Influence of plant growth-promoting endophytes Colletotrichum siamense and Diaporthe masirevici on tomato plants (Lycopersicon esculentum Mill.).

The protective and growth-promoting activities of Colletrotrichum and Diaporthe endophytes on tomato plants (Lycopersicon esculentum Mill.) are underexplored. We screened 40 endophytic fungi associated with Mexican shrimp plant (Justicia brandegeana) using an in vitro dual culture assay for Fusarium oxysporum, one of the most important phytopathogens of tomato plants. The three best antagonists, Colletotrichum siamense (JB224.g1), C. siamense (JB252.g1), and Diaporthe masirevicii (JB270), were identified based on multilocus sequence analysis. They were assessed in vitro for their inhibition of F. oxysporum and phosphate solubilisation capacity, and for the production of indole acetic acid. Greenhouse experiments verified the growth-promoting effects of these endophytes and the suppression of F. oxysporum symptoms in tomato plants.   Under greenhouse conditions, the JB252.g1 and JB270 isolates showed positive results for seedling emergence speed. The radicular system depth of plants inoculated with JB270 was greater than that in uninoculated plants (27.21 vs 21.95 cm). The soil plant analysis development chlorophyll metre (SPAD) index showed statistically significant results, especially for the endophyte JB224.g1 (36.99) compared to the control plants (30.90) and plants infected solely with F. oxysporum (33.64).

Metagenomic analyses, isolation and characterization of endophytic bacteria associated with Eucalyptus urophylla BRS07-01 in vitro plants.

Eucalyptus is the main species for the forestry industry in Brazil. Biotechnology and, more recently, gene editing offer significant opportunities for rapid improvements in Eucalyptus breeding programs. However, the recalcitrance of Eucalyptus species to in vitro culture is also a major limitation for commercial deployment of biotechnology techniques in Eucalyptus improvement. We evaluated various clones of Eucalyptus urophylla for their in vitro regeneration potential identified a clone, BRS07-01, with considerably higher regeneration rate (85%) in organogenesis, and significantly higher than most works described in literature. Endophytic bacteria are widely reported to improve in vitro plant growth and development. Hence, we believe that inclusion of endophytic plant growth promoting bacteria enhanced was responsible for the improved plantlets growth and development of this clone under in vitro culture. Metagenomic analysis was performed to isolate and characterize the prominent endophytic bacteria on BRS07-01 leaf tissue in vitro micro-cultures, and evaluate their impact on plant growth promotion. The analysis revealed the presence of the phyla Firmicutes (35%), Proteobacteria (30%) and much smaller quantities of Actinobacteria, Bacteroidetes, Gemmatimonadetes, Crenarchaeota, Euryarchaeota and Acidobacteria. Of the thirty endophytic bacterial strains isolated, eleven produced indole-3-acetic acid. Two of the isolates were identified as Enterobacter sp. and Paenibacillus polymyxa, which are nitrogen-fixing and capable of phosphate and produce ammonium. These isolates also showed similar positive effects on the germination of common beans (Phaseolus spp.). The isolates will now be tested as a growth promoter in Eucalyptus in vitro cultures. Graphical abstract for the methodology using cultivation independent and dependent methodologies to investigate the endophytic bacteria community from in vitro Eucalyptus urophylla BRS07-01.

Optimization of theoretical maximal quantity of cells to immobilize on solid supports in the rational design of immobilized derivatives strategy.

Current worldwide challenges are to increase the food production and decrease the environmental contamination by industrial emissions. For this, bacteria can produce plant growth promoter phytohormones and mediate the bioremediation of sewage by heavy metals removal. We developed a Rational Design of Immobilized Derivatives (RDID) strategy, applicable for protein, spore and cell immobilization and implemented in the RDID1.0 software. In this work, we propose new algorithms to optimize the theoretical maximal quantity of cells to immobilize (tMQCell) on solid supports, implemented in the RDIDCell software. The main modifications to the preexisting algorithms are related to the sphere packing theory and exclusive immobilization on the support surface. We experimentally validated the new tMQCell parameter by electrostatic immobilization of ten microbial strains on AMBERJET® 4200 Cl- porous solid support. All predicted tMQCell match the practical maximal quantity of cells to immobilize with a 10% confidence. The values predicted by the RDIDCell software are more accurate than the values predicted by the RDID1.0 software. 3-indolacetic acid (IAA) production by one bacterial immobilized derivative was higher (~ 2.6 µg IAA-like indoles/108 cells) than that of the cell suspension (1.5 µg IAA-like indoles/108 cells), and higher than the tryptophan amount added as indole precursor. Another bacterial immobilized derivative was more active (22 µg Cr(III)/108 cells) than the resuspended cells (14.5 µg Cr(III)/108 cells) in bioconversion of Cr(VI) to Cr(III). Optimized RDID strategy can be used to synthesize bacterial immobilized derivatives with useful biotechnological applications.

Genetic and Phenotypic Characterization of Indole-Producing Isolates of Pseudomonas syringae pv. actinidiae Obtained From Chilean Kiwifruit Orchards.

In recent years, Chilean kiwifruit production has been affected by the phytopathogen Pseudomonas syringae pv. actinidiae (Psa), which has caused losses to the industry. In this study, we report the genotypic and phenotypic characterization of 18 Psa isolates obtained from Chilean kiwifruits orchards between 2012 and 2016 from different geographic origins. Genetic analysis by multilocus sequence analysis (MLSA) using four housekeeping genes (gyrB, rpoD, gltA, and gapA) and the identification of type III effector genes suggest that the Chilean Psa isolates belong to the Psa Biovar 3 cluster. All of the isolates were highly homogenous in regard to their phenotypic characteristics. None of the isolates were able to form biofilms over solid plastic surfaces. However, all of the isolates formed cellular aggregates in the air-liquid interface. All of the isolates, except for Psa 889, demonstrated swimming motility, while only isolate Psa 510 demonstrated swarming motility. The biochemical profiles of the isolates revealed differences in 22% of the tests in at least one Psa isolate when analyzed with the BIOLOG system. Interestingly, all of the isolates were able to produce indole using a tryptophan-dependent pathway. PCR analysis revealed the presence of the genes aldA/aldB and iaaL/matE, which are associated with the production of indole-3-acetic acid (IAA) and indole-3-acetyl-3-L-lysine (IAA-Lys), respectively, in P. syringae. In addition, IAA was detected in the cell free supernatant of a representative Chilean Psa strain. This work represents the most extensive analysis in terms of the time and geographic origin of Chilean Psa isolates. To our knowledge, this is the first report of Psa being able to produce IAA. Further studies are needed to determine the potential role of IAA in the virulence of Psa during kiwifruit infections and whether this feature is observed in other Psa biovars.

Assessment of Rice Associated Bacterial Ability to Enhance Rice Seed Germination and Rice Growth Promotion

ABSTRACT The application of beneficial bacteria has recently been used for sustainable agriculture. In current research, 71 bacterial isolates were obtained from rice plant and the rhizosphere soil of different paddy fields in Guilan province, Iran. After primitive investigation, 40 bacteria with typical predominant characteristics were selected. By PCR-RFLP of their 16S r-DNA gene, 8 Operational Taxonomic Units (OTUs) totally consisted of 33 isolates were obtained. From all of them, 8 isolates were selected for rice seed germination experiment, then, effective isolates were used for pot experiment to evaluate their ability for promoting rice growth. All of them were able to increase rice growth and yield, but in different potential. These tested isolates were identified as Alcaligenes faecalis (DEp8, O1R4), Pantoea ananatis (AEn1), Bacillus vietnamensis (MR5), Bacillus idriensis (MR2) and Stenotrophomonas maltophilia by partial sequencing of their 16S r-DNA gene. Among them, AEn1 and MR5 produced indole-3- acetic acid (IAA) in larger amounts than the other isolates and the isolates AEn1 and O1R4 were able to solubilize phosphate in higher amounts. According to the results obtained, it can be concluded that AEn1, O1R4 and MR5 can be considered as bacterial inoculants to use as alternatives for chemical fertilizers.

Isolation and identification by 16S rRNA sequence analysis of plant growth-promoting azospirilla from the rhizosphere of wheat

ABSTRACT The main objective of the present study was to isolate phytohormone-producing, phosphate-solubilizing strains of Azospirillum from wheat to be used as inoculants for plant growth promotion. Five Azospirillum strains were isolated from the rhizosphere of field-grown wheat (Triticum aestivum L.), and it was confirmed by BOX-polymerase chain reaction (PCR) that the isolates were different and not re-isolates of the same strain. Sequence analysis of the PCR-amplified 16S rRNA gene indicated that four isolates showed maximum similarity to Azospirillum brasilense and one isolate showed maximum similarity to Azospirillum zeae. This is the first report indicating the presence of an A. zeae like isolate in the wheat rhizosphere in Pakistan. The bacterial isolates were characterized for their plant growth-promoting traits, phosphate solubilization, and indole-3-acetic acid (IAA) production. None of the isolates showed phosphate solubilization activity in the commonly used Pikovskaya medium. However, all strains (except AzoK4) exhibited ability to solubilize tricalcium phosphate (TCP) in modified Pikovskaya medium in which sucrose was replaced by Na-malate, as well as in TCP-supplemented Luria-Bertani (LB) medium. Organic acids, such as acetic, citric, lactic, malic, and succinic acids, were detected in culture supernatants of the tested Azospirillum strains. All strains exhibited ability to produce IAA in the growth medium, except Azospirillum sp. AzoK1. Among the strains tested, the maximum IAA production (30.49 ± 1.04 mg L-1) and phosphate solubilization (105.50 ± 4.93 mg L-1) were shown by a pure culture of Azospirillum sp. AzoK2. In pot experiments, single-strain inocula of Azospirillum sp. AzoK1 and AzoK2 improved wheat plant growth.

Isolation and identification by 16S rRNA sequence analysis of plant growth-promoting azospirilla from the rhizosphere of wheat

The main objective of the present study was to isolate phytohormone-producing, phosphate-solubilizing strains of Azospirillum from wheat to be used as inoculants for plant growth promotion. Five Azospirillum strains were isolated from the rhizosphere of field-grown wheat (Triticum aestivum L.), and it was confirmed by BOX-polymerase chain reaction (PCR) that the isolates were different and not re-isolates of the same strain. Sequence analysis of the PCR-amplified 16S rRNA gene indicated that four isolates showed maximum similarity to Azospirillum brasilense and one isolate showed maximum similarity to Azospirillum zeae. This is the first report indicating the presence of an A. zeae like isolate in the wheat rhizosphere in Pakistan. The bacterial isolates were characterized for their plant growth-promoting traits, phosphate solubilization, and indole-3-acetic acid (IAA) production. None of the isolates showed phosphate solubilization activity in the commonly used Pikovskaya medium. However, all strains (except AzoK4) exhibited ability to solubilize tricalcium phosphate (TCP) in modified Pikovskaya medium in which sucrose was replaced by Na-malate, as well as in TCP-supplemented Luria-Bertani (LB) medium. Organic acids, such as acetic, citric, lactic, malic, and succinic acids, were detected in culture supernatants of the tested Azospirillum strains. All strains exhibited ability to produce IAA in the growth medium, except Azospirillum sp. AzoK1. Among the strains tested, the maximum IAA production (30.49 ± 1.04 mg L-1) and phosphate solubilization (105.50 ± 4.93 mg L-1) were shown by a pure culture of Azospirillum sp. AzoK2. In pot experiments, single-strain inocula of Azospirillum sp. AzoK1 and AzoK2 improved wheat plant growth.(AU)

Isolation and identification by 16S rRNA sequence analysis of plant growth-promoting azospirilla from the rhizosphere of wheat.

The main objective of the present study was to isolate phytohormone-producing, phosphate-solubilizing strains of Azospirillum from wheat to be used as inoculants for plant growth promotion. Five Azospirillum strains were isolated from the rhizosphere of field-grown wheat (Triticum aestivum L.), and it was confirmed by BOX-polymerase chain reaction (PCR) that the isolates were different and not re-isolates of the same strain. Sequence analysis of the PCR-amplified 16S rRNA gene indicated that four isolates showed maximum similarity to Azospirillum brasilense and one isolate showed maximum similarity to Azospirillum zeae. This is the first report indicating the presence of an A. zeae like isolate in the wheat rhizosphere in Pakistan. The bacterial isolates were characterized for their plant growth-promoting traits, phosphate solubilization, and indole-3-acetic acid (IAA) production. None of the isolates showed phosphate solubilization activity in the commonly used Pikovskaya medium. However, all strains (except AzoK4) exhibited ability to solubilize tricalcium phosphate (TCP) in modified Pikovskaya medium in which sucrose was replaced by Na-malate, as well as in TCP-supplemented Luria-Bertani (LB) medium. Organic acids, such as acetic, citric, lactic, malic, and succinic acids, were detected in culture supernatants of the tested Azospirillum strains. All strains exhibited ability to produce IAA in the growth medium, except Azospirillum sp. AzoK1. Among the strains tested, the maximum IAA production (30.49±1.04mgL(-1)) and phosphate solubilization (105.50±4.93mgL(-1)) were shown by a pure culture of Azospirillum sp. AzoK2. In pot experiments, single-strain inocula of Azospirillum sp. AzoK1 and AzoK2 improved wheat plant growth.