Aqueous extracts prepared from the powdered entire pomaces and their parts (seeds and peels) of two Iraqi grape cultivars (Shada and Des-Alanez) were evaluated for their ovicidal, larvicidal, and pupicidal activities against housefly, Musca domestica (Linnaeus 1758) and compared them with chemical insecticide (Agita 10 WG; Austria). The highest insecticidal activity was shown by the aqueous extracts of powdered seeds followed by the entire pomace and then peels. The water extracts from the seeds, peels and pomaces were able to inhibit completely the hatching of the eggs at a concentration of 150 μg/ml while the Agita insecticide was able to inhibit the hatching process at a concentration of 50 μg/ml. The water extracts also were able to kill the L3 larvae and the activity was increasing with the increasing of the concentration. It is very interesting to note that the addition of the powdered entire pomaces, seeds, and peels on top or at the bottom of the cow manure was able to inhibit the development of up to 50% and 47.5% of L1 to pupae and to adult flies, respectively. Taking into consideration the huge amounts of grape pomaces that produced annually, the cheap price and the ease in dealing with, make them a good candidate as alternative insecticides and environmentally more friendly than the chemical insecticides.
Kaempferia parviflora is an ethnomedicinally important plant. Conventional propagation of K. parviflora is hindered by slow growth rate, long dormancy periods and dual use of rhizomes for seeds as well as marketable produce. In our study, we developed a promising dual-phase micropropagation protocol to increase number of plantlets, survivability, biomass and quality plantlets for mass production. Multiple shoot regeneration was found most successful on Murashige and Skoog (MS) media supplemented with 35.52 μM N6-benzyladenine (BA) in terms of highest number of shoots (22.4 ± 1.84), leaves (29.27 ± 1.30), and roots (17.8 ± 1.72) per explant. High survivability was observed with an acclimatisation percentage of 100% in sterile perlite medium. This method was shown to be preferable compared to conventional propagation in terms of propagation time and number of plantlets. Regenerated in vitro plantlets were then successfully induced to form microrhizomes in MS media with an optimal concentration of 6% (w/v) sucrose. Increase in microrhizome biomass (35.7 ± 2.59 g per flask), number of microrhizomes (5.2 ± 0.78), shoots (8.5 ± 1.58) and roots (8.5 ± 1.58) were observed for this treatment. This investigation successfully highlights the manipulation of single factors in short time frame to produce a simple and efficient alternative propagation method for K. parviflora.
The production of nitrogenase enzyme and auxins by free living diazotrophs has the potential to influence the growth of host plants. In this study, diazotrophs were grown in the presence of various concentrations of nitogen (N) to determine the optimal concentration of N for microbial growth stimulation, promotion of gaseous N (N2) fixation, and phytohormone production. Therefore, we investigate whether different levels of N supplied to Herbaspirillum seropedicae (Z78) have significant effects on nitrogenase activity and auxin production. The highest nitrogenase activity and the lowest auxin production of H. seropedicae (Z78) were both recorded at 0 gL(-1) of NH4Cl. Higher levels of external N caused a significant decrease in the nitrogenase activity and an increased production of auxins. In a subsequent test, two different inoculum sizes of Z78 (10(6) and 10(12) cfu/ml) were used to study the effect of different percentages of acetylene on nitrogenase activity of the inoculum via the acetylene reduction assay (ARA). The results showed that the optimal amount of acetylene required for nitrogenase enzyme activity was 5% for the 10(6) cfu/ml inoculum, whereas the higher inoculum size (10(12) cfu/ml) required at least 10% of acetylene for optimal nitrogenase activity. These findings provide a clearer understanding of the effects of N levels on diazotrophic nitrogenase activity and auxin production, which are important factors influencing plant growth.
Heterologous prime-boost immunisation strategies can evoke powerful antibody responses and may be of value in developing an improved malaria vaccine. Herein, we show that an immunisation protocol that primes Balb/c mice with a recombinant Bacille Calmette-Guérin (rBCG) vaccine consisting of a plasmid encoding a synthetic fragment of the ESAT-6 epitope of Mycobacterium tuberculosis, the fragment 2 region II of erythrocyte-binding antigen (F2RIIEBA) and the three repeat sequences of the circumsporozoite protein (NANP)3 of Plasmodium falciparum before subsequently boosting the mice with either two doses of the rBCG clone or with a DNA vaccine expressing the native form of F2RIIEBA generating higher serum anti-F2RIIEBA antibody levels than an immunisation protocol that calls for a homologous prime-boost with two doses of rBCG. These results demonstrate the potential of DNA vaccination in boosting the antibody response to a recombinant vaccine expressing multiple epitopes.
Treatment with hypovirulent binucleate Rhizoctonia (HBNR) isolates induced systemic resistance against anthracnose infected by Colletotrichum orbiculare in cucumber, as there were no direct interaction between HBNR and C. orbiculare. This is because of the different distances between HBNR and C. orbiculare, where the root was treated with HBNR isolate and C. orbiculare was challenged and inoculated in leaves or first true leaves were treated with HBNR isolate and C. orbiculare was challenged and inoculated in second true leaves. The use of barley grain inocula and culture filtrates of HBNR significantly reduced the lesion diameter compared to the control (p = 0.05). The total lesion diameter reduction by applying barley grain inoculum of HBNR L2, W1, W7, and Rhv7 was 28%, 44%, 39%, and 40%, respectively. Similar results was also observed in treatment using culture filtrate, and the reduction of total lesion diameter by culture filtrate of HBNR L2, W1, W7, and Rhv7 was 45%, 46%, 42%, and 48%, respectively. When cucumber root was treated with culture filtrates of HBNR, the lignin was enhanced at the pathogen penetration, which is spread along the epidermis tissue of cucumber hypocotyls. Peroxidase activity in hypocotyls in the treated cucumber plant with culture filtrates of HBNR significantly increased before and after inoculation of pathogens as compared to the control. Significant enhancement was also observed in the fast-moving anodic peroxidase isozymes in the treated plants with culture filtrates of HBNR. The results showed the elicitor(s) contained in culture filtrates in HBNR. The lignin deposition as well as the peroxidase activity is an important step to prevent systemically immunised plants from pathogen infection.
Sponge-associated bacteria are considered a rich source of bioactive compounds particularly to reduce the risk of Vibrio harveyi and Vibrio parahaemolyticus infection. The present study aimed to analyse the effectiveness of 19 isolates to control Vibrio infection in vivo. All 19 isolates displayed a non-pathogenic characteristic on shrimps (cell density of 106 cells/mL) as analysed using the pathogenicity test. The mortality caused by both Vibrio spp. on 50% of the shrimp population (LC50 value) had a cell density of 105 cells/mL as determined using the proportion interval method. On the basis of the challenge test, all isolates improved the survival rate of infected shrimps in diverse effectivities up to 89%, which was nearly 30% higher than the infected control. Two isolates coded as D6.9, and P5.20 reduced shrimp mortality after infection with Vibrio spp. 16S rRNA-based identification showed these isolates were closely similar to different genera of Bacillus and Staphylococcus. The extract derived from the most prospective isolate, D6.9, was dominated by 1-hydroxy-6-(3-isopropenyl-cycloprop-1-enyl)-6-methyl-heptan-2-one, hexadecanoic acid, 4-epicyclomusalenone [(24S)-24-methyl-28-norcycloart-25-en-3-one], and 2,4-dimethyl acetoacetanilide. This observation suggested these isolates characterised by in vivo anti-Vibrio activity need to be further developed as biocontrol candidates.
Gastrointestinal nematode infections can cause great losses in revenue due to decrease livestock production and animal death. The use of anthelmintic to control gastrointestinal nematode put a selection pressure on nematode populations which led to emergence of anthelmintic resistance. Because of that, this study was carried out to investigate the efficacy of aqueous and methanol extract of Cassia siamea against the motility of C. elegans Bristol N2 and C. elegans DA1316. Caenorhabditis elegans Bristol N2 is a susceptible strain and C. elegans DA1316 is an ivermectin resistant strain. In vitro bioassay of various concentrations of (0.2, 0.6, 0.8, 1.0 and 2.0 mg mL-1) aqueous and methanol extracts of C. siamea was conducted against the motility of L4 larvae of C. elegans Bristol N2 and C. elegans DA1316. The L4 larvae were treated with 0.02 μg mL-1 of ivermectin served as positive control while those in M9 solution served as negative control. The activity of the extracts was observed after 24 h and 48 h. A significant difference was recorded in the extract performance compared to control at (P < 0.001) after 48 h against the motility of the larvae of both strains. The methanol extracts inhibited the motility of C. elegans Bristol N2 by 86.7% as well as DA1316 up to 84.9% at 2.0 mg mL-1 after 48 h. The methanol extract was more efficient than aqueous extract (P < 0.05) against the motility of both strains of C. elegans. Cassia siamea may be used as a natural source of lead compounds for the development of alternative anthelmintic against parasitic nematodes as well ivermectin resistant strains of nematodes.
Citrus medica is a horticultural crop grown in different parts of the world. The plant leaves have medicinal importance in traditional medicine for the treatment of various diseases. The leaves are an underutilised part of the plant, despite having various bioactive compounds with health benefits, with phytochemical analysis having revealed the presence of flavonoids, fatty acids, alkaloids, terpenoids, glycosides, carbohydrates and phytosterols. The biochemical constituents were identified using Fourier-transform infrared spectroscopy (FTIR) and gas chromatography-mass spectrometry (GC-MS), which confirmed the presence of terpenoids, alcohols, alkanes, phytosterols and fatty acids. Among these, methyl 8, 11, 14-heptadecatrienoate is a linolenic acid, and α-linolenic acid, trimethylsilyl ester and levulinic acid are the predominant compounds belonging to the omega-3 fatty acid group, which has known health benefits. Further, the antimicrobial activity of C. medica plant leaves were tested against certain food-borne pathogens and showed significant results. The minimum inhibitory concentrations ranged from 6.09 mg/mL to 390 mg/mL for bacterial organisms and 48.75 mg/mL to 390 mg/mL for fungal organisms. The antioxidant activity values were 300 μg/mL and 450 μg/mL by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assay, respectively. The methanolic extract from the C. medica leaves also showed anticancer activity against MCF7 breast cancer cell lines, with an IC50 value of material for developing a healthy processed food such as nutraceuticals and functional foods.
Malaria still remains a life-threatening parasitic disease with universal targets set for control and elimination. This study aimed to evaluate the in vitro antimalarial susceptibility of Plasmodium falciparum isolates and Plasmodium berghei to selected antimalarial agents and column chromatographic subfractions of Glyphaea brevis leaves extract and FTIR and GCMS of SF8. Trager and Jensen as well as World Health Organisation (WHO) standardised in vitro micro-test system methods were used to determine susceptibility on the patients' blood samples; Column chromatographic procedure was carried out to obtain 11 pooled fractions; FTIR and GCMS were used to determine functional groups and phytochemicals respectively. In vitro anti-plasmodial activity against P. falciparum clinical isolates had IC50 range of 1.03 μg/mL-7.63 μg/mL while their IC50 against P. berghei ranges from 4.32 μg/mL-7.89 μg/mL. Subfraction 8 (SF8) had the least IC50 of 4.32 μg/mL. The FTIR spectrum showed the presence of isoprenoid, alcohol, phenol, alkane, alkenes, ester, carboxylic acids, aromatics and nitro compounds while GCMS identified dodecanoic acid, methyl ester; carotol; hexadecanoic acid, methyl ester; 9-octadecenoic acid (Z)-, methyl ester (oleic acid); methyl stearate; heptadecanoic acid, 16-methyl-, methyl ester; all with their antimalarial reported activities. In conclusion, G. brevis has a great potential for drug development against malaria parasite since it inhibited schizont growth and possesses phytocompounds with antimalarial report.
Malaria continues to be a major public health issue in a number of countries, particularly in tropical regions-the emergence of drug-resistant Plasmodium falciparum encourages new drug discovery research. The key to Plasmodium falciparum survival is energy production up to 100 times greater than other parasites, primarily via the PfLDH. This study targets PfLDH with natural bioactive compounds from the Zingiberaceae family through molecular docking and molecular dynamic studies. Sulcanal, quercetin, shogosulfonic acid C, galanal A and naringenin are the Top 5 compounds with a lower binding energy value than chloroquine, which was used as a control in this study. By binding to NADH and substrate binding site residues, the majority of them are expected to inhibit pyruvate conversion to lactate and NAD+ regeneration. When compared to sulcanal and control drugs, the molecular dynamics (MD) simulation study indicated that quercetin may be the most stable molecule when interacting with PfLDH.
Probiotics are a non-digestible food ingredient that promotes the growth of beneficial microorganisms in the intestines. One of the functional food ingredients, Momala corn flour, is a source of prebiotics with a resistant starch content of 4.42%. Thi s study aimed to improve the prebiotic properties and resistant starch content of modified corn flour (MCF) Momala Gorontalo by using physical, chemical, and enzymatic modification processes. The research methods include physical modification (heat moisture treatment, annealing, autoclaving-cooling cycling, microwave), chemical modification (acid hydrolysis), and enzymatic modification (debranching pullulanase). The results showed that the modified by heat moisture treatment (HMT) increased RS levels 1-fold, annealing modification (ANN) 8.9-fold, autoclaving-cooling one cycle modification (AC-1C) 2.9-fold, autoclaving-cooling two cycles modification (AC-2C) 2.0-fold, microwave modification (MW) 1.3-fold, acid hydrolysis (HA) modification 5.0-fold, and debranching pullulanase (DP) modification 3.8-fold compared with corn flour control without modification. The value of the prebiotic activity of MCF hydrolysed acid (HA) is 0.03, and debranching pullulanase (DP) is 0.02 against Enteropathogenic Escherichia coli (EPEC). The prebiotic effect value of MCF HA and DP were 0.76 and 0.60, respectively. The prebiotic index value of MCF HA and DP were 0.60 and 0.48, respectively. This study confirms that MCF HA and DP are good prebiotic candidates because they have resistant starch content, low starch digestibility, and resistance to simulated gastric fluid hydrolysis than unmodified corn flour.
Macrophages provide the first line of defense against Shigella flexneri infection in the gastrointestinal tract by inducing a variety of inflammatory and antimicrobial responses. Secondary metabolites of plants are used as drugs against infections that are resistant to common antibiotics. In this study, the innate effects of asiaticoside on the proinflammatory activity of mouse macrophages infected with S. flexneri were investigated. The viability of the infected mouse macrophages were examined using viability assay, while the pro-inflammatory cytokines productions were determined using the enzyme-linked immunosorbent assay (ELISA) for determination of IL-1β, IL-12 p40 and TNF-α levels. The production of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS) protein were determined using the Griess assay and western blot, respectively. Statistical analyses were performed using the Statistical Package of Social Sciences (SPSS) software, version 20. The data obtained from independent experiments (n = 3) were presented as the mean ± standard error of mean (SEM). The results showed that, asiaticoside stimulated the infected macrophages by stimulating increased production of TNF-α, IL-12 p40 and NO as well as increased expression of iNOS in a dose-dependent manner. In contrast the viability of the cells and the production of IL-1β and were reduced also in a dose-dependent manner when compared to untreated cells. These results indicate that asiaticoside has immunomodulatory effects on the innate immune function of infected macrophages, showing the potential use of this compound to reduce the clinical symptoms of the infections.
Marine sponges are acknowledged as bacterial hotspots in the oceanic biome. Aquatic bacteria are being investigated comprehensively for bioactive complexes and secondary metabolites. Cultivable bacteria associated with different species of sea sponges in South China Sea waters adjacent to Bidong Island, Terengganu were identified. Molecular identification was accomplished using 16S rRNA gene cloning and sequencing. Fourteen bacterial species were identified and their phylogenetic relationships were analysed by constructing a neighbour-joining tree with Molecular Evolutionary Genetics Analysis 6. The identified species encompassed four bacterial classes that were Firmicutes, Actinobacteria, Alphaproteobacteria and Gammaproteobacteria known to have been associated with sponges. The potential biotechnological applications of the identified bacteria were compared and reviewed based on relevant past studies. The biotechnological functions of the 14 cultivable isolates have been previously reported, hence reinforcing that bacteria associated with sponges are an abundant resource of scientifically essential compounds. Resilience of psychrotolerant bacteria, Psychrobacter celer, in warm tropical waters holds notable prospects for future research.
Indole-3-acetic acid (IAA) is one of the most physiologically active auxins produced by rhizobacteria and is potentially applied for agriculture. Two endophytic bacteria, VR2 and MG9, isolated from the root of Chrysopogon zizanioides (L.) collected at Cha-Am, and the leaf of Bruguiera cylindrica (L.) Blume collected from a mangrove forest at Ban Laem, Phetchaburi Province, Thailand, were taxonomic characterised based on their phenotypic characteristics and 16S rRNA gene analysis. Strain VR2 was closely related to Enterobacter hormaechei CIP 103441T (99.6% similarity), while strain MG9 was closely related to Bacillus aryabhattai B8W22T (99.9% similarity). Consequently, they were identified as Enterobacter hormaechei and Bacillus aryabhattai, respectively. The IAA production of VR2 and MG9 strains are determined and applied to rice seeds for their root and shoot germination. Strains VR2 and MG9 greatly produced a yield of IAA, 246.00 and 195.55 μg/mL in 1,000 μg/mL of L-tryptophan at pH 6 for 48 h. They showed no significant differences in IAA to root and shoot development. However, the bacterial IAA exhibited potential nearby synthetic IAA, which had a significant effect compared to the control. IAA produced from these two strains might preferably trim down the use of synthetic IAA and could contribute to sustainable agriculture.
The coconut is an important economic crop in the Philippines which currently ranks as the world's second largest producer. This study characterised and identified endophytes from coconut tissue culture in order to gain an initial understanding of their potential uses as sources of bioproducts. The isolates were evaluated using morphological, biochemical and molecular methods. Gram staining results revealed that four out of five bacteria isolated were Gram positive. Isolate CEB 1 fermented all three sugars in the Triple Sugar Iron Test while the other four did not. 16S rDNA gene fragments were amplified from genomic DNA using the universal primers 16F27 and 16R1542. The 16S rDNA sequence were found to be homologous to Bacillus subtilis and Pantoea dispersa. Phylogenetic analyses showed significant clustering of bacterial isolates together with archived DNA of B. subtilis and P. dispersa. All isolated bacteria matched the characteristics of their molecular homologies. Isolate CEB 5, identified as B. subtilis, produced red pigments which are possibly pulcherrimin. Literature reports that pulcherrimin possesses antimicrobial activity against yeast species, microscopic fungi, and postharvest pathogens. P. dispera, on the other hand, has been reported to convert insoluble phosphorus into soluble form to enable plants to take up more phosphorus. Determination of the bioactivities of endophytes reported in this study may enable the discovery of novel bioproducts.
Keradangan hati berkait rapat dengan perubahan dalam ekspresi lipoprotein dan apolipoprotein. Interferon-γ (IFN-γ), wakil tunggal jenis kedua IFN, memainkan peranan yang penting dalam memodulasi dan mempergiatkan tindak balas keradangan. Justeru itu, kajian ini direka untuk mengenal pasti kesan IFN-γ terhadap apolipoprotein A-I (APOA-I) dan penglibatan nuclear factor–kappa B (NF-κB) dalam laluan isyarat tersebut. Tindak balas rantai polymerase transkripsi berbalik kuantitatif (qRT-PCR) dan analisis blot western telah dilaksanakan untuk menguantifikasi ekspresi APOA-I dalam sel-sel HepG2 selepas dirawat dengan IFN-γ. Kajian ini menunjukkan bahawa 50 ng/mL IFN-γ merangsangkan ekspresi mRNA dan protein APOA-I. Walau bagaimanapun, pra-rawatan sel dengan inhibitor laluan isyarat NF-κB mengurangkan tahap ekspresi APOA-I. Kajian ini juga mendemonstrasikan penglibatan langsung isyarat NF-κB dalam ekspresi APOA-I akibat rangsangan IFN-γ, di mana IFN-γ meningkatkan tahap fosforilasi NF-κB p65 Ser468 dan Ser536 kepada 2.59-ganda and 1.63-ganda. Namun demikian, pra-rawatan sel dengan perencat laluan isyarat NF-κB melumpuhkan peningkatkan tersebut dan kemudian mengurangkan ekspresi APOA-I dalam sel HepG2. Sebagai rumusan, kajian ini berjaya mengenalpasti peranan isyarat NF-κB dan pengaktifan p65 Ser468 dan Ser536 sebagai pengantara IFN-γ ke atas induksi APOA-I di dalam sel-sel HepG2.
Burkholderia pseudomallei is a soil-dwelling bacterium that causes a globally emerging disease called melioidosis. Approximately one third of the in silico annotated genes in its genome are classified as hypothetical genes. This group of genes is difficult to be functionally characterised partly due to the absence of noticeable phenotypes under conventional laboratory settings. A bioinformatic survey of hypothetical genes revealed a gene designated as BPSL3393 that putatively encodes a small protein of 11 kDA with a CoA binding domain. BPSL3393 is conserved in all the B. pseudomallei genomes as well as various in other species within the genus Burkholderia. Taking into consideration that CoA plays a ubiquitous metabolic role in all life forms, characterisation of BPSL3393 may uncover a previously over-looked metabolic feature of B. pseudomallei. The gene was deleted from the genome using a double homologous recombination approach yielding a null mutant. The BPSL3393 mutant showed no difference in growth rate with the wild type under rich and minimal growth conditions. An extensive metabolic phenotyping test was performed involving 95 metabolic substrates. The deletion mutant of BPSL3393 was severely impaired in its ethanolamine metabolism. The growth rate of the mutant was attenuated when ethanolamine was used as the sole carbon source. A transcriptional analysis of the ethanolamine metabolism genes showed that they were down-regulated in the BPSL3393 mutant. This seemed to suggest that BPSL3393 functions as a positive regulator for ethanolamine metabolism.
This study evaluated the histopathological changes in the gill, liver and kidney of African catfish (Clarias gariepinus) intoxicated with a sub-lethal dose of Melaleuca cajuputi leaves extract (MCLE) for 96 h. The acute toxicity test has been determined previously with a value of 96-h LC50 = 127 mg/L, hence the selection of sub-lethal ranges from 60 mg/L to 160 mg/L of MCLE. Degenerative alterations were prominent in all tested organs, particularly after exposure to a high concentration of MCLE. Gill exhibited haemorrhage, epithelial lifting, lamellar disorganisation, and necrosis after exposure to a high MCLE concentration. Alterations in the liver include congestion, hydropic degeneration, and vacuolation, whereas lesions in the kidney were pyknosis, vacuolation, hydropic degeneration, and tubular necrosis. The obtained data showed that the organs experienced severe changes proportional to the increase in MCLE concentration. In addition, fish exposed to higher concentrations than the LC50 value experienced irreversible lesions. The present study suggests that the use of MCLE below the LC50 is recommended to avoid severe alterations to organs, particularly in African catfish. This study demonstrated that the use of MCLE above the LC50 promotes severe damage to the gills, liver and kidney of African catfish. However, further investigations are needed to define the causing-mechanisms underlying these effects.
A survey was carried out at Gunung Panti Forest Reserve, Johor from 3-7 August 2006, 2-5 June 2008, and 28-31 July 2008 to inventory the herpetofauna therein. An updated checklist for the area which incorporates findings from previous studies is provided. In total, 37 species of frogs, 1 turtle, 27 lizards, and 11 snakes have been recorded from Gunung Panti Forest Reserve, Johor.
Hematological evaluation of fish is essential to the assessment of their physiological status. This study describes the morphometric analysis and comparison of blood cell characteristics in Zanarchopterus sp., Gerres filamentosus Cuvier, 1829 and Leiognathus decorus (De Vis, 1884). The species were collected at two locations off the coast of Trang Province, Thailand. A comparative hematological evaluation was made to assess the effects of environmental conditions on the blood of the fish. Ten individuals of each species were collected from a seagrass bed at Libong Island, where human activities are increasing, and from a secluded sandy beach. Their blood samples were analysed using the blood smear technique. Erythrocytes of all the studied fishes were either elliptical or oval. The morphometric data from both locations showed that erythrocytes were of similar size, except for those of Zanarchopterus sp. Fish from both stations showed several types of leukocytes, including neutrophils and lymphocytes.The highest proportion of leukocytes was made up of lymphocytes, followed by neutrophils. However, monocytes were only observed in fish from Libong Island and the erythrocytic nuclei of fish collected from Libong Island were both reniform and lobate. Our results show the potential of hematological evaluation as an early warning signal of environmental impacts on aquatic animals. The determination of baseline parameters could provide a tool for the monitoring of environmental quality.