<Emphasis Type="Italic">Streptomyces sanglieri</Emphasis> which colonised and enhanced the growth of <Emphasis Type="Italic">Elaeis guineensis</Emphasis> Jacq. seedlings was antagonistic to <Emphasis Type="Italic">Ganoderma boninense</Emphasis> in in vitro studies

Actinomycete strain AUM 00500 was 99.5 % similar to Streptomyces sanglieri NBRC 100784^T and was evaluated for antagonistic activity towards Ganoderma boninense, the causative fungus of basal stem rot of oil palm. The strain showed strong antifungal activity towards G. boninense in in vitro and SEM analysis showed various modes of inhibition of the fungus. Ethyl acetate extracts of single culture and inhibition zone of cross-plug culture by HPLC indicated that strain AUM 00500 produced two different antibiotics of the glutarimide group namely cycloheximide and actiphenol. In greenhouse trials, oil palm seed treated with spores of S. sanglieri strain AUM 00500 at 10⁹ cfu/ml showed significant (P < 0.05) increase in oil palm seedlings growth when compared to the control. Streptomyces sanglieri strain AUM 00500 successfully colonised the epidermal surface of the roots of treated oil palm seedlings and it was recovered from root fragments plated on starch casein agar.     

An <Emphasis Type="Italic">in vitro</Emphasis> study of the antifungal activity of <Emphasis Type="Italic">Trichoderma virens</Emphasis> 7b and a profile of its non-polar antifungal components released against <Emphasis Type="Italic">Ganoderma boninense</Emphasis>

Ganoderma boninense is the causal agent of a devastating disease affecting oil palm in Southeast Asian countries. Basal stem rot (BSR) disease slowly rots the base of palms, which radically reduces productive lifespan of this lucrative crop. Previous reports have indicated the successful use of Trichoderma as biological control agent (BCA) against G. boninense and isolate T. virens 7b was selected based on its initial screening. This study attempts to decipher the mechanisms responsible for the inhibition of G. boninense by identifying and characterizing the chemical compounds as well as the physical mechanisms by T. virens 7b. Hexane extract of the isolate gave 62.60% ± 6.41 inhibition against G. boninense and observation under scanning electron microscope (SEM) detected severe mycelial deformation of the pathogen at the region of inhibition. Similar mycelia deformation of G. boninense was observed with a fungicide treatment, Benlate^® indicating comparable fungicidal effect by T. virens 7b. Fraction 4 and 5 of hexane active fractions through preparative thin layer chromatography (P-TLC) was identified giving the best inhibition of the pathogen. These fractions comprised of ketones, alcohols, aldehydes, lactones, sesquiterpenes, monoterpenes, sulphides, and free fatty acids profiled through gas chromatography mass spectrometry detector (GC/MSD). A novel antifungal compound discovery of phenylethyl alcohol (PEA) by T. virens 7b is reported through this study. T. virens 7b also proved to be an active siderophore producer through chrome azurol S (CAS) agar assay. The study demonstrated the possible mechanisms involved and responsible in the successful inhibition of G. boninense.     

Genomic structure,QTL mapping,and molecular markers of lipase genes responsible for palm oil acidity in the oil palm (<Emphasis Type="Italic">Elaeis guineensis</Emphasis> Jacq.)

The degradation of triglycerides in oil palm fruit due to an endogenous lipase in the pulp is the main reason for acidification of palm oil, which causes major economic losses and is currently mainly associated with the FLL1 gene. We designed this study to identify all the major genes controlling differences in acidity and lipase activity in the oil palm fruit mesocarp and determine a molecular markers kit to allow marker-assisted selection of commercial varieties with low acidity. Not only one gene (FLL1) but three closely linked genes including FLL1 were found and characterized in LM2T_EgCIR184O12c, a bacterial artificial chromosome sequence of 231 kb. Intra-gene PCR-based markers were designed for these genes. A QTL gene co-localization analysis for oil acidity (percentage of fatty acids released) was performed on two mapping populations. It evidenced a single major QTL at our lipase gene loci, explaining 84 to 92% of phenotypic variation, and validating the main genetic control of palm oil acidification by FLL1 and/or by the two new lipase genes. The three lipase genes had high homology to demonstrated triacylglycerol lipases. While FLL1 shows the highest expression levels, the two other genes may also contribute to oil acidity. Our molecular markers of lipase genes and the associated major QTL is an important step towards marker-assisted selection of commercial varieties with low acidity.     

Novel aromatic polyketides from soil <Emphasis Type="Italic">Streptomyces</Emphasis> spp.: purification,characterization and bioactivity studies

Aromatic polyketides are important therapeutic compounds which include front line antibiotics and anticancer drugs. Since most of the aromatic polyketides are known to be produced by soil dwelling Streptomyces, 54 Streptomyces strains were isolated from the soil samples. Five isolates, R1, B1, R3, R5 and Y8 were found to be potent aromatic polyketide producers and were identified by 16S rRNA gene sequencing as Streptomyces spectabilis, Streptomyces olivaceus, Streptomyces purpurascens, Streptomyces coeruleorubidus and Streptomyces lavendofoliae respectively. Their sequences have been deposited in the GenBank under the accession numbers KF468818, KF681280, KF395224, KF527511 and KF681281 respectively. The Streptomyces strains were cultivated in the media following critically optimised culture conditions. The resulting broth extracts were fractionated on a silica gel column and preparative TLC to obtain pure compounds. The pure compounds were tested for bioactivity and the most potent compound from each isolate was identified by UV–Vis, IR and NMR spectroscopic methods. Isolated S. spectabilis (R1), yielded one potent compound identified as dihydrodaunomycin with an MIC of 4 µg/ml against Bacillus cereus and an IC₅₀ value of 24 µM against HeLa. S. olivaceus (B1), yielded a comparatively less potent compound, elloramycin. S. purpurascens (R3) yielded three compounds, rhodomycin, epelmycin and obelmycin. The most potent compound was rhodomycin with an MIC of 2 µg/ml against B. cereus and IC₅₀ of 15 µM against HeLa. S. coeruleorubidus (R5), yielded daunomycin showing an IC₅₀ of 10 µM and also exhibiting antimetastatic properties against HeLa. S. lavendofoliae (Y8), yielded a novel aclacinomycin analogue with IC₅₀ value of 2.9 µM and potent antimetastatic properties at 1 µM concentration against HeLa. The study focuses on the characterization of aromatic polyketides from soil Streptomyces spp., which can serve as potential candidates for development of chemotherapeutic drugs in future.     

Characterization and antimicrobial activity of the bioactive metabolites in streptomycete isolates

Twenty different streptomycete isolates were obtained from soils of southeast Serbia. Five isolates identified as Streptomyces hygroscopicus (SH100, SH101, SH102, SH103, and SH104) showed strong activity against Botrytis cinerea, a parasite found in domestic vines. These isolates were extensively studied for their in vitro antimicrobial activity against Gram-positive bacteria, Gram-negative bacteria, yeasts and fungi, and also antiviral activity against Herpes simplex. The results indicated that the obtained isolates were highly active against Botrytis cinerea, Candida albicans, and Herpes simplex, with an inhibition zone at ≥31 mm. The structure of the bioactive components was determined using elemental analysis, as well as UV/VIS, FTIR, and TLC.     

<Emphasis Type="Italic">Streptomyces xiangluensis</Emphasis> sp. nov., a novel actinomycete isolated from soil

A novel actinomycete, designated strain NEAU-LA29^T, was isolated from soil collected from Xianglu Mountain and subjected to a polyphasic taxonomic study. Based on a polyphasic taxonomic approach comprising chemotaxonomic, phylogenetic, morphological and physiological characterisation, the isolate has been affiliated to the genus Streptomyces. 16S rRNA gene sequence analysis showed that the isolate is closely related to Streptomyces vastus JCM4524^T (98.8% identity) and Streptomyces cinereus DSM43033^T (97.9%). However, multilocus sequence analysis based on five other house-keeping genes (atpD, gyrB, rpoB, recA and trpB) and low DNA–DNA relatedness values enabled the strain to be differentiated from these closely related species of the genus Streptomyces. Thus, strain NEAU-LA29^T is concluded to represent a novel species of the genus Streptomyces, for which the name Streptomyces xiangluensis sp. nov. is proposed. The type strain is NEAU-LA29^T (=?CGMCC 4.7466^T?=?DSM 105786^T).     

<Emphasis Type="Italic">Streptomyces gamaensis</Emphasis> sp. nov., a novel actinomycete with antifungal activity isolated from soil in Gama,Chad

During an investigation exploring potential sources of novel species and natural products, a novel actinomycete with antifungal activity, designated strain NEAU-Gz11^T, was isolated from a soil sample, which was collected from Gama, Chad. The isolate was found to have morphological and chemotaxonomic characteristics typical of members of the genus Streptomyces. 16S rRNA gene sequence similarity studies showed that strain NEAU-Gz11^T belongs to the genus Streptomyces with high sequence similarity to Streptomyces hiroshimensis JCM 4098^T (98.0 %). Similarities to other type strains of the genus Streptomyces were lower than 98.0 %. However, the physiological and biochemical characteristics and low levels of DNA–DNA relatedness could differentiate the isolate genotypically and phenotypically from S. hiroshimensis JCM 4098^T. Therefore, the strain is concluded to represent a novel species of the genus Streptomyces, for which the name Streptomyces gamaensis sp. nov. is proposed. The type strain is NEAU-Gz11^T (=CGMCC 4.7304^T=DSM 101531^T).     

A review of the palm genus <Emphasis Type="Italic">Acrocomia</Emphasis>: Neotropical green gold

The genus Acrocomia, popularly known as macaw palm or macaúba, occurs in savanna areas and open forests of tropical America, with distribution from Central to southern South America. They are important oleaginous palm trees, due to their role in ecosystems and local economies and their potential for biofuel production and vegetable oil. Although the taxonomy of the genus is not resolved because of observed phenotypic diversity in A. aculeata (Jacq.) Lodd. ex Mart., there are several conflicting treatments. Some authors recognize three caulescent spp. occurring in South America: A. aculeata, A. intumescens Drude, and A. totai Mart, although a new one was described recently—Acrocomia corumbaensis. Because some Latin American governments want to expand production of macaw palm in their territory as raw material for agro-energy, several groups have been encouraged to study this genus, focusing on the production of biodiesel, seed germination, phenotypic aspects, and genetic diversity. The goal of this review is to compile key information available in the literature and herbarium data, focusing on South American populations of the genus.     

Plant growth promotion and suppression of <Emphasis Type="Italic">Phytophthora drechsleri</Emphasis> damping-off in cucumber by cellulase-producing <Emphasis Type="Italic">Streptomyces</Emphasis>

Phytophthora drechsleri damping-off is one of the most important diseases of cucumber (Cucumis sativus). Salinity is a serious problem for crop production and affects diversity and activity of soil microorganisms. Application of salt-tolerant biocontrol agents may be beneficial in order to protect plants against pathogenic fungi in saline soils. In this study, a total of 717 Streptomyces isolates were isolated from the rhizosphere of cucumber, out of which two isolates showed more than 70% inhibitory effect against P. drechsleri and had cellulase activity in the presence and absence of NaCl. In a greenhouse experiment, two Streptomyces isolates with the highest antagonistic activity, strains C 201 and C 801, reduced seedling damping-off of cucumber caused by P. drechsleri by 77 and 80%, respectively, in artificially infested soils. Strain C 201 increased dry weight of seedlings up to 21% in greenhouse experiments. Phylogenetic analyses of 16S rRNA gene sequence reveals that strains C 201 and C 801 are closely related to S. rimosus and S. monomycini respectively. Increased activity of polyphenol oxidase (PPO) and peroxidase (POX) enzymes in Streptomyces-treated plants proved the biocontrol-induced systemic resistance (ISR) in cucumber plants against P. drechsleri.     

Enhancement of rapamycin production by metabolic engineering in <Emphasis Type="Italic">Streptomyces hygroscopicus</Emphasis> based on genome-scale metabolic model

Rapamycin, as a macrocyclic polyketide with immunosuppressive, antifungal, and anti-tumor activity produced by Streptomyces hygroscopicus, is receiving considerable attention for its significant contribution in medical field. However, the production capacity of the wild strain is very low. Hereby, a computational guided engineering approach was proposed to improve the capability of rapamycin production. First, a genome-scale metabolic model of Streptomyces hygroscopicus ATCC 29253 was constructed based on its annotated genome and biochemical information. The model consists of 1003 reactions, 711 metabolites after manual refinement. Subsequently, several potential genetic targets that likely guaranteed an improved yield of rapamycin were identified by flux balance analysis and minimization of metabolic adjustment algorithm. Furthermore, according to the results of model prediction, target gene pfk (encoding 6-phosphofructokinase) was knocked out, and target genes dahP (encoding 3-deoxy-d-arabino-heptulosonate-7-phosphate synthase) and rapK (encoding chorismatase) were overexpressed in the parent strain ATCC 29253. The yield of rapamycin increased by 30.8% by knocking out gene pfk and increased by 36.2 and 44.8% by overexpression of rapK and dahP, respectively, compared with parent strain. Finally, the combined effect of the genetic modifications was evaluated. The titer of rapamycin reached 250.8 mg/l by knockout of pfk and co-expression of genes dahP and rapK, corresponding to a 142.3% increase relative to that of the parent strain. The relationship between model prediction and experimental results demonstrates the validity and rationality of this approach for target identification and rapamycin production improvement.     

Mapping QTL for leaf area in oil palm using genotyping by sequencing

Leaf area is an important parameter in oil palm breeding as it is positively correlated with oil yield. However, measurement of leaf area is tedious and also destructive. In the present study, a breeding population with 145 palms derived from a cross between Deli Dura and Avros Pisifera was used to construct a high-density linkage map and identify quantitative trait loci (QTL) for leaf area in oil palm. Using genotyping by sequencing, a linkage map containing 2413 SNPs was constructed. The total length of the linkage map was 1161.89 cM, with an average marker spacing of 0.48 cM. Based on the continuous phenotyping of leaf area from 2010 to 2015, two suggestive QTL for leaf area were mapped on chromosomes (Chr) 3 and 9. The allelic effects of the QTL associated with leaf area in the mapping population were estimated by linear regression using ordinary least squares method. The QTL on Chr 9 explained 13.3% of phenotypic variation for leaf area. A candidate gene, ARC5, within the narrow interval of QTL on Chr 9 was identified. The gene was significantly higher expressed in leaf than root and fruit of oil palm. This high-quality and SNP-based map supplies a base to fine map QTL for agronomic traits in oil palm, and the markers closely linked to the stable QTL may be used in marker-assisted selection in oil palm breeding.     

Anionic carbohydrate-containing cell wall polymers of <Emphasis Type="Italic">Streptomyces melanosporofaciens</Emphasis> and related species

The structures of cell wall anionic carbohydrate-containing polymers in Streptomyces melanosporofaciens VKM Ac-1864^T and phylogenetically close organisms—S. hygroscopicus subsp. hygroscopicus VKM Ac-831^T, S. violaceusniger VKM Ac-583^T, S. endus VKM Ac-1331^T, S. endus VKM Ac-129, and S. rutgersensis subsp. castelarensis VKM Ac-832^T—have been comparatively studied by chemical and NMR spectroscopic methods. The natural polymer of a new, previously unknown structure, Kdn (3-deoxy-D-glycero-D-galacto-non-2-ulopyranosonic acid) with β-galactose residues at C-9, has been found in the cell walls of all the strains under study. The cell walls of all the studied organisms contain three teichoic acids (TA): a predominant TA (1,3-poly(glycerol phosphate) with N-acetylated α-glucosaminyl substitutes by C-2 of glycerol, and minor TAs, 1,3-and 2,3-poly(glycerol phosphate) polymers without substitution. Their chains have O-acetyl and O-lysyl groups. Microorganisms of the above-mentioned species differ in the number of α-glucosaminyl substitutes and in the degree of their acetylation in the predominant teichoic acid.     

Two <Emphasis Type="Italic">Streptomyces</Emphasis> Species Producing Antibiotic,Antitumor, and Anti-Inflammatory Compounds Are Widespread Among Intertidal Macroalgae and Deep-Sea Coral Reef Invertebrates from the Central Cantabrian Sea

Streptomycetes are widely distributed in the marine environment, although only a few studies on their associations to algae and coral ecosystems have been reported. Using a culture-dependent approach, we have isolated antibiotic-active Streptomyces species associated to diverse intertidal marine macroalgae (Phyllum Heterokontophyta, Rhodophyta, and Chlorophyta), from the central Cantabrian Sea. Two strains, with diverse antibiotic and cytotoxic activities, were found to inhabit these coastal environments, being widespread and persistent over a 3-year observation time frame. Based on 16S rRNA sequence analysis, the strains were identified as Streptomyces cyaneofuscatus M-27 and Streptomyces carnosus M-40. Similar isolates to these two strains were also associated to corals and other invertebrates from deep-sea coral reef ecosystem (Phyllum Cnidaria, Echinodermata, Arthropoda, Sipuncula, and Anelida) living up to 4.700-m depth in the submarine Avilés Canyon, thus revealing their barotolerant feature. These two strains were also found to colonize terrestrial lichens and have been repeatedly isolated from precipitations from tropospheric clouds. Compounds with antibiotic and cytotoxic activities produced by these strains were identified by high-performance liquid chromatography (HPLC) and database comparison. Antitumor compounds with antibacterial activities and members of the anthracycline family (daunomycin, cosmomycin B, galtamycin B), antifungals (maltophilins), anti-inflamatory molecules also with antituberculosis properties (lobophorins) were identified in this work. Many other compounds produced by the studied strains still remain unidentified, suggesting that Streptomyces associated to algae and coral ecosystems might represent an underexplored promising source for pharmaceutical drug discovery.     

Diversity and occupancy of small carnivores within oil palm plantations in central Sumatra,Indonesia

In Southeast Asia, the conversion of native forests to oil palm plantations threatens tropical biodiversity, but very little is known about the impacts of oil palm cultivation on small carnivore species. To determine the diversity and occupancy of small carnivores within oil palm plantations and to investigate possible factors that might affect their presence within oil palm, we used camera-traps within two oil palm plantations in central Sumatra, analysed the data using occupancy modelling and tested whether two covariates (distance to the edge of the oil palm habitat and distance from extensive areas of lowland forest) affected the model parameters for each small carnivore species. From 3164 camera-trap days, we detected only three small carnivores: leopard cat (Prionailurus bengalensis), common palm civet (Paradoxurus hermaphroditus) and Malay civet (Viverra tangalunga), which indicates that there was a low diversity of small carnivores within the oil palm plantations. Both the leopard cat and common palm civet were found deep within the oil palm, whereas the Malay civet was only detected near the edge in one of the plantations. The leopard cat and common palm civet had very high occupancy values, whereas the Malay civet had low values for both occupancy and detection probability. Neither covariate affected occupancy of the leopard cat and common palm civet, but distance from the edge of the oil palm habitat did influence their detection probabilities. Malay civet occupancy decreased with distance from the oil palm edge, and detection probability was affected by distance from extensive areas of lowland forest. Forests and rest/den site availability are suggested to be important features for small carnivores with oil palm-dominated landscapes.     

Development and application of a T7 RNA polymerase-dependent expression system for antibiotic production improvement in <Emphasis Type="Italic">Streptomyces</Emphasis>

Objective

To develop a reliable and easy to use expression system for antibiotic production improvement of Streptomyces.

Results

A two-compound T7 RNA polymerase-dependent gene expression system was developed to fulfill this demand. In this system, the T7 RNA polymerase coding sequence was optimized based on the codon usage of Streptomyces coelicolor. To evaluate the functionality of this system, we constructed an activator gene overexpression strain for enhancement of actinorhodin production. By overexpression of the positive regulator actII-ORF4 with this system, the maximum actinorhodin yield of engineered strain was 15-fold higher and the fermentation time was decreased by 48 h.

Conclusion

The modified two-compound T7 expression system improves both antibiotic production and accelerates the fermentation process in Streptomyces. This provides a general and useful strategy for strain improvement of important antibiotic producing Streptomyces strains.