Factors affecting the production of Trichoderma harzianum secondary metabolites during the interaction with different plant pathogens

Aims: Strains of Trichoderma spp. produce numerous bioactive secondary metabolites. The in vitro production and antibiotic activities of the major compounds synthesized by Trichoderma harzianum strains T22 and T39 against Leptosphaeria maculans, Phytophthora cinnamomi and Botrytis cinerea were evaluated. Moreover, the eliciting effect of viable or nonviable biomasses of Rhizoctonia solani, Pythium ultimum or B. cinerea on the in vitro production of these metabolites was also investigated. Methods and Results: T22azaphilone, 1‐hydroxy‐3‐methyl‐anthraquinone, 1,8‐dihydroxy‐3‐methyl‐anthraquinone, T39butenolide, harzianolide, harzianopyridone were purified, characterized and used as standards. In antifungal assays, T22azaphilone and harzianopyridone inhibited the growth of the pathogens tested even at low doses (1–10 μg per plug), while high concentrations of T39butenolide and harzianolide were needed (>100 μg per plug) for inhibition. The in vitro accumulation of these metabolites was quantified by LC/MS. T22azaphilone production was not enhanced by the presence of the tested pathogens, despite its antibiotic activity. On the other hand, the anthraquinones, which showed no pathogen inhibition, were stimulated by the presence of P. ultimum. The production of T39butenolide was significantly enhanced by co‐cultivation with R. solani or B. cinerea. Similarly, viable and nonviable biomasses of R. solani or B. cinerea increased the accumulation of harzianopyridone. Finally, harzianolide was not detected in any of the interactions examined. Conclusions: The secondary metabolites analysed in this study showed different levels of antibiotic activity. Their production in vitro varied in relation to: (i) the specific compound; (ii) the phytopathogen used for the elicitation; (iii) the viability of the elicitor; and (iv) the balance between elicited biosynthesis and biotransformation rates. Significance and Impact of the Study: The use of cultures of phytopathogens to enhance yields of Trichoderma metabolites could improve the production and application of novel biopesticides and biofertilizers based on the active compounds instead of the living microbe. This could have a significant beneficial impact on the management of diseases in crop plants.     

Flooding tolerance of Tabebuia cassinoides: Metabolic, morphological and growth responses

Tabebuia cassinoides (Lam.) DC (Bignoniaceae) is a tree species that occurs in swampy areas of the coastal “restinga” in SE Brazil (a coastal sandy plains scrub and forest formation). To elucidate possible adaptive strategies that enable this species to occupy areas subjected to seasonal or perennial waterlogging, metabolic, morphological and growth responses of plants under flooding conditions were studied. The root system of T. cassinoides plants presented elevated amounts of ethanol (10.6 μmol g⁻¹ fresh wt) only in the first 5 d of soil water saturation. The two-fold increase in ethanol production under flooding was corroborated by an increase in ADH activity in the same period. Lactic acid concentrations did not change significantly during four months of flooding treatment. The decrease of alcoholic fermentation under hypoxia was associated with the appearing of new roots. The induction of aerenchyma formation in roots developed under flooding conditions, allowed oxygen transport from the shoot to these organs, thus maintaining an aerobic respiration. We conclude that this characteristic and the capacity to oxidize the rhizosphere are probably responsible for the survival and growth of plants while flooded and for their success in an environment, which restricts the presence of the majority of competing tree species.     

Selenium supplementation influences growth performance, antioxidant status and immune response in lambs

To investigate and compare the effect of inorganic and organic Se supplementation, 18 male lambs (24.68 ± 2.89 kg mean body weight, about 8–9 months of age) were divided into three groups of six animals in each, following randomized block design. While animals in the control group (Gr I) were fed a standard TMR containing 195 g/kg crushed maize grain, 175.5 g/kg soybean meal, 260 g/kg wheat bran, 13 g/kg mineral mixture (without Se), 6.5 g/kg common salt and 350 g/kg wheat straw, animals in Gr II and Gr III were additionally supplemented with 0.15 mg Se/kg of diet through sodium selenite (inorganic Se) and Jevsel-101 (organic Se), respectively. Experimental feeding was done for a period of 90 days. To assess the humoral immune response, all the lambs were intramuscularly inoculated with a single dose (2 mL) of Haemorrhagic septicaemia oil adjuvant vaccine on day 0; and blood samples were collected on day 0, 30, 60 and 90. Supplementation of Se had no effect on serum total cholesterol, total protein, albumin, globulin, albumin:globulin ratio, T₃, T₄, T₄:T₃ ratio; serum Ca and P levels and SGOT and SGPT activity. However, there was a significant increase in the serum Se level, RBC GSH-Px activity and humoral immune response in both the Se supplemented groups as compared to control group. Average daily gain (g) was highest (110) in Gr III, followed by Gr II (98.2) and lowest in Gr I (89.1). Thus, supplementation of organic as well as inorganic Se was found to improve the growth rate, humoral immune response and antioxidant status of the lambs; and between two sources, organic Se was more effective than inorganic Se.     

Effects of isobutyrate on rumen fermentation, lactation performance and plasma characteristics in dairy cows

The objective of this study was to evaluate effects of isobutyrate supplementation on rumen fermentation, lactation performance and plasma characteristics of dairy cows. Twenty multiparous second filial generation (F2) cows of a cross between Chinese Jinnan Yellow and Holstein cows at 148 ± 4.5 days in milk and 22.3 ± 0.81 kg milk production were used in a replicated 4 × 4 Latin square experiment. The treatments were: control (without isobutyrate), low (LIB), medium (MIB) and high (HIB) isobutyrate supplementation of 20, 40 and 60 g per cow per day, respectively. Experimental periods were 30 days with 15 d of adaptation and 15 d of data collection. Dry matter (DM) intake was not affected by increasing isobutyrate supplementation, but milk yields were highest for the 40 g/d isobutyrate supplementation level, where proportion of milk fat, true protein and lactose were minimized. Ruminal pH (6.38–6.24) and ammonia N (13.8–11.1 mg/100 ml) were linearly (P<0.01) decreased, whereas total VFA concentration (124–131 mM) increased at a decreasing rate with increasing isobutyrate supplementation. The ratio of acetate to propionate increased linearly (P<0.01) from 2.77 to 4.43 as isobutyrate supplementation increased due to the increase in acetate production and decrease in propionate production. Digestibilities of OM in the total tract increased linearly (P<0.01) as isobutyrate supplementation increased, digestibilities of DM and EE were highest for the 40 g/d isobutyrate supplementation level, digestibilities of CP, aNDF and ADF increased at a decreasing rate with increasing isobutyrate supplementation. Plasma concentrations of glucose and growth hormone linearly (P<0.03) increased, whereas concentrations of non-esterified fatty acids linearly (P<0.01) decreased. Results indicate that supplementation of this diet with isobutyrate changed the rumen fermentation pattern towards acetate production, improved digestion and modified plasma concentrations of glucose and growth hormone. This suggests that isobutyrate stimulated digestive microorganisms or enzymes in a dose-dependent manner with the optimum isobutyrate dose at about 40 g per cow per day in terms of improved digestion.     

Alternaria alternata AT Toxin Induces Programmed Cell Death in Tobacco

Detached tobacco leaves were infiltrated with an AT toxin preparation from the foliar pathogen Alternaria alternata tobacco pathotype. The AT toxin preparation caused formation of necrotic lesions within 5 days post-infiltration in a concentration-dependent manner. Cell death was accompanied by increased levels of the stress metabolites hydrogen peroxide, malondialdehyde, free proline and by enhanced total protease activity. Lesion development and the production of stress metabolites were suppressed if the infiltration site was pre-infiltrated with caspase-specific peptide inhibitors (irreversible caspase-1 inhibitor acyl-Tyr-Val-Ala-Asp-chloromethylketone (Ac-YVAD-CMK) and the broad range caspase inhibitor benzyoxycarbonyl-Asp-2,6-dichlorobenzoyloxymethylketone (Z-Asp-CH2-DCB)), the serine protease inhibitor Nα-p-tosyl- l -lysine chloromethylketone and the polyamine spermine. Extensive accumulation of reactive oxygen species (ROS), as determined by staining with 3-3'-diaminobenzidine and 2',7'-dichlorofluorescein diacetate, was found in the AT toxin-challenged lesions. The data show that AT toxin-induced cell death in tobacco is a type of programmed cell death in which caspase-like proteases and ROS signalling play a prominent role.     

Taxol biosynthesis: Identification and characterization of two acetyl CoA:taxoid-O-acetyl transferases that divert pathway flux away from Taxol production

Two cDNAs encoding taxoid-O-acetyl transferases (TAX 9 and TAX 14) were obtained from a previously isolated family of Taxus acyl/aroyl transferase cDNA clones. The recombinant enzymes catalyze the acetylation of taxadien-5α,13α-diacetoxy-9α,10β-diol to generate taxadien-5α,10β,13α-tri-acetoxy-9α-ol and taxadien-5α,9α,13α-triacetoxy-10β-ol, respectively, both of which then serve as substrates for a final acetylation step to yield taxusin, a prominent side-route metabolite of Taxus. Neither enzyme acetylate the 5α- or the 13α-hydroxyls of taxoid polyols, indicating that prior acylations is required for efficient peracetylation to taxusin. Both enzymes were kinetically characterized, and the regioselectivity of acetylation was shown to vary with pH. Sequence comparison with other taxoid acyl transferases confirmed that primary structure of this enzyme type reveals little about function in taxoid metabolism. Unlike previously identified acetyl transferases involved in Taxol production, these two enzymes appear to act exclusively on partially acetylated taxoid polyols to divert the Taxol pathway to side-route metabolites.     

一株放线菌代谢产物除草活性的初步研究

从土壤中分离筛选出1株编号为Z40的链霉菌菌株,其代谢产物具有明显的除草活性。室内生物测定结果表明,Z40菌株发酵液干物质的除草活性主要表现为抑制生长作用。对反枝苋的主根和主茎生长抑制中浓度(EC50)分别为276.76 mg/L和1 609.37 mg/L,对狗尾草的主根和主茎生长抑制中浓度分别为501.81mg/L和629.01 mg/L。浓度为10 000 mg/L对油菜的萌发抑制率和主茎生长抑制率均为100.00%,而对小麦的生长发育没有明显影响。田间试验结果表明,Z40菌株代谢产物对稗、狗尾草、铁苋菜的防治效果均在80%以上,抑制率分别为80.30%,81.59%和88.71%。     

Control of blast and sheath blight diseases of rice using antifungal metabolites produced by Streptomyces sp. PM5

Two antifungal aliphatic compounds, SPM5C-1 and SPM5C-2 with a lactone and ketone carbonyl unit, respectively obtained from Streptomyces sp. PM5 were evaluated under in vitro and in vivo conditions against major rice pathogens, Pyricularia oryzae and Rhizoctonia solani. These compounds were dissolved in distilled water/medium to get the required concentrations. The well diffusion bioassay indicated that the of SPM5C-1 remarkably inhibited the mycelial growth of P. oryzae and R. solani in comparison to SPM5C-2. Though SPM5C-2 showed low antifungal activity against P. oryzae, it was not active against R. solani. Further, SPM5C-1 completely inhibited the growth of P. oryzae and R. solani at concentrations of 25, 50, 75 and 100 μg/ml. Greenhouse experiments revealed that spraying of SPM5C-1 at 500 μg/ml on rice significantly decreased blast and sheath blight development by 76.1% and 82.3%, respectively, as compared to the control with a corresponding increase in rice grain yield.