Antimalarials and Phytotoxins from Botryosphaeria dothidea Identified from a Seed of Diseased Torreya taxifolia

The metabolic pathways in the apicoplast organelle of Plasmodium parasites are similar to those in plastids in plant cells and are suitable targets for malaria drug discovery. Some phytotoxins released by plant pathogenic fungi have been known to target metabolic pathways of the plastid; thus, they may also serve as potential antimalarial drug leads. An EtOAc extract of the broth of the endophyte Botryosphaeria dothidea isolated from a seed collected from a Torreya taxifolia plant with disease symptoms, showed in vitro antimalarial and phytotoxic activities. Bioactivity-guided fractionation of the extract afforded a mixture of two known isomeric phytotoxins, FRT-A and flavipucine (or their enantiomers, sapinopyridione and (-)-flavipucine), and two new unstable γ-lactam alkaloids dothilactaenes A and B. The isomeric mixture of phytotoxins displayed strong phytotoxicity against both a dicot and a monocot and moderate cytotoxicity against a panel of cell lines. Dothilactaene A showed no activity. Dothilactaene B was isolated from the active fraction, which showed moderate in vitro antiplasmodial activity with high selectivity index. In spite of this activity, its instability and various other biological activities shown by related compounds would preclude it from being a viable antimalarial lead

Applying an internal transcribed spacer as a single molecular marker to differentiate between Tetraselmis and Chlorella species

In the realm of applied phycology, algal physiology, and biochemistry publications, the absence of proper identification and documentation of microalgae is a common concern. This poses a significant challenge for non-specialists who struggle to identify numerous eukaryotic microalgae. However, a promising solution lies in employing an appropriate DNA barcoding technique and establishing comprehensive databases of reference sequences. To address this issue, we conducted a study focusing on the molecular characterization and strain identification of Tetraselmis and Chlorella species, utilizing the internal transcribed spacer (ITS) barcode approach. By analyzing the full nuclear ITS region through the Sanger sequencing approach, we obtained ITS barcodes that were subsequently compared with other ITS sequences of various Tetraselmis and Chlorella species. To ensure the reliability of our identification procedure, we conducted a meticulous comparison of the DNA alignment, constructed a phylogenetic tree, and determined the percentage of identical nucleotides. The findings of our study reveal the significant value of the ITS genomic region as a tool for distinguishing and identifying morphologically similar chlorophyta. Moreover, our results demonstrate that both the ITS1 and ITS2 regions are capable of effectively discriminating isolates from one another; however, ITS2 is preferred due to its greater intraspecific variation. These results underscore the indispensability of employing ITS barcoding in microalgae identification, highlighting the limitations of relying solely on morphological characterization

Optimized construction of microsatellite-enriched libraries

The construction of microsatellite-enriched libraries is an indispensable tool to search for molecular markers as complete genome sequences are still not available for the majority of species of interest. Numerous protocols are available in the literature for the construction of these libraries; however, sometimes their low efficiency or lack of optimization in the protocols can restrict their efficacy. We have designed and tested various adapters and ligation methods; we also tested oligo-repeat combinations and hybridization temperatures, and created libraries with this new protocol for four organisms: Ipomoea batatas (L.) Lam, Chionanthus retusus Lindley & Paxton, Rotylenchulus reniformis Linford & Olivera and Puccinia kuehnii W. Kruger. The number of microsatellites detected for these species ranged from 2494 to 3919 per Mb of nonredundant sequence, that was 0.86 and 1.53 microsatellites per contig, with 37–66% of di-nucleotide motifs and 21–49% of tri- to octa-nucleotide repeats combined. A simplified protocol is provided for the successful generation of SSR-enriched libraries

Antiplasmodial and cytotoxic cytochalasins from an endophytic fungus, Nemania sp. UM10M, Isolated from a Diseased Torreya taxifolia Leaf

© 2019 American Chemical Society and American Society of Pharmacognosy. Cranberry (Vaccinium macrocarpon) juice is traditionally used for the prevention of urinary tract infections. Human urine produced after cranberry juice consumption can prevent Escherichia coli adhesion, but the antiadhesive urinary metabolites responsible have not been conclusively identified. Adult female sows were therefore fed spray-dried cranberry powder (5 g/kg/day), and urine was collected via catheter. Urine fractions were tested for antiadhesion activity using a human red blood cell (A+) anti-hemagglutination assay with uropathogenic P-fimbriated E. coli. Components were isolated from fractions of interest using Sephadex LH-20 gel filtration chromatography followed by HPLC on normal and reversed-phase sorbents with evaporative light scattering detection. Active urine fractions were found to contain a complex series of oligosaccharides but not proanthocyanidins, and a single representative arabinoxyloglucan octasaccharide was isolated in sufficient quantity and purity for full structural characterization by chemical derivatization and NMR spectroscopic methods. Analogous cranberry material contained a similar complex series of arabinoxyloglucan oligosaccharides that exhibited antiadhesion properties in preliminary testing. These results indicate that oligosaccharides structurally related to those found in cranberry may contribute to the antiadhesion properties of urine after cranberry consumption

Optimized construction of microsatellite-enriched libraries

The construction of microsatellite-enriched libraries is an indispensable tool to search for molecular markers as complete genome sequences are still not available for the majority of species of interest. Numerous protocols are available in the literature for the construction of these libraries; however, sometimes their low efficiency or lack of optimization in the protocols can restrict their efficacy. We have designed and tested various adapters and ligation methods; we also tested oligo-repeat combinations and hybridization temperatures, and created libraries with this new protocol for four organisms: Ipomoea batatas (L.) Lam, Chionanthus retusus Lindley & Paxton, Rotylenchulus reniformis Linford & Olivera and Puccinia kuehnii W. Kruger. The number of microsatellites detected for these species ranged from 2494 to 3919 per Mb of nonredundant sequence, that was 0.86 and 1.53 microsatellites per contig, with 37–66% of di-nucleotide motifs and 21–49% of tri- to octa-nucleotide repeats combined. A simplified protocol is provided for the successful generation of SSR-enriched libraries

Antiplasmodial and Cytotoxic Cytochalasins from an Endophytic Fungus, <i>Nemania</i> sp. UM10M, Isolated from a Diseased <i>Torreya taxifolia</i> Leaf

Bioassay-guided fractionation of an EtOAc extract of the broth of the endophytic fungus Nemania sp. UM10M (Xylariaceae) isolated from a diseased Torreya taxifolia leaf afforded three known cytochalasins, 19,20-epoxycytochalasins C (1) and D (2), and 18-deoxy-19,20-epoxy-cytochalasin C (3). All three compounds showed potent in vitro antiplasmodial activity and phytotoxicity with no cytotoxicity to Vero cells. These compounds exhibited moderate to weak cytotoxicity to some of the cell lines of a panel of solid tumor (SK-MEL, KB, BT-549, and SK-OV-3) and kidney epithelial cells (LLC-PK11). Evaluation of in vivo antimalarial activity of 19,20-epoxycytochalasin C (1) in a mouse model at 100 mg/kg dose showed that this compound had weak suppressive antiplasmodial activity and was toxic to animals

Essential Oils of Five Baccharis Species: Investigations on the Chemical Composition and Biological Activities

This paper provides a comparative account of the essential oil chemical composition and biological activities of five Brazilian species of Baccharis (Asteraceae), namely B. microdonta, B. pauciflosculosa, B. punctulata, B. reticularioides, and B. sphenophylla. The chemical compositions of three species (B. pauciflosculosa, B. reticularioides, and B. sphenophylla) are reported for the first time. Analyses by GC/MS showed notable differences in the essential oil compositions of the five species. &alpha;-Pinene was observed in the highest concentration (24.50%) in B. reticularioides. Other major compounds included &alpha;-bisabolol (23.63%) in B. punctulata, spathulenol (24.74%) and kongol (22.22%) in B. microdonta, &beta;-pinene (18.33%) and limonene (18.77%) in B. pauciflosculosa, and &beta;-pinene (15.24%), limonene (14.33%), and spathulenol (13.15%) in B. sphenophylla. In vitro analyses for antimalarial, antitrypanosomal, and insecticidal activities were conducted for all of the species. B. microdonta and B. reticularioides showed good antitrypanosomal activities; B. sphenophylla showed insecticidal activities in fumigation bioassay against bed bugs; and B. pauciflosculosa, B. reticularioides, and B. sphenophylla exhibited moderate antimalarial activities. B. microdonta and B. punctulata showed cytotoxicity. The leaves and stems of all five species showed glandular trichomes and ducts as secretory structures. DNA barcoding successfully determined the main DNA sequences of the investigated species and enabled authenticating them