Degradation of indole by enrichment culture and Pseudomonas aeruginosa Gs isolated from mangrove sediment

Microbial degradation of indole was investigated using enrichment culture and Pseudomonas aeruginosa Gs obtained from mangrove sediment of Hong Kong Nature Reserve as inocula. Degradation of indole using this enrichment culture and Ps. aeruginosa Gs was quantified on reversed-phase high-performance liquid chromatography. Initial concentrations of indole affected degradation and the results conformed to the zero-order kinetic model. The optimum pH and salinity were 7.0 and 5‰, respectively, for indole degradation by Ps. aeruginosa Gs. Two major metabolites of indole degradation were detected. This study suggests that indole can be rapidly degraded by indigenous microorganisms of the mangrove environment.     

Production of indole-3-acetic acid and related indole derivatives from L-tryptophan by <Emphasis Type="Italic">Rubrivivax benzoatilyticus</Emphasis> JA2

Rubrivivax benzoatilyticus JA2 produces indoles with simultaneous utilization of L-tryptophan. Fifteen chromatographically distinct indole derivatives were detected from the L-tryptophan-supplemented cultures of R. benzoatilyticus JA2. Nine of these were identified as, indole 3-acetamide, Methoxyindole-3-aldehyde, indole 3-aldehyde, methoxyindole-3-acetic acid, indole 3-acetic acid, indole-3-carboxylic acid, indole-3-acetonitrile, indole, and trisindoline. Tryptophan stable isotope feeding confirmed the indoles produced are from the supplemented L-tryptophan. Indole 3-acetic acid is one of the major products of L-tryptophan catabolism by R. benzoatilyticus JA2 and its production was influenced by growth conditions. Identification of indole 3-acetamide and tryptophan monooxygenase activity suggests indole 3-acetamide routed IAA biosynthesis in R. benzoatilyticus JA2. The study also indicated the possible multiple pathways of IAA biosynthesis in R. benzoatilyticus JA2.     

Biodegradation of indole at high concentration by persolvent fermentation with Pseudomonas sp. ST-200

Pseudomonas sp. strain ST-200 grew on indole as a sole carbon source. The minimal inhibitory concentration of indole was 0.3 mg/ml for ST-200. However, ST-200 grew in a persolvent fermentation system containing a large amount of indole (a medium containing 20% by vol. diphenylmethane and 4 mg/ml indole), because most of the indole was partitioned in the organic solvent layer. When the organism was grown in the medium containing indole at 1 mg/ml in the presence of diphenylmethane, more than 98% of the indole was consumed after 48 h. Isatic acid (0.4 mg/ml) and isatin (0.03 mg/ml) were produced as the metabolites in the aqueous medium layer. Received: September 12, 1996 / Accepted: January 2, 1997     

A novel regulatory circuit to control indole biosynthesis protects Escherichia coli from nitrosative damages during the anaerobic respiration of nitrate

Indole is a widely distributed microbial secondary metabolite. It mediates a broad range of physiological processes in both its producing and surrounding species. Yet, indole biosynthesis during the anaerobiosis of bacteria remains largely uncharacterized. Here, we find that while indole production is promoted during fermentation and anaerobic respiration of fumarate and trimethylamine N‐oxide in E. coli, its biosynthesis is repressed during anaerobic respiration of nitrate especially during exponential growth. We show that expression of the indole biosynthetic operon tnaCAB is repressed under this condition by the two component systems NarXL and NarPQ in the global regulator FNR dependent manner. During stationary growth phase of nitrate respiration, indole biosynthesis is derepressed. However, cellular indole concentration remains low. We demonstrate that this is due to the rapid conversion of indole into mutagenic indole nitrosative derivatives under this condition. Consistent with this, a supplement of exogenous indole during nitrate respiration causes elevated mutation frequencies in E. coli cells lacking the detoxifying efflux genes mdtEF, and ectopic over‐expression of tnaAB genes decreases the fitness of E. coli to this physiological condition. Together, these results suggest that indole production is tuned to the bioenergetics activities of E. coli to facilitate its adaptation and fitness.     

Production of a novel indole ester from 2-aminobenzoate by Rhodobacter sphaeroides OU5

Culture supernatants of Rhodobacter sphaeroides OU5 grown in the presence of 2-aminobenzoate gave an orange-red color-reaction with Salpers reagent, suggesting the presence of an indole derivative. This production was light-dependent and inducible only with 2-aminobenzoate. Replacement of 2-aminobenzoate with other 2-substituted benzoates did not result in the formation of indole. Fumarate appeared to be the conjugating molecule with 2-aminobenzoate, resulting in the formation of an indole derivative. The purified indole derivative was orange-brown in color, with a yields 0.34 mM from 1 mM 2-aminobenzoate. Infrared analysis suggested an indole ester and ¹H NMR analysis indicated an indole carboxylate, esterified with a terpenoid alcohol. The indole ester has a mass of 441 with the molecular formula C₂₇H₃₉NO₄. The IUPAC name of the compound is (3 E,5 E)-14-hydroxy-3,7,11-trimethyl-3,5-tetradecadienyl 2-(hydroxymethyl)-1 H-indole-3-carboxylate; and the common name given to this compound is sphestrin.     

Indole analogues decreasing the virulence of Vibrio campbellii towards brine shrimp larvae

Indole signalling has been proposed as a potential target for the development of novel virulence inhibitors to control bacterial infections. However, the major structural features of indole analogues that govern antivirulence activity remain unexplored. Therefore, we investigated the impact of 26 indole analogues on indole-regulated virulence phenotypes in Vibrio campbellii and on the virulence of the bacterium in a gnotobiotic brine shrimp model. The results demonstrated that 10 indole analogues significantly increased the fluorescence of indole reporter strain Vibrio cholerae S9149, 21 of them decreased the swimming motility of V. campbellii, and 13 of them significantly decreased the biofilm formation of V. campbellii. Further, we found that 1-methylindole, indene, 2,3-benzofuran, thianaphthene, indole-3-acetonitrile, methyl indole-3-carboxylate, 3-methylindole, and indole-2-carboxaldehyde exhibited a significant protective effect on brine shrimp larvae against V. campbellii infection, resulting in survival rates of challenged brine shrimp above 80%. The highest survival of shrimp larvae (98%) was obtained with indole-3-acetonitrile, even at a relatively low concentration of 20 μM. Importantly, the indole analogues did not affect bacterial growth, both in vitro and in vivo. These results indicate the potential of indole analogues in applications aiming at the protection of shrimp from vibriosis.     

Transformation of indole and quinoline by Desulfobacterium indolicum (DSM 3383)

Degradation of indole and quinoline by Desulfobacterium␣indolicum was studied in batch cultures. The first step in the degradation pathway of indole and quinoline was a hydroxylation at the 2 position to oxindole and 2-hydroxyquinoline respectively. These hydroxylation reactions followed saturation kinetics. The kinetic parameters for indole were an apparent maximum specific transformation rate (V _Amax) of 263 μmol mg total protein⁻¹ day⁻¹ and an apparent half-saturation constant (K _Am) of 139 μM. The V _Amax for quinoline was 170 μmol mg total protein⁻¹ day⁻¹ and K _Am was 92 μM. Oxindole inhibited indole hydroxylation whereas 2-hydroxyquinoline stimulated quinoline hydroxylation. An adaptation period of approximately 20 days was required before transformation of 2-hydroxyquinoline in cultures previously grown on quinoline. Indole and quinoline were hydroxylated with a lag phase shorter than 4 h in a culture adapted to ethanol. Chloramphenicol inhibited the hydroxylation of indole and quinoline in ethanol-adapted cells, indicating an inducible enzyme system. Chloramphenicol had no effect on the hydroxylation of indole in quinoline-adapted cells or on the hydroxylation of quinoline in indole-adapted cells. This indicated that it was the same inducible enzyme system that hydroxylated indole and quinoline. Received: 16 July 1996 / Received revision: 23 September 1996 / Accepted: 29 September 1996     

Transformation of indole by methanogenic and sulfate-reducing microorganisms isolated from digested sludge

In the present study, mineralization of an aromaticN-heterocyclic molecule, indole, by microorganisms present in anaerobically digested sewage sludge was examined. The first step in indole mineralization was the formation of a hydroxylated intermediate, oxindole. The rate of transformation of indole to oxindole and its subsequent disappearance was dependent on the concentration of inoculum and indole and the incubation temperature. Methanogenesis appeared to be the dominant process in the mineralization of indole in 10% digested sludge even in the presence of high concentrations of sulfate. Enrichment of the digested sludge with sulfate as an electron acceptor allowed the isolation of a metabolically stable mixed culture of anaerobic bacteria which transformed indole to oxindole and acetate, and ultimately to methane and carbon dioxide. This mixed culture exhibited a predominance of sulfate-reducers over methanogens with more than 75% of the substrate mineralized to carbon dioxide. The investigation demonstrates that indole can be transformed by both methanogenic and sulfate-reducing microbial populations.     

Chaunopycnis pustulata sp. nov., a new clavicipitalean anamorph producing metabolites that modulate potassium ion channels

Two morphologically similar strains of hyphomycetous fungi, MF5785 and MF5638, produced potent indole diterpene antagonists of the calcium-gated potassium ion channel, Maxi-K, and a diterpene blocker (nalanthalide) of the voltage-gated potassium channel, Kv 1.3, respectively. The two strains were tentatively identified in the literature as Nalanthamala species. Analyses of their secondary metabolite profiles by HPLC and mass spectroscopy demonstrated that both produced a series of indole diterpenes, of which at least one was produced by both strains. Another strain, Chaunopycnis alba (MF6799), that produced the diterpene, nalanthalide, also produced indole diterpene metabolites. Morphological comparisons and phylogenetic studies based on 28S and ITS rDNA sequences indicated that MF5785 and MF5368, and another soil-derived strain GB6597, belonged to a monophyletic clade of the Clavicipitaceae that included the anamorph Chaunopycnis alba and several Cordyceps and Tolypocladium species. A new species of Chaunopycnis is therefore proposed to accommodate MF5785, MF5368, and GB6597. The possible synonymy of Albophoma yamanashiensis with C. alba also is discussed. Within the Clavicipitaceae, indole diterpenoid compounds have only been known from the graminicolous species (subfamily Clavicipitoideae); therefore delineation of a Chaunopycnis clade has revealed a previously unrecognized lineage of indole diterpene-producing fungi among the subfamily Cordycipitoideae.     

水稻内生菠萝泛菌YJ76吲哚产量上调突变株的鉴定及生理性状

菠萝泛菌(Pantoea ananatis)YJ76是从水稻"越富"品种中分离的优势内生菌,与宿主水稻互作时具有多种促生作用,其分泌的吲哚作为细菌种内及种间的信号分子参与调控多种生理生化行为。[目的]筛选获得与吲哚调控相关的突变株,鉴定突变位点并研究突变基因对菌株的生存适应性以及对宿主水稻定殖和促生的影响,为研究吲哚调控通路奠定基础。[方法]用双亲本接合法构建YJ76的mTn5转座子插人突变文库,以染色体步移TAIL-PCR技术鉴定突变基因,最后探究基因突变对菌体产生的影响。[结果]筛选到1株吲哚产量大幅上升的YJ76突变株M04,鉴定突变位点为一个长度195 bp未报道过的新基因,将其命名为ipc(indole production control),基因突变后增强了YJ76对重金属、四环素和酸的抗性,也增强了菌体对宿主水稻定殖和促生的能力。[结论]吲哚产量上调的ipc突变株能够提高菌体生存适应性并增强其对宿主水稻定殖和促生的能力。     

Light-dependent transformation of anthranilate to indole by Rhodobacter sphaeroides OU5

Rhodobacter sphaeroides OU5 transformed anthranilate (2 mM) to an indole (0.7 mM) in a light-dependent process. Photobiotransformation was enhanced by tricarboxylic acid cycle intermediates and the indole formed was identified as 2,3 dihydroxy indole. Journal of Industrial Microbiology & Biotechnology (2000) 24, 219–221. Received 16 September 1999/ Accepted in revised form 20 December 1999     

Molecular characterization of two anthranilate synthase alpha subunit genes in <Emphasis Type="Italic">Camptotheca acuminata</Emphasis>

The potent anticancer and antiviral compound camptothecin (CPT) is a monoterpene indole alkaloid produced by Camptotheca acuminata. In order to investigate the biosynthetic pathway of CPT, we studied the early indole pathway, a junction between primary and secondary metabolism, which generates tryptophan for both protein synthesis and indole alkaloid production. We cloned and characterized the alpha subunit of anthranilate synthase (ASA) from Camptotheca (designated CaASA), catalyzing the first committed reaction of the indole pathway. CaASA is encoded by a highly conserved gene family in Camptotheca. The two CaASA genes are differentially regulated. The level of CaASA2 is constitutively low in Camptotheca and was found mainly in the reproductive tissues in transgenic tobacco plants carrying the CaASA2 promoter and -glucuronidase gene fusion. CaASA1 was detected to varying degrees in all Camptotheca organs examined and transiently induced to a higher level during seedling development. The spatial and developmental regulation of CaASA1 paralleled that of the previously characterized Camptotheca gene encoding the beta subunit of tryptophan synthase as well as the accumulation of CPT. These data suggest that CaASA1, rather than CaASA2, is responsible for synthesizing precursors for CPT biosynthesis in Camptotheca and that the early indole pathway and CPT biosynthesis are coordinately regulated.     

Indole Glucosinolates and Camalexin do not Influence the Development of the Clubroot Disease in Arabidopsis thaliana

Root galls of Brassicaceae caused by Plasmodiophora brassicae are dependent on increased auxin and cytokinin formation. In this study we investigated whether indole glucosinolates are involved in indole‐3‐acetic acid (IAA) biosynthesis in root galls, by using a genetic approach. The cytochrome P450 enzymes, CYP79B2 and CYP79B3, convert tryptophan to indole‐3‐acetaldoxime (IAOx), which is a precursor for indole glucosinolates and the phytoalexin camalexin in Arabidopsis thaliana. Root galls of the Arabidopsis ecotypes Wassilewskija (WS) and Columbia (Col) accumulated camalexin, WS at levels up to 320 μg/g dry weight. By contrast, camalexin was absent in root galls of cyp79b2/b3 double mutants. Infection rate and disease index as a measure of club development in mutant and wild‐type plants of the two ecotypes were investigated and no differences were found in gall formation. This demonstrates that camalexin is an ineffective inhibitor of P. brassicae and indole glucosinolates are not the source of elevated levels of IAA in galls, because free IAA levels in mutant galls were comparable with those in wild type.     

Biodegradation of indole at high concentration by persolvent fermentation with the thermophilic fungus Sporotrichum thermophile

Indole and its derivatives form a class of toxic recalcitrant environmental pollulants. Sporotrichum thermophile was grown in a persolvent fermentation system containing a large amount of indole. The medium contained up to 20% by volume soybean oil and up to 2 g L⁻¹ indole. Most of the indole was partitioned in the organic solvent layer. When the organism was grown in the medium containing indole at 1 g L⁻¹, indole was totally consumed after 6 days. Under a fed–batch fermentation process where daily batches of indole (1 g L⁻¹) supplemented the microbial culture for 4 days, the biodegradation level was 3.0 g L⁻¹. These values make this process promising and worthy of further investigation for the microbial degradation of other toxic compounds.     

Synthesis and Antiviral Evaluation of Some Novel Tricyclic Pyrazolo[3,4‐b]indole Nucleosides

Novel pyrazolo[3,4‐b]indole nucleoside analogs were synthesized from the corresponding 3‐formyl‐2‐chloroindole and 3‐cyano‐2‐chloroindole nucleosides by treatment with hydrazine. Very few examples of pyrazolo[3,4‐b]indole heterocycles have been published in the literature and this is the first synthesis of nucleoside analogs containing this heterocycle. These new pyrazolo[3,4‐b]indole nucleosides were active against human cytomegalovirus and herpes simplex virus type 1, but this activity was not well separated from cytotoxicity.     

Indole glucosinolate breakdown and its biological effects

Most species in the Brassicaceae produce one or more indole glucosinolates. In addition to the parent indol-3-ylmethylglucosinolate (IMG), other commonly encountered indole glucosinolates are 1-methoxyIMG, 4-hydroxyIMG, and 4-methoxyIMG. Upon tissue disruption, enzymatic hydrolysis of IMG produces an unstable aglucone, which reacts rapidly to form indole-3-acetonitrile and indol-3-ylmethyl isothiocyanate. The isothiocyanate, in turn, can react with water, ascorbate, glutathione, amino acids, and other plant metabolites to produce a variety of physiologically active indole compounds. Myrosinase-initiated breakdown of the substituted indole glucosinolates proceeds in a similar manner to that of IMG. Induction of indole glucosinolate production in response to biotic stress, experiments with mutant plants, and artificial diet assays suggest a significant role for indole glucosinolates in plant defense. However, some crucifer-feeding specialist herbivores recognize indole glucosinolates and their breakdown products as oviposition and/or feeding stimulants. In mammalian diets, IMG can have both beneficial and deleterious effects. Most IMG breakdown products induce the synthesis of phase 1 detoxifying enzymes, which may in some cases prevent carcinogenesis, but in other cases promote carcinogenesis. Recent advances in indole glucosinolate research have been fueled by their occurrence in the well-studied model plant Arabidopsis thaliana. Knowledge gained from genetic and biochemical experiments with A. thaliana can be applied to gain new insight into the ecological and nutritional properties of indole glucosinolates in other plant species.     

Plant regeneration from hypocotyls,cotyledons and stem segments of the C3–C4 intermediate species Moricandia arvensis

Techniques have been developed for the regeneration of Moricandia arvensis from complex explants. Hypocotyl segments and cotyledonary explants regenerated shoots, but the most efficient plant regeneration was from stem sections taken from in vitro grown shoots. Regeneration from these three explant types was tested on a range of concentrations of benzylaminopurine and either naphthylene acetic acid or indole acetic acid. Regeneration from all three explants was much higher on indole acetic acid than on naphthylene acetic acid and the ratio of auxin to cytokinin was also significant in determining the response of explants. Optimum regeneration was on 1mg/l IAA with 1mg/l BAP. Plants could be transferred to soil and grown to flowering in the glasshouse.Abbreviations GDC glycine decarboxylase - BAP benzyl aminopurine - NAA naphthalene acetic acid - IAA indole acetic acid