Preventive effect of Terminalia bellirica (Gaertn.) Roxb. extract on mice infected with Salmonella Typhimurium.

Terminalia bellirica (Gaertn.) Roxb. (TB) is a traditional herbal combination used in Chinese medicine for the treatment of a broad range of diseases. In this study, thirty KM mice were randomly divided into control (N), infection group (NS), and the TB protection group (HS). Based on its digestive feature, intestinal physical barrier, immunological barrier and gut microbiota effects in vivo on challenged with S.typhimurium mice were investigated after oral administration of 600 mg/kg b.wt of TB for 13 days. The results show that the extract could improve the level of serum immunoglobulins (IgA and IgG), decrease the intestinal cytokine secretion to relieve intestinal cytokine storm, reinforce the intestinal biochemical barrier function by elevating the sIgA expression, and strengthen the intestinal physical barrier function. Simultaneously, based on the V3-V4 region of the 16S rRNA analyzed, the results of the taxonomic structure of the intestinal microbiota demonstrated that the TB prevention effect transformed the key phylotypes of the gut microbiota in S. Typhimurium-challenged mice and promoted the multiplication of beneficial bacteria. Furthermore, the abundance of Firmicutes and Deferribacteres increased, while that of Bacteroidetes and Actinobacteria decreased. At the genus level, the abundance of Ruminococcus and Oscillospira was substantially enhanced, while the other dominant genera showed no significant change between the vehicle control groups and the TB prevention groups. In summary, these results provide evidence that the administration of TB extract can prevent S. Typhimurium infection by alleviating the intestinal physical and immunological barriers and normalizing the gut microbiota, highlighting a promising application in clinical treatment. Thus, our results provide new insights into the biological functions of TB for the preventive effect of intestinal inflammation.

The combination of ITS2 and psbA-trnH region is powerful DNA barcode markers for authentication of medicinal Terminalia plants from Thailand.

The dried fruits of Terminalia plant (Combretaceae) called "Samo" have been used as herbal medicine in Thai traditional medicine. Four "Samo" crude drugs, namely, Samo thai, Samo thed, Samo dee-ngu, and Samo phiphek, are used as the main ingredients in Triphala and Trisamo recipes. Their commercial products are available in processed and powdered form, but are difficult to authenticate by conventional methods. In this study, we aimed to discriminate species of genus Terminalia for the identification of their crude drugs by a DNA barcoding technique. A total of 208 closely related nucleotide sequences were obtained from nine Terminalia species collected from Thailand and the DDBJ/EMBL/GenBank database. An effective DNA barcode marker was selected from six DNA loci (matK, rbcL, psbA-trnH, ITS, ITS1, and ITS2) and their two-locus combination. All sequences were analyzed by three major methods: (1) BLAST search; (2) the genetic divergence method using Kimura 2-parameter (K2P) distance matrices; and (3) tree topology analysis based on the neighbor-joining method. Comparison of the six candidate DNA loci indicated that ITS identified Terminalia with 100% accuracy at the species and genus levels in the BLAST1 method. ITS2 showed the highest K2P variability. The data from the single markers and the two-locus combinations revealed that only the two-locus combinations, namely, the combinations of rbcL, ITS, ITS1, and ITS2 with psbA-trnH, clearly discriminated all the species. From the results of DNA sequence analysis and the three methods, ITS2 is recommended for the identification of Terminalia species to supplement psbA-trnH.

History of the fragmentation of the African rain forest in the Dahomey Gap: insight from the demographic history of Terminalia superba.

Paleo-environmental reconstructions show that the distribution of tropical African rain forests was affected by Quaternary climate changes. They suggest that the Dahomey Gap (DG)-the savanna corridor that currently separates Upper Guinean (UG, West Africa) and Lower Guinean (LG, western Central Africa) rain forest blocks-was forested during the African Humid Holocene period (from at least 9 ka till 4.5 ka), and possibly during other interglacial periods, while an open vegetation developed in the DG under drier conditions, notably during glacial maxima. Nowadays, relics of semi-deciduous forests containing UG and LG forest species are still present within the DG. We used one of these species, the pioneer tree Terminalia superba (Combretaceae), to study past forest fragmentation in the DG and its impact on infraspecific biodiversity. A Bayesian clustering analysis of 299 individuals genotyped at 14 nuclear microsatellites revealed five parapatric genetic clusters (UG, DG, and three in LG) with low to moderate genetic differentiation (Fst from 0.02 to 0.24). Approximate Bayesian Computation analyses inferred a demographic bottleneck around the penultimate glacial period in all populations. They also supported an origin of the DG population by admixture of UG and LG populations around 54,000 (27,600-161,000) years BP, thus before the Last Glacial Maximum. These results contrast with those obtained on Distemonanthus benthamianus where the DG population seems to originate from the Humid Holocene period. We discuss these differences in light of the ecology of each species. Our results challenge the simplistic view linking population fragmentation/expansion with glacial/interglacial periods in African forest species.

Molecular Analysis of Terminalia spp. Distributed in Thailand and Authentication of Crude Drugs from Terminalia Plants.

Terminalia, a large genus of Combretaceae, is distributed in Tropical Asia, Africa, and America. Some Terminalia plants are used in folk medicine because they possess powerful medicinal properties. Dried fruits of Terminalia bellirica and Terminalia chebula are used as the main ingredient in Triphala, a famous polyherbal formulation in Ayurvedic medicine and Thai folk medicine, because of their laxative, detoxifying, and rejuvenating effects. To clarify the phylogenetic relationships of medicinal Terminalia species (T. bellirica, T. chebula, and T. catappa) and authenticate their crude drugs, "Samo" and Triphala, nucleotide sequencing alignments in the internal transcribed spacer one-two (ITS 1-2) regions of Terminalia plants collected in Thailand were performed. The amplified fragments of Terminalia species were approximately 800 bp in length. To compare these sequences and DDBJ registered data, a molecular phylogenetic tree was constructed. Phylogenetic analysis clearly separated the sequences into two groups: Asian Terminalia and African Terminalia with some exceptions. In the analyzed sequences, the length of the ITS1-5.8S-ITS2 region was 674 bp in T. chebula, and 677 bp in T. bellirica and T. catappa. Eighty-one single nucleotide polymorphisms (SNPs) and nine insertion-deletions (indels) were observed, and the nucleotide sequences of this region showed species-specific sequences. Based on these differences, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system (ARMS) were applied to identify medicinal Terminalia species. Moreover, the ARMS method was chosen for fingerprinting analysis of Samo crude drugs and Triphala formulations because it was a fast, cost-effective, and reproducible approach.

In vitro propagation, encapsulation, and genetic fidelity analysis of Terminalia arjuna: a cardioprotective medicinal tree.

The present study described an improved and reproducible in vitro regeneration system for Terminalia arjuna using nodal segment explants obtained from a mature plant. Shoot tips excised from in vitro proliferated shoots were encapsulated in 3 % sodium alginate and 100 mM CaCl2[Symbol: see text]2H2O for the development of synthetic seeds which may be applicable in short-term storage and germplasm exchange of elite genotype. Shoot multiplication was significantly influenced by a number of factors, namely types and concentrations of plant growth regulators, medium composition, repeated transfer of mother explants, subculturing of in vitro regenerated shoot clumps, agar concentrations, and temperature. Maximum numbers of shoots (16.50 ± 3.67) were observed on modified Murashige and Skoog (MMS) medium containing 0.5 mg l(-1) of benzylaminopurine (BAP) and 0.1 mg l(-1) of naphthalene acetic acid (NAA). To shortening the regeneration pathway, rooting of micropropagated shoots under in vitro condition was excluded and an experiment on ex vitro rooting was conducted and it was observed that the highest percentage of shoots rooted ex vitro when treated with indole-3-butyric acid (IBA, 250 mg l(-1)) + 2-naphthoxy acetic acid (NOA, 250 mg l(-1)) for 5 min. The well-developed ex vitro rooted shoots were acclimatized successfully in soilrite under greenhouse conditions with 80 % survival of plants. Randomly amplified polymorphic DNA (RAPD) analysis confirmed that all the regenerated plants were genetically identical to the mother plant, suggesting the absence of detectable genetic variation in the regenerated plantlets. To the best of our knowledge, this is the first report on synthetic seed production as well as ex vitro rooting and genetic fidelity assessment of micropropagated shoots of T. arjuna.

Estimation of genetic diversity and evaluation of relatedness through molecular markers among medicinally important trees: Terminalia arjuna, T. chebula and T. bellerica.

Terminalia trees are being over-exploited because of their medicinal and economical importance leading to loss of valuable genetic resources. For sustainable utilization and conservation, assessment of genetic diversity therefore becomes imperative. We report a comprehensive first study on estimation and analysis of genetic variation through Amplified fragment length polymorphism (AFLP), inter simple sequence repeat polymorphism (ISSR) and random amplification of polymorphic DNA (RAPD) across three species of Terminalia. The study included (i) characterization of genetic diversity at interspecific level, and (ii) comparison of efficiency of the marker systems. That the three species are genetically distinct was revealed by all the three marker systems as unique DNA fingerprints were obtained. This led to identification of several species-specific amplification products. Further analysis helped in species-wise clustering. The species specific bands obtained from the present investigation can be used as diagnostic markers to identify the raw materials for herbal drug preparations for authentication purposes.

Phylogeographic structure of Terminalia franchetii (combretaceae) in southwest China and its implications for drainage geological history.

Following the rapid uplift of the Qinghai-Tibetan Plateau, the reorganization of the major river drainages in southwest China was primarily caused by river capture events. However, the impact of these past changes in drainage patterns on the current distribution and genetic structure of the endemic flora of this region remains largely unknown. Here we report a survey of amplified fragment length polymorphism (AFLP) in Terminalia franchetii, an endemic shrub or small tree of the deep and dry-hot river valleys of this region. We surveyed AFLP variation within and among 21 populations (251 individuals) of T. franchetii, distributed disjunctively between northern and southern drainage systems. Using STRUCTURE, principal coordinates analysis, and genetic distance methods, we identified two main population genetic groups (I and II) and four subgroups within the species, as follows: (I) the Upper Jinshajiang Valley (subgroup I((north))) and the Honghe drainage area (subgroup I((south))); (II) the Middle and Lower Jinshajiang and Yalongjiang Valleys (subgroup II((north))) and the Nanpanjiang drainage area (subgroup II((south))). Genetic diversity was lower in group I than in group II. According to the genetic diversity and genetic structure results, we suggest that the modern disjunctive distribution and associated patterns of genetic structure of T. franchetii result from vicariance caused by several historical drainage capture events, involving the separation of the Upper Jinshajiang, Yalongjiang and Daduhe from the Honghe or Nanpanjiang in southwest China.

A standardized protocol for genomic DNA isolation from Terminalia arjuna for genetic diversity analysis

For studying genetic diversity in natural populations of Terminalia, a medicinal plant, our attempts to isolate high quality DNA using several previously reported protocols and even modifications were unsuccessful. We therefore combined CTAB based isolation, and column based purification step, to isolate DNA from Terminalia arjuna. The DNA isolated using this standardized protocol was high in quality and suitable for restriction digestion and generation of random amplification of polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP).

High-resolution mapping and chromosome landing at the root-know nematode resistance locus Ma from Myrobalan plum using a large-insert BAC DNA library.

The Ma gene for root-knot nematode (RKN)resistance from Myrobalan plum (Prunus cerasifera L.)confers a complete-spectrum and a heat-stable resistance to Meloidogvne spp., conversely to Mi-I from tomato,which has a more restricted spectrum and a reduced efficiency at high temperature. This gene was identified from a perennial self-incompatible near-wild rootstock species and lies in cosegregation with the SCAR marker SCAFLP2 on the Prunus linkage group 7 in a 2.3 cM interval between the SCAR SCAL19 and SSR pchgms6 markers. We initiated a map-based cloning of Ma and report here the strategy that rapidly led to fine mapping and direct chromosome landing at the locus. Three pairs of bulks, totaling 90 individuals from half-sibling progenies derived from the Ma-heterozygous resistant accession P.2175, were constructed using mapping data, and saturation of the Ma region was performed by bulked segregant analysis (BSA) of 320 AFLP primer pair combinations. The closest three AFLP markers were transformed into codominant SCARs or CAPS designatedSCAFLP3, SCAFLP4 and SCAFLP5. By completing the mapping population up to 1,332 offspring from P.2175,Ma and SCAFLP2 were mapped in a 0.8 cM interval between SCAFLP3 and SCAFLP4. A large-insert bacterial artificial chromosome (BAC) DNA library of P.2175,totaling 30,720 clones with a mean insert size of 145 kb and a 14-15x Prunus haploid genome coverage was constructed and used to land on the Ma spanning interval with few BAC clones. As P.2175 is heterozygous for the gene, we constructed the resistant and susceptible physical contigs by PCR screening of the library with codominant markers. Additional microsatellite markers were then designed from BAC subcloning or BAC end sequencing. In the resistant contig, a single 280 kb BAC clone was shown to carry the Ma gene; this BAC contains two flanking markers on each side of the gene as well as two cosegregating markers. These results should allow future cloning of the Ma gene in this perennial species.

Genetic diversity of the tropical tree Terminalia amazonia (Combretaceae) in naturally fragmented populations.

The effect of long-term fragmentation on the genetic diversity of populations of the neotropical tree species, Terminalia amazonia, was studied using random amplified polymorphic DNA (RAPD) analysis. Leaf material from 104 trees was collected from three naturally fragmented gallery forest patches and three plots in nearby continuous forest in the Mountain Pine Ridge, Belize. In total, 30 RAPD bands generated by five decamer primers were used to compare the genetic diversity of the six populations in the two groups. Genetic variation within the populations (H0), as estimated by the Shannon diversity index, ranged from 0.32 to 0.38, with an overall diversity of 0.38 (Hspecies). Analysis of molecular variation revealed that most (94.4%, P<0.001) of the variation was attributable to differences among individuals within populations. Population differentiation was significantly (P=0.038) lower among the fragmented populations than among continuous forest populations. On average, the fragmented populations also had slightly, but statistically significant (P=0.046) lower levels of genetic diversity. However, one gallery forest site had a higher level of genetic diversity than two of the continuous forest sites. We suggest that the long-term effect of fragmentation on the genetic diversity of tropical trees will depend upon the amount of local forest cover in proximity to the fragmented populations.