the rest of the accessions and the BDJ plant in set II also showed least similarity estimate. Fragments of 5.62 Kb and 4.47 Kb with primer GN59 was found to be unique for BDP and BD2 having ovate leaf character, whereas ovoid leaf genotype exhibited 0.79 Kb (GN34 primer) fragment. Similarly a unique band type (0.35 Kb) with primer GN83 was present in BDL and BD1 that share light pink flower. Jaccard's, and Nei and Li similarity coefficient values amongst the accessions were in the range of 0.22 to 0.89 and 0.33 to 0.93, respectively. Association of RAPD markers with the leaf characteristics, flower colour as well as with geographical locations has been made. This shows that RAPD markers are also useful for the study of genetic structure of Boerhavia populations.     

Genetic diversity of the African hexaploid species Solanum scabrum Mill. and Solanum nigrum L. (Solanaceae)

Two hexaploid species of Solanum sect. Solanum are present in Africa: Solanum scabrum and S. nigrum. Solanum scabrum is a widely cultivated species and is used as a leafy vegetable, as a source of medicine and as a source of ink dye. In previous studies a wide range of morphological diversity has been reported in this species and in some studies subspecies have been proposed. Subspecies are also recognized in S. nigrum. However, it has not been established whether or not the morphological differences are reflected at the genomic level. The present study applies AFLPs to study the genetic diversity in S. scabrum and its relationship to geographical provenance, morphological differences and the possible existence of subspecies within S. scabrum and S. nigrum. The data obtained were analyzed with cluster analysis (using UPGMA and NJ). The results indicate that the genetic variation within S. scabrum was higher within accessions than between accessions. Accessions did not cluster according to their geographical provenance, indicating that accessions from different geographical areas were not significantly different genetically. The clustering reflected neither morphological differences nor domestication status (cultivated or wild). The morphological differences exhibited by S. scabrum could be due to selection by farmers for different plant types. The AFLP derived clustering pattern did not segregate the subspecies recognized in S. scabrum and S. nigrum into separate subclusters.     

Analysis of genetic diversity in Indian taro [Colocasia esculenta (L.) Schott] using random amplified polymorphic DNA (RAPD) markers

Taro [Colocasia esculenta (L.) Schott] germplasm accessions collected from different parts of India were subjected to RAPD (Random Amplified Polymorphic DNA) analysis to assess the genetic diversity prevalent and also to test the genetic basis of morphotypic classification. Thirteen random decamer primers out of the 22 tested were used to analyse 32 taro accessions belonging to 28 morphotypes. Three out of these thirteen primers analysed showed 100 per cent polymorphism. Per cent polymorphism varied from 60 to 100 among the polymorphic primers. High genetic diversity was revealed as the similarity coefficient values ranged from 0.50 to 0.98. No two accessions analysed in the present study showed a similarity coefficient value of one thereby indicating their distinctness and presence of high genetic diversity in Indian taro germplasm. Dendrogram obtained from UPGMA analysis grouped 32 accessions in four clusters and three accessions were placed as outliers. Clustering pattern did not show any strict relationship with geographical distribution, morphotype classification and genotypic diversity. Further, accessions classified, as belonging to the same morphotypic group did not always cluster together. Presence of a very close genepool of the wild, weedy and cultivated forms with extreme levels of phenotypic and genotypic variation is suggested as the reason for high genetic diversity reported. Usefulness of DNA markers such as RAPD in characterising and assessing the genetic diversity in Indian taro germplasm is hereby demonstrated.     

Genetic diversity of Pakistan wheat germplasm as revealed by RAPD markers

Wheat breeding in Pakistan started in 1930s before partition in the United India and so far has released more than 68 cultivars, but no systematic analyses of the genetic diversity of Pakistan wheat have been made. Twenty Pakistan wheat cultivars released from 1933 to 2002 were examined for genetic diversity and relationships using random amplified polymorphic DNA (RAPD) markers. Forty-two RAPD primers were applied and 184 polymorphic bands were generated for each cultivar. Most of the cultivars were genetically interrelated, although six of them displayed some genetic distinctness. The RAPD variation observed among these cultivars was low. Only 40.7% of the total scorable bands were polymorphic, and 26.1% of the polymorphic bands were observed most frequently (f = 0.95) among the 20 cultivars. The proportions of polymorphic bands for each cultivar ranged from 0.67 in ‘Yecora’ to 0.84 in ‘C-250’ with an average of 0.76. About 1.4% of the RAPD variation might have been fixed over the 69 years of wheat breeding, but such fixation was not statistically significant. These results are significant for future improvement and conservation of Pakistan wheat.     

Evaluation of genetic diversity and relationships within an on-farm collection of Perilla frutescens (L.) Britt. using microsatellite markers

The present study demonstrates utilization of 11 microsatellite markers to explore genetic diversity held in Perilla frutescens (L.) Britt. landrace accessions growing on farms in different parts of Korea and Japan and to assess their genetic relationships. All microsatellite loci were polymorphic and produced a total of 96 alleles ranging from 4 to 20, with an average of 8.7 alleles per locus. Of the 96 alleles found, a total of 15 unique landrace-specific alleles were observed at 9 different loci. The locus GBPFM203 provided the highest number of alleles (20), of which five were unique and each specific to a particular landrace accession. The occurrence of unique, accession-specific alleles presented molecular evidence for the generation of new alleles within on-farm collection of Perilla. The mean values of observed (H _O) and expected heterozygosity (H _E) were 0.39 and 0.68, respectively, indicating a considerable amount of polymorphism within this collection. A genetic distance-based phylogeny grouped the two Perilla varieties, var. frutescens and var. crispa (Thunb.) Decne into two distinct groups. Accessions belonging to var. frutescens could also be divided into two subgroups at a close genetic distance (GD = 0.432). The overall clustering pattern did not strictly follow the grouping of accessions according to their geographic origins. These observations are indicative of extensive germplasm exchange among farms from different geographical regions. The genetic similarity observed among the Perilla landraces may be useful for future Perilla crop variety identification, conservation, and improvement programs.     

Microsatellite markers and genetic diversity assessment in Lolium temulentum

Lolium and Festuca are two important genera of cool-season forage and turf grasses worldwide. Lolium temulentum L. (darnel ryegrass) has been proposed as a model species for genomics studies of cool-season forage and turf grasses. A study with 41 darnel ryegrass, three tall fescue (Festuca arundinacea Schreb.), two tetraploid fescue (F. glaucescens), and two meadow fescue (F. pratensis) genotypes was initiated to (i) identify a set of microsatellite (simple sequence repeats) markers useful for L. temulentum L., and (ii) to utilize such markers for assessing the genetic variability of L. temulentum accessions collected from different geographical regions of the world. A total of 40 tall fescue (TF) EST-SSRs and 60 Festuca–Lolium (F × L) genomic SSRs were screened on a subset of eight genotypes. The selected 30 tall fescue EST-SSRs and 32 F × L genomic SSRs were used for further analysis of genotypes. The TF-EST- and the F × L genomic-SSRs identified 10.3 and 9.3 alleles per marker, respectively with an average polymorphic information content (PIC) value of 0.66. The phenogram based on 319 EST-SSR and 296 genomic SSR fragments, grouped L. temulentum accessions into three major clusters except for accession ABY-BA 8892.78. Lolium temulentum accession ABY-BA 8892.78 did not cluster with any other accession. The Festuca clusters were distantly related with darnel ryegrass clusters with a similarity coefficient of 0.26. The selected set of tall fescue EST- and F × L genomic SSRs were useful in assessing L. temulentum genetic diversity and could benefit the genetic improvement of members of the Festuca–Lolium complex.     

Genetic variation in wild and cultivated artichoke revealedby RAPD markers

Determination of intraspecific genetic diversity is an important first step for the utilization of genetic resources and canprovide useful information on crop evolution. A living collection of Italian and foreign artichoke varieties and ecotypes is maintained at the Germplasm Institute, Bari, Italy. A total of 32 accessions of cultivated (Cynara cardunculus L. var.scolymus (L.) Fiori), three of wild (Cynara cardunculus var.cardunculus L.) artichoke and two ofcultivated cardoon (Cynara cardunculus var.altilis L.) were analysed in order to studygenetic variation and relationships within the species, using RAPDmarkers. Cultivated accessions were selected according tomorphological variation and geographical distribution available inthe collection. Fifty arbitrary decamer primers were initially tested, 18 ofwhich showed from 3 to 10 unambiguously interpretable fragments. Anintra-accession analysis using 4 varieties and 8 polymorphicprimers revealed that no RAPD variation was detected amongindividuals. Jaccard's similarity index (JSI) was comprisedamong 1 and 0.693 when all accessions were considered, on the otherhand, within the cultivated artichoke, JSI ranged between 1 and0.817. A UPGMA dendrogram based on the similarity matrix showed thatwild artichokes were clearly separated from cultivated accessions.Moreover, within cultivated artichokes, several groups could bedistinguished.     

Application of random amplified polymorphic DNA markers to evaluate intraspecific genetic variation in the Elymus alaskanus complex (Poaceae)

Random amplified polymorphic DNA (RAPD) markers were used to evaluatethe levels and patterns of genetic diversity in ten Elymusalaskanus populations, which were collected from Canada, USA,Greenland and Russia. Ten arbitrarily chosen decamer primers were used in thisstudy. The results revealed high levels of variation. The mean number of allelesper locus (Ap) was 1.5, ranging from 1.4 to 1.6, the meanpercent of polymorphic loci (Pp) was 49.5%, ranging from35.1% to 64.9%, and the mean gene diversity (Hep) was0.162, varying from 0.142 to 0.262. The total variation wasH _T = 0.403. When partitioned(G _ST), 60% of the total variation was foundamong the populations. Although the genetic diversity values obtained with RAPDsare much higher than for allozymes, they are similar regarding how the geneticvariation is distributed among populations. In addition, a similar geneticpattern of population differentiation, where populations from Greenland andthe USA (violaceus and latiglumis)were clearly separated from the others (hyperarcticus,komarovii and sajanensis), wasrevealed by both the cluster and principal coordinates analyses.     

Genetic diversity in cowpea [Vigna unguiculata (L.) Walp.] as revealed by RAPD markers

The present study, using RAPD analysis, was undertaken to characterize genetic variation in domesticated cowpea and its wild progenitor, as well as their relationships. The materials used consisted of 26 domesticated accessions, including accessions from each of the five cultivar-group, and 30 wild/weedy accessions, including accessions from West, East and southern Africa. A total of 28 primers generated 202 RAPD bands. One hundred and eight bands were polymorphic among the domesticated compared to 181 among wild/weedy cowpea accessions. Wild accessions were more diverse in East Africa, which is the likely area of origin of V. unguiculata var. spontanea. Var. spontanea is supposed to have spread westward and southward, with a loss of variability, loss counterbalanceed in southern Africa by introgressions with local perennial subspecies. Although the variabilty of domesticated cowpea was the highest ever recorded, cultivar-groups were poorly resolved, and several results obtained with isozyme data were not confirmed here. However primitive cultivars were more diverse than evolved cultivars, which still suggests two consecutive bottlenecks within domesticated cowpea evolution. As isozymes and AFLP markers, although with a larger number of markers, RAPD data confirmed the single domestication hypothesis, the gap between wild and domesticated cowpea, and the widespread introgression phenomena between wild and domesticated cowpea.     

Assessment of genetic relationships between <Emphasis Type="Italic">Rhus</Emphasis> L. species using RAPD markers

Genetic diversity of seven Rhus L. species was assessed using random amplified polymorphic DNA (RAPDs) markers. Initially, 90 primers were screened, of which 25 produced reproducible amplification products. These primers generated a total of 296 bands, with an average of 11.8 bands per primer. Out of 296 bands scored, 236 (80%) were polymorphic and 62 (20%) were monomorphic. Primers OPC-05 and OPD-05 generated 100% polymorphic bands. The resolving power of primers ranged from 9.4 to 26.8. Similarity matrix values ranged from 0.45 to 0.63. The dendrogram generated using Unweighted Pair Group Method using Arithmetic Averages (UPGMA) grouped all the species of Rhus in one major group with two sister groups, whilst R. pyroides Burch. and R. dentata Thunb. were outliers. R. gerrardii (Harv. ex Engl.) Diels, R. glauca Thunb. and R. pentheri Zahlbr. constituted one sister group, while R. natalensis Bernh. ex C. Krauss and R. gueinzii Sond. were included in the other. The degree of genetic diversity observed between seven species of Rhus with RAPD markers suggest that this approach could be used for studying the phylogeny of the genus.     

Characterization of genetic variation and relationships among Choix germplasm accessions using RAPD markers

Choix, a plant in the tribe Maydeae of the grass family, has been cultivated in Asia for several thousand years. It is a potential gene resource for improvement of other cereal crops because of its nutritional value and tolerance to stress. Genetic variation and relationships among 21 Choix lachryma-jobi L. accessions were characterized by random amplified polymorphic DNA (RAPD) markers. A total of 205 DNA fragments across all materials were amplified with 31 random primers, averaging 6.61 per primer. Among amplified fragments, 115 showed polymorphism averaging 3.71 per primer. Of amplified markers, 56.1% were polymorphic, indicating considerable variation at the DNA level among these accessions. Some fragments were accession-specific. Pair-wise genetic similarity (GS) among 21 accessions ranged from 0.809 to 0.301. The 21 accessions clustered into two major groups. Three exotic Choix accessions clustered together. Three other Choix accessions, collected from Guangxi, China, clustered into a cohesive subgroup. Four wild types of Choix clustered into the same subgroup. These results indicated that the classification by RAPD data reflected the differences in geographic origins and evolution in Choix.     

Characterization of under-utilized fruits by molecular markers. A case study of loquat

The usefulness of RAPD markers for genotyping a minor fruit species such as loquat has been tested in order to assess their ability for identifying accessions and to provide a set of markers suitable for use by different groups of scientists and curators. Twenty-nine polymorphic markers selected from a previous study of 33 accessions were tested in 46 new accessions added to the collection. Using the same PCR standard conditions, only 20 markers out of 29 selected in the previous study gave consistent amplifications in the new set of plant material. The rest required optimization of reaction conditions. This fact pointed out that RAPD markers were sensitive to the experimental conditions, hence a standard technique did not guarantee reproducibility. To overcome this problem markers for plant fingerprinting should be selected after comparison across accession sets. Only those markers reproducible with different sampling and checked in several sets of accessions are suitable for germplasm fingerprinting. In the present study we propose 31 RAPDs for fingerprinting loquat that accomplished these characteristics.The markers obtained were sufficient for determining origin and relationships of cultivars, for identifying synonyms and derived varieties from bud-sports. All bud sports were identical for all RAPDs selected.     

Studying the extent of genetic diversity among Gossypium arboreum L. genotypes/cultivars using DNA fingerprinting

Genetic diversity is an area of concern for sustaining crop yield. Information on genetic relatedness/diversity among Gossypium arboreum L. cultivars/genotypes is scanty. We have used random amplified polymorphic DNA (RAPD) analysis to assess the genetic divergence/relationship among 30 genotypes/cultivars of G. arboreum. Of 45 primers surveyed, 63% were polymorphic. Out of the total number of loci amplified, 36% were polymorphic. The calculated genetic similarity between the cultivars/genotypes was in the range of 47.05–98.73%. Two genotypes, HK-244 and Entry-17, were the most distantly related. The average genetic relatedness among all the genotypes was 80.46%. However, most of the cultivated varieties showed a close genetic relationship, indicating a narrow genetic base in comparison to the non-cultivated germplasm. The calculated coefficients were used to construct a dendrogram using the unweighted pair group of arithmetic means (UPGMA) algorithm, which grouped the genotypes/cultivars into two major and three smaller clusters. The study is the first comprehensive analysis of the genetic diversity of G. arboreum germplasm and identifies cultivars that will be useful in extending the genetic diversity of cultivated varieties and future genome mapping projects.     

Analysis of genetic diversity among European and Asian fig varieties (<Emphasis Type="Italic">Ficus carica</Emphasis> L.) using ISSR,RAPD, and SSR markers

Nineteen fig varieties and lines from Europe and Asia have been fingerprinted by ISSR, RAPD, and SSR markers, respectively, using 13, 19, and 13 primer combinations. All primers produced 258 loci, with the highest number of loci (119) generated by RAPD (R _p: 48.42). Clustering analysis was applied to the three marker datasets to elucidate the genetic structure and relationships among these varieties. Mean genetic similarities were 0.787, 0.717, and 0.749, respectively, as determined using ISSR, RAPD, and SSR. Each marker system produced incompletely separated clusters, although a weak binding group based on race type appeared in the combined dataset. Comparisons of coefficients revealed no correlation between different similarity matrices; congruence was observed between similarity matrices and co-phenetic matrices in all markers. Analysis of molecular variance (AMOVA) showed that most of the total polymorphism was attributable to within-group variance (ISSRs + RAPDs, 97.41%; SSRs, 90.18%). These results suggest that the genetic diversity of this fig population is low and that multiple marker utilization is critical to estimate the relatedness of figs at the variety level. Additionally, it was presumed that ‘Houraihi’, the oldest variety in Japan, was disseminated independently of other foreign varieties in the 17th century or before then.     

Genetic diversity in the Olive tree (Olea europaea L. subsp. europaea) cultivated in Portugal revealed by RAPD and ISSR markers

To assess the genetic diversity of the most important olive cultivars used in Portugal, a base collection was established with two hundred and one accessions of eleven cultivars from the different agro-ecological-regions (AER) of olive oil production. Inter-cultivar diversity was evaluated using seven RAPD primers producing fifty-nine polymorphic markers that enable cultivar distinction. Discriminant analysis according to fruit use and AER revealed a genetic structure associated with local selection both for fruit exploitation and agro-ecological adaptation. Intra-cultivar diversity of the ancient cultivar Galega was also investigated. Three RAPD and five ISSR primers produced ninety-three polymorphic markers upon seventy-seven accessions from five AERs. Total accession discrimination was achieved. UPGMA clustering and discriminant analysis revealed that the genetic diversity was predominantly structured according to accessions origin. The within and among AER variation revealed by AMOVA supported this genetic structure and showed a high proportion of intra-AER variability. These evidences suggest that Galega is composed by a mixture of different genotypes adapted to local conditions, indicating that this cultivar is in an early stage of domestication and should be treated as a landrace instead of a uniform cultivar. The assessment of Galega genetic diversity within each of the five AERs indicated the highest significant level (H_g = 6.23 at p< 0.001) in Ribatejo-Santarém. This finding associated with the distinctiveness of Galega in relation to other Portuguese cultivars and with the recent insights of olive tree domestication allowed us to hypothesize that Ribatejo-Santarém was the ecological region of origin and dispersion of this ancient cultivar.     

Detection of genetic diversity in Libyan wheat genotypesusing wheat microsatellite markers

Twenty-four wheat microsatellites (WMS) wereused to estimate the extent of genetic diversity among 15 Libyanwheat genotypes. The WMS used determined 26 loci located on 20different chromosomes, and were capable of detecting 116 alleles withan average of 4.5 alleles per locus. Only two markers located on 2DSand 4DL, were monomorphic. The results indicated that the B genome(5.9 alleles per locus) was more variable than the A and Dgenomes (4.1 and 2.7 alleles per locus, respectively).Furthermore, the results obtained suggest that a relatively smallnumber of primers can be used to distinguish all genotypes used andto estimate their genetic diversity. Genetic dissimilarity valuesbetween genotypes, calculated by the WMS derived data, were used toproduce a dendrogram. The diversity within the analysed germplasm isdiscussed.     

Genetic diversity in sweetpotato [Ipomoea batatas (L.) Lam.] in relationship to geographic sources as assessed with RAPD markers

Available evidence shows that sweetpotato originated from either Central or South American lowlands with subsequent dispersal to North America, Europe, Africa, Asia and the pacific islands. A total of 71 polymorphic RAPD molecular markers were used to assess the genetic relationships amongst 74 sweetpotato varieties originating from a total of 23 sweetpotato producing countries within six geographical regions, namely, South America, Central America/Caribbean, United States of America (USA), East Africa, Asia and Oceania. An Analysis of Molecular Variance (AMOVA) indicated that 93.4% of the total variance was due to the differences between genotypes within regions. The difference between regions was significant (P < 0.001) but only contributed 6.6% to the variance. Genetic distance (PhiST) calculated with AMOVA and multidimensional scaling (MDS) revealed that the South American and the Central American/Caribbean genotypes formed two separate clusters. East African varieties, which have unique characteristics from other traditional varieties, were distinct from other traditional varieties from South America and Oceania. These results support the reported hypothesis of the origin and dispersal of the sweetpotato and indicate that the primary centre of diversity probably has two distinct genepools. It is proposed that the dispersal of the sweetpotato from its origin may have mainly involved varieties from Central America/Caribbean as opposed to varieties from South America. There is an indication that new genepools may be evolving in Africa and Asia due to hybridisation and adaptation to the local environments.     

Low levels of genetic diversity detected by RAPD analysis in geographically distinct accessions of Bacopa monnieri

Brahmi (Bacopa monnieri) is atraditional Indian medicinal plant known for its natural nootropicaction of saponins present in large amount in its shoots. Acollection of 24 B.monnieri accessions from differentagro-climatic zones of India and an introduction from Malaysiamaintained in the field genebank at CIMAP was analysed for RAPDvariation. Among the 40 random primers tested, 29 primers generatedone or more polymorphic bands. The number of polymorphic bandsgenerated was primer dependent, ranging from 2 to maximum of 8.Similarity matrices were generated from the RAPD data on the basis ofNei's estimates of similarity indices and dendrograms wereconstructed based on UPGMA clustering. All the accessions were foundto be in the range of 0.8–1.0 of similarity, which isindicative of a narrow genetic base among the various accessions witha medium level of polymorphism. It was possible to differentiateindividual accessions, showing differences in morphological andgrowth properties at DNA level. The observed low levels of geneticvariation were attributed to interplay of sexual and vegetative modesof reproduction and similarity of local environments in habitats ofB. monnieri.