Genetic diversity of Eurycoma longifolia inferred from single nucleotide polymorphisms

Eurycoma longifolia Jack. is a treelet that grows in the forests of Southeast Asia and is widely used throughout the region because of its reported medicinal properties. Widespread harvesting of wild-grown trees has led to rapid thinning of natural populations, causing a potential decrease in genetic diversity among E. longifolia. Suitable genetic markers would be very useful for propagation and breeding programs to support conservation of this species, although no such markers currently exist. To meet this need, we have applied a genome complexity reduction strategy to identify a series of single nucleotide polymorphisms (SNPs) within the genomes of several E. longifolia accessions. We have found that the occurrence of these SNPs reflects the geographic origins of individual plants and can distinguish different natural populations. This work demonstrates the rapid development of molecular genetic markers in species for which little or no genomic sequence information is available. The SNP markers that we have developed in this study will also be useful for identifying genetic fingerprints that correlate with other properties of E. longifolia, such as high regenerability or the appearance of bioactive metabolites.     

Effects of immunomodulatory drugs on plasma inflammatory markers in a rabbit model of atherosclerosis

Atherosclerosis is a chronic disease of the arterial wall where both innate and adaptive immuno-inflammatory mechanisms are involved. Inflammatory cytokines are implicated in the development and progression of atherosclerotic lesions. Immunomodulatory therapies have been proposed for the treatment of atherosclerosis. Therefore, the aim of this study was to investigate the systemic anti-inflammatory and immunomodulatory effects of atorvastatin, cyclosporine A (CsA), and tacrolimus (FK506) on plasma inflammatory markers in atherosclerotic rabbits. Male New Zealand rabbits were randomized into five groups each of 12 animals. Standard diet-fed group served as control, and the cholesterol-fed group received a diet supplemented with 1% cholesterol alone, cholesterol + atorvastatin, cholesterol + FK506, and cholesterol + CsA. Serum levels of lipid profile parameters (triglycerides, cholesterol, and high-density lipoprotein) were measured using colorimetric methods. Serum levels of C-reactive protein (CRP), interleukin-6 (Il-6), and interferon-gamma (INF-γ) were measured in all studied groups using ELISA techniques. Our results revealed a significant decrease (p < 0.001) in the serum levels of lipid profile parameters, CRP, Il-6, and INF-γ in atorvastatin-treated group compared with the cholesterol-fed group. On the other hand, a non-significant difference was observed for the same parameters in either FK506- or CsA-treated groups compared with the cholesterol-fed group. In conclusion, atorvastatin has a systemic anti-inflammatory role that far surpassed the cholesterol reduction effect alone. FK506 or CsA failed to suppress elevated plasma inflammatory markers. Thus, low doses of these two immunomodulating drugs could not have generalized systemic anti-inflammatory or immunosuppressive effects.     

Immunogenicity of synthetic peptides derived from the sequences of a Streptococcus mutans cell surface antigen in nonhuman primates

The immunogenicity and antigenicity of synthetic peptides (SP) derived from the sequences of a cell surface Ag of Streptococcus mutans were investigated in macaque monkeys. Immunization with the free peptides of 11, 17, and 21 residues failed to elicit serum antibodies or T cell responses. However, immunization with the SP17 and SP21 linked to tetanus toxoid (TT) as a carrier elicited serum antibodies and proliferative responses of lymphocytes, not only to the SP but also to the native streptococcal Ag. In vivo recall of SP-TT immunized monkeys with suboptimal doses of the native streptococcal Ag resulted in a significant increase in antibodies, both to the SP and the streptococcal Ag, confirming that the SP shares antigenic epitopes with the native Ag. B and T cell epitopes were then determined and a B cell epitope was found in residues 8-13, whereas an overlapping T cell epitope was located in residues 7-15. The T cell epitope has an amino-terminal leucine and carboxy-terminal glycine and alanine added to residues 8-13 of the B cell epitope. In spite of the B and T cell epitopes being expressed in SP17 (residues 1-15), the monomer failed to induce serum antibodies without a carrier. However, immunization with a dimer of SP17 elicited both serum antibodies and proliferative responses of lymphocytes without a carrier. The results suggest that the monomeric SP17 is not immunogenic and needs to be dimerised in order to elicit antibodies and T cell responses, both to the SP and to the streptococcal Ag.     

Microstructures,interactions and dynamics properties studies of N-methyldiethanolamine + guanidinium triflate ionic liquid + water tertiary system at the standard temperature

Molecular dynamics simulations with an all-atom force field have been carried out in order to understand the phase equilibrium behaviour of ternary aqueous mixtures containing guanidinium triflate ionic liquid [gua][OTf] and water mixed with N-methyldiethanolamine (MDEA) in different function composition at the standard temperature of 298.15 K. A very good numerical agreement has been obtained for the prediction of the mixture densities. The analysis of structural and dynamic properties showed that the molecular level of ternary mixtures is slightly affected by the presence of MDEA and [gua][OTf] molar fractions. For MDEA–water interactions in [gua][OTf] media, we found that MDEA prefers to be surrounded by water molecules rather than by MDEA molecules even at a high MDEA molar fraction. While for [gua][OTf]–water interaction in MDEA media, as [gua][OTf] molar fraction increases, water molecules replace counterions in the coordination shell of both ions, thus weakening their interaction. On the other hand, for MDEA–[gua][OTf] interactions in water media, we have found that as the molar fraction of [gua][OTf] increases, a sulfonate group from anion appears to have a stronger association by making hydrogen bonding with MDEA molecules. The chemical process using ionic liquids (ILs) as solvents is commonly limited by their high viscosity. Based on their physical properties such as viscosities, these ternary solvents can be applied in natural gas industry, such as removing carbon dioxide using aqueous MDEA and IL at high pressure.     

Microchirita ruthiae sp. nov. (Gesneriaceae) from Peninsular Malaysia

A new species Microchirita ruthiae Rafidah (Gesneriaceae) from limestone in Kelantan, Peninsular Malaysia, is described and illustrated. This new species is characterized by having a branched stem, inflorescences with 1–4 flowers, the flowers with a white corolla (rarely faintly tinged with pale purple), glistening pale brown glandular hairs inside the throat, free anthers, and hairy capsules. Its conservation status following IUCN categories and criteria is assessed as ‘Endangered’ (EN).     

Modulation of ethanol toxicity by Asian ginseng (Panax ginseng) in Japanese ricefish (Oryzias latipes) embryogenesis

Alcohol consumption by women during pregnancy often induces fetal alcohol spectrum disorder (FASD) in children who have serious central nervous system (CNS), cardiovascular, and craniofacial defects. Prevention of FASD, other than women abstaining from alcohol drinking during pregnancy, is not known. A limitation of the use of synthetic anti-alcoholic drugs during pregnancy led us to investigate herbal products. In particular, many plants including Asian ginseng (Panax ginseng) have therapeutic potential for the treatment of alcoholism. We used Japanese ricefish (medaka) (Oryzias latipes), an animal model of FASD, for identifying herbal medicines that can attenuate ethanol toxicity. Fertilized eggs in standard laboratory conditions were exposed to ginseng (PG) root extract (0–2 mg/mL) either 0–2 (group A) or 1–3 (group B) day post fertilization (dpf) followed by maintenance in a clean hatching solution. The calculated IC₅₀ as determined 10 dpf in A and B groups were 355.3 ± 1.12 and 679.7 ± 1.6 μg/mL, respectively. Simultaneous exposure of embryos in sub-lethal concentrations of PG (50–200 μg/mL) and ethanol (300 mM) for 48 h disrupted vessel circulation and enhanced mortality. However, PG (100 μg/mL) may partially protect trabecular cartilage (TC) deformities in the neurocranium in B group embryos induced by ethanol (300 mM). To understand the mechanism, embryonic ethanol concentration was measured at 2 dpf and adh5, adh8, aldh2, aldh9a, catalase, GST, and GR mRNAs were analyzed at 6 dpf. It was observed that although ethanol is able to reduce adh8 and GST mRNA contents, the simultaneous addition of PG was unable to alter ethanol level as well as mRNA contents in these embryos. Therefore, antagonistic effects of PG on ethanol toxicity are mediated by a mechanism which is different from those regulating ethanol metabolism and oxidative stress.     

Development of a Real-Time Microchip PCR System for Portable Plant Disease Diagnosis

Rapid and accurate detection of plant pathogens in the field is crucial to prevent the proliferation of infected crops. Polymerase chain reaction (PCR) process is the most reliable and accepted method for plant pathogen diagnosis, however current conventional PCR machines are not portable and require additional post-processing steps to detect the amplified DNA (amplicon) of pathogens. Real-time PCR can directly quantify the amplicon during the DNA amplification without the need for post processing, thus more suitable for field operations, however still takes time and require large instruments that are costly and not portable. Microchip PCR systems have emerged in the past decade to miniaturize conventional PCR systems and to reduce operation time and cost. Real-time microchip PCR systems have also emerged, but unfortunately all reported portable real-time microchip PCR systems require various auxiliary instruments. Here we present a stand-alone real-time microchip PCR system composed of a PCR reaction chamber microchip with integrated thin-film heater, a compact fluorescence detector to detect amplified DNA, a microcontroller to control the entire thermocycling operation with data acquisition capability, and a battery. The entire system is 25×16×8 cm³ in size and 843 g in weight. The disposable microchip requires only 8-µl sample volume and a single PCR run consumes 110 mAh of power. A DNA extraction protocol, notably without the use of liquid nitrogen, chemicals, and other large lab equipment, was developed for field operations. The developed real-time microchip PCR system and the DNA extraction protocol were used to successfully detect six different fungal and bacterial plant pathogens with 100% success rate to a detection limit of 5 ng/8 µl sample.     

A preliminary study on the effects of E-Z Mixin® and EquiPlus® extenders supplemented with Edible Bird’s Nest on the quality of chilled Arabian stallion semen

The aim of this study was to evaluate the effects of adding different concentrations of edible bird’s nest (EBN) which is secreted by swiftlet birds (Aerodramus fuciphagus), into EquiPlus® and E-Z Mixin® extenders on the quality of chilled Arabian stallion semen at various storage times (0, 24 and 48 h). Ten ejaculates were collected from five stallions, and diluted using the two extenders containing 0% (control), 0.12%, 0.24% and 0.24% of EBN + seminal plasma (SP). All the diluted semen samples were then cooled and stored at 5 °C, and examined at 0, 24 and 48 h. Sperm kinetic parameters were assessed using computer assisted sperm analysis (CASA) and viability were assessed using Hoechst33342/PI stain. In both extenders, total motility (TM) and progressive motility (PM) were significantly higher at 0.12% and 0.24% compared to 0.24% + SP at 24 and 48 h. At 0.12%, E-Z mixin® treated semen had significantly higher TM and PM than EquiPlus^® at 24 and 48 h. At 0.12% and 0.24%, average path velocity (VAP), straight-line velocity (VSL) and curvilinear velocity (VCL) were significantly higher in E-Z mixin® treated semen compared to EquiPlus^® at 24 and 48 h. Comparisons between the two extender types at different concentrations of EBN showed no significant difference in lateral head amplitude (ALH), linearity (LIN), straightness (STR), beat cross frequency (BCF) and viability, irrespective of the storage time. The percentage of viable was significantly higher in E-Z mixin® than EquiPlus® at 0 and 48 h in control and 0.12%. Supplementation of the E-Z mixin^® extender with 0.12% and 0.24% EBN concentrations in the absence of SP provided better CASA parameters such as TM, PM, VAP, VSL, and VCL at 24 and 48 h storage time. In conclusion, the results of this study indicated that chilled semen from Arabian stallion that was extended using E-Z mixin® and supplemented with 0.12% and 0.24% EBN concentrations performed better and yielded superior results in sperm kinetic parameters and % viable compared to EquiPlus® at 24 and 48 h storage time.