Grain yield of common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.) varieties is markedly increased by rhizobial inoculation and phosphorus application in Ethiopia

A field experiment was conducted to assess plant growth, symbiotic performance and grain yield of common bean in response to rhizobial incoculation and phosphorus application at Galalicha in Southern Ethiopia during the 2012 and 2013 cropping seasons under rain-fed conditions. The treatments consisted of 2 released common bean varieties (Hawassa Dume and Ibbado), 3 levels of Rhizobium inoculation (uninoculated, inoculated with strain HB-429 or GT-9) and 4 levels of phosphorus application (0, 10, 20 and 30 kg P ha^?1) using a split-split plot design with four replications. Here, phosphorus levels, Rhizobium inoculation and common bean varieties were assigned as main, sub- and sub-sub treatments, respectively. The results revealed marked varietal differences in plant growth, grain yield and symbiotic performance. Of the two common bean varieties studied, Hawassa Dume generally showed superior performance in most measured parameters in 2013. Rhizobium inoculation significantly (p?≤?0.05) increased plant growth, symbiotic performance and grain yield. Applying Rhizobium strain HB-429 to bean crop respectively increased plant growth, %Ndfa, amount of N-fixed and grain yield by 19, 17, 54 and 48% over uninoculated control. Similarly, the application of 20 kg P ha^?1 to bean plants respectively resulted in 36, 20, 96 and 143% increase in plant growth, %Ndfa, N-fixed and grain yield when compared to the control. These results clearly indicate that plant growth, symbiotic performance and grain yield of common bean can be significantly increased by Rhizobium inoculation and phosphorus fertilization in Ethiopia. Rhizobium inoculants are a cheaper source of nitrogen than chemical fertilizers and when combined with moderate phosphorus application can markedly increase grain yield for resource-poor farmers.     

<Emphasis Type="Italic">Rhizobium cellulosilyticum</Emphasis> as a co-inoculant enhances <Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> grain yield under greenhouse conditions

The Rhizobium-legume symbiosis is a complex partnership with many factors, with initial bacterial colonization of the plant root surface and primary infection as key early stages. Two molecules are strongly involved in these processes: the structural carbohydrate cellulose and the enzyme cellulase, which breaks down the former and allows rhizobia to infect the roots. Here, we report the effect on common bean (Phaseolus vulgaris L.) after co-inoculation of the non-nodulating, cellulase-overproducing strain Rhizobium cellulosilyticum ALA10B2^T and the P. vulgaris-nodulating R. leguminosarum strain TPV08. In order to elucidate the effect of combined inoculation with both strains, we designed greenhouse assays, including single inoculation with strain TPV08, co-inoculation with both strains and an uninoculated treatment in non-sterile peat. Chemical fertilizers were not added. Chlorophyll content in the leaves was measured after the flowering stage by spectrophotometry and was considered to be indicative of the nutrient status of the plants. Nodule formation was observed on roots of the inoculated plants, while no nodulation was observed on roots of the uninoculated plants. The results indicate a synergistic effect between the two Rhizobium strains. Co-inoculated plants exhibited significant increases in seed yield and nitrogen content in comparison with the uninoculated control plants and with plants inoculated with a single strain. It is suggested that co-inoculation with strain ALA10B2^T greatly increased the efficiency of N fixation by strain TPV08.     

Analysis of fatty acid composition of PGPR <Emphasis Type="Italic">Klebsiella</Emphasis> sp. SBP-8 and its role in ameliorating salt stress in wheat

A halotolerant plant-growth-promoting rhizobacteria (PGPR) can ameliorate salt stress in associated plants by various mechanisms. Therefore, the present study aimed to characterize a PGPR Klebsiella sp. SBP-8 for its ability to tolerate salt stress and to study the mechanism of PGPR-mediated mitigation of salt stress in the wheat plant. The abiotic stressors result in multiple changes in the fatty acid composition of Klebsiella sp. SBP-8, helping the membrane to keep its integrity, fluidity, and function for its growth under salt (NaCl) stress conditions. The changes in fatty acid composition of test organism were analyzed by fatty acid methyl ester (FAME) analysis under varying saline conditions. The spectroscopy (GC-MS) profile of cell extract at different salt concentrations was comprised of hydrocarbons, and fatty alcohols with varying carbon chain length. Inoculation of Klebsiella sp. SBP-8 to wheat seedling showed increase in proline, total soluble sugar, and total protein content of treated plants. Bacterial inoculation also decreased the concentration of salinity-induced malondialdehyde (MDA) content. In addition, bacterial inoculation also increased the various antioxidative enzymes like superoxide dismutase (SOD), catalase (CAT), and peroxidase (POX) in treated plants. It is likely that bacterial inoculation alleviated the salt stress to wheat plant by co-ordination of antioxidative machinery, and improvement in osmolyte contents. Therefore, the present study suggests that bacterial-inoculated wheat plants were able to cope better with salt stress than uninoculated control, therefore it can serve as a promising bio-inoculant for enhancing the growth of wheat like cereal crops under saline stress.     

Inoculation with indigenous <Emphasis Type="Italic">rhizobium</Emphasis> strains increases yields of common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.) in northern Spain,although its efficiency is affected by the tillage system

Common bean (Phaseolus vulgaris L.) crops hold the potential to obtain higher yields by enhancing their biological nitrogen fixation (BNF) with Rhizobium. However in contrast to other legumes, common bean has shown a lack of positive response to inoculation with Rhizobium in many cases. This has led to a limited use of rhizobial inoculants in this crop, especially in Europe. The adaptation of bacterial strains to the rhizosphere is a key factor in the success of any inoculant, especially in a promiscuous legume such as common bean. This research aimed at increasing common bean yields via inoculation with effective indigenous Rhizobium leguminosarum strains. Three highly effective strains (LCS0306, LBM1123 and ZBM1008) which were selected according to their effectiveness at BNF in hydroponic conditions were separately inoculated onto common bean in a field experiment. The experiment was carried out under three environments and three tillage systems: conventional-tillage (CONVT), no-tillage (NT) and a cover-crop (CC). The grain yield observed with seed inoculation was significantly higher than the yield obtained with uninoculated seed under CONVT and CC. However, under NT inoculation had no effect. Furthermore, under CONVT and CC, inoculation with R. leguminosarum LCS0306 produced even higher yields than those obtained in nitrogen-fertilised or control plots. This is the first attempt to explain the inoculation performance of common bean under different tillage systems in Europe. A gene–based hypothesis has been used to explain the effectiveness of indigenous common bean rhizobia as nitrogen fixers in this crop.     

A novice <Emphasis Type="Italic">Achromobacter</Emphasis> sp. EMCC1936 strain acts as a plant-growth-promoting agent

Fifteen bacterial isolates were isolated from a watering canal at Al Hadady-Damrou, Kafr El-Sheikh Governorate, Egypt (31.3°N 30.93°E). The screening process was achieved based on nitrogenase activity. The most potent bacterial isolate (B9) was tested as plant-growth-promoting rhizobacteria (PGPR). Ultrastructural, cultural, biochemical characteristics and 16S rDNA partial sequence were used for the isolate identification and characterization. From the 16S rRNA gene sequencing results, the nearest bacterial species to our isolate was Achromobacter marplatensis B2 (T), EU150134.1, with 97% matching. The sequence was submitted to the NCBI website with the accession number GenBank: KM491552.1. In vitro analysis revealed that the isolate under study is non-pathogenic (virulence factors-free) and capable of producing indole acetic acid (IAA), gibberellin (GA3) and solubilizing rock phosphate. Under greenhouse conditions, tomato inoculation with the obtained Achromobacter sp. EMCC1936 significantly increased vegetative growth, yield parameters and endogenous phytohormones content as compared with common free diazotrophic PGPR, Azotobacter chroococcum EMCCN1458. It was deposited in Microbiological Resource Center for public use with number (EMCC1936). Data revealed the importance of soil inoculation with the obtained isolate and of its role in increasing soil enzymatic activity. These features fulfill the isolate to be used as a PGPR for various crops.     

<Emphasis Type="Italic">Rhizobium</Emphasis> strains in the biological control of the phytopathogenic fungi <Emphasis Type="Italic">Sclerotium</Emphasis> (<Emphasis Type="Italic">Athelia</Emphasis>) <Emphasis Type="Italic">rolfsii</Emphasis> on the common bean

Aims

To identify Rhizobium strains’ ability to biocontrol Sclerotium rolfsii, a fungus that causes serious damage to the common bean and other important crops, 78 previously isolated rhizobia from common bean were assessed.

Methods

Dual cultures, volatiles, indole-acetic acid (IAA), siderophore production and 16S rRNA sequencing were employed to select strains for pot and field experiments.

Results

Thirty-three antagonistic strains were detected in dual cultures, 16 of which were able to inhibit ≥84% fungus mycelial growth. Antagonistic strains produced up to 36.5 μg mL^?1 of IAA, and a direct correlation was verified between IAA production and mycelium inhibition. SEMIA 460 inhibited 45% of mycelial growth through volatile compounds. 16S rRNA sequences confirmed strains as Rhizobium species. In pot condition, common bean plants grown on S. rolfsii-infested soil and inoculated with SEMIA 4032, 4077, 4088, 4080, 4085, or 439 presented less or no disease symptoms. The most efficient strains under field conditions, SEMIA 439 and 4088, decreased disease incidence by 18.3 and 14.5% of the S. rolfsii-infested control.

Conclusions

Rhizobium strains could be strong antagonists towards S. rolfsii growth. SEMIA 4032, 4077, 4088, 4080, 4085, and 439 are effective in the biological control of the collar rot of the common bean.

     

Molecular diversity of 1-aminocyclopropane-1-carboxylate (ACC) deaminase producing PGPR from wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) rhizosphere

Aims

The present study was planned to investigate the diversity of 1-aminocyclopropane-1-carboxylate (ACC) deaminase producing bacteria from the rhizosphere of wheat plants and subsequent evaluation of selected PGPR on growth enhancement of wheat seedlings under drought and saline conditions.

Methods

ACC deaminase producing plant growth promoting rhizobacteria (PGPR) were isolated from the rhizosphere of wheat and identified using 16S rRNA gene sequence analysis. Isolates were evaluated for various direct and indirect plant growth promoting (PGP) traits. Plant inoculation experiment was conducted using isolates IG 19 and IG 22 in wheat to assess their plant growth promotion potential under salinity and drought stress.

Results

Thirty-eight ACC deaminase producing PGPR were isolated which belonged to 12 distinct genera and falling into four phyla γ-proteobacteria, β-proteobacteria, Flavobacteria and Firmicutes. Klebsiella sp. was the most abundant genera and followed by Enterobacter sp. The isolates exhibited ACC deaminase activities ranging from 0.106–0.980 μM α- ketobutyrate μg protein^?1 h^?1. The isolates showed multiple PGP traits such as IAA production, phosphate, zinc, potassium solubilization and siderophore production. Enterobacter cloacae (IG 19) and Citrobacter sp. (IG 22) inoculated wheat seedlings showed notable increases in fresh and dry biomass under non-stress as well as under stressed condition.

Conclusion

To the best of our knowledge this is the first report of presence of ACC deaminase activity and other PGP traits from the genus Citrobacter and Empedobacter. Our finding revealed that the γ-proteobacteria group dominated the wheat rhizosphere. Plant inoculation with PGPR could be a sustainable approach to alleviate abiotic stresses in wheat plants. These native PGPR isolates could be used as potential biofertilizers for sustainable agriculture.

     

Reduction of soil-borne pathogen <Emphasis Type="Italic">Fusarium solani</Emphasis> reproduction in soil enriched with phenolic acids by inoculation of endophytic fungus <Emphasis Type="Italic">Phomopsis liquidambari</Emphasis>

The increase of soil-borne pathogens induced by phenolic acids that accumulate in continuous cropping soil reduces the yield and quality of crops. The aims of this study were to investigate (i) the biological control of Fusarium solani, in soil enriched with phenolic acids, by the inoculation of the endophytic fungus Phomopsis liquidambari, and (ii) the biocontrol mechanisms involved. Inoculation of P. liquidambari significantly inhibited the reproduction of F. solani. The prompt degradation of soil phenolic acids by P. liquidambari was determined, but no direct antagonism relationship was observed between P. liquidambari and F. solani, implying the alleviated stimulation of phenolic acids was a major factor in controlling F. solani. Moreover, the presence of glucose did not significantly impact the biocontrol function of P. liquidambari, and P. liquidambari inoculation significantly alleviated disease severity of peanut. Therefore, P. liquidambari could be an effective means to control F. solani in phenolic acids-rich continuous cropping soils.     

<Emphasis Type="Italic">Stenotrophomonas maltophilia</Emphasis> HW2 enhanced cucumber resistance against cucumber green mottle mosaic virus

Cucumber green mottle mosaic virus (CGMMV) is a major limiting factor in the production of cucumber plants worldwide. In the present study, we use plant growth-promoting rhizobacteria (PGPR) to control this virus effectively. Stenotrophomonas maltophilia HW2 was isolated from healthy cucumber root, exhibited a good biocontrol efficacy against CGMMV. Here, it is documented that 20 d after virus inoculation, the biocontrol efficacy of HW2 reached 52.61%. HW2 can effectively colonize in cucumber rhizosphere, and also promoted cucumber plants growth. We also examined the effect of HW2 on viral replication and its mechanism. Compared with the control, HW2 pre-treated plants could delay virus replication for more than 3 d and inhibit viral protein genes (CP, MP, Rep) expression in the cucumber leaf. The expression of antioxidant enzyme genes (SOD and CAT) and defense-related genes (PR1 and PR5) were quickly induced by HW2. These results suggest that HW2 induced plant defense responses to CGMMV by increasing the expression of defense response genes. We report for the first time that Stenotrophomonas maltophilia improved cucumber resistance against CGMMV, which highlights the applying of PGPR on controlling of virus diseases.     

Functional Analysis of <Emphasis Type="Italic">VvBG1</Emphasis> During Fruit Development and Ripening of Grape

β-glucosidase (BG) was believed to take part in abscisic acid (ABA) synthesis via hydrolysis of ABA glucose ester to release active ABA during plant growth and development. However, there is no genetic evidence available to indicate the role of genes during fruit ripening. Here, the expression patterns of three genes (VvBG1, VvBG2, and VvBG3) encoding β-glucosidase were analyzed during grape fruit development, and it was found that β-glucosidase activity increased in grape fruit in response to various stresses. Furthermore, to verify the function of β-glucosidase during fruit ripening, heterogeneous expression of the VvBG1 gene in strawberry fruit was validated, and the results showed that the VvBG1 over-expression increased β-glucosidase and promoted the fruit ripening process in strawberry. In addition, we found that ABA contents increased in the VvBG1 over-expression of strawberry fruit, which induced fruit anthocyanin, soluble solid accumulation, and fruit softening. Moreover, genes related to coloring (CHS, CHI, F3H, and UFGT), softening (PG1, PL1, and EXP1), and aroma (SAAT, and QR) were up-regulated. This work will elucidate the specific roles of VvBGs in the synthesis of ABA and provide some new insights into the ABA-controlled grape ripening mechanism.     

A new method of propagation of arbuscular mycorrhizal fungi in field cropped sesame (<Emphasis Type="Italic">Sesamum indicum</Emphasis> L.)

Though arbuscular mycorrhizal (AM) fungi are indigenous to agricultural soils, their beneficial effects to host plants could be further improved by inoculation with efficient species. The method of AM propagation described in the present study uses oil cake as a supporting medium for the simultaneous delivery of sesame seeds and AM inoculum to the field. Experiment was conducted in a farmer’s field located at Avoor, Kerala, India where sesame was cultivated as a winter crop in rice fallows. Oil cake entrapped with sesame seeds (var. Tilatara) and AM fungus (Funneliformis dimorphicus) inoculum was prepared by thoroughly mixing sterilized coconut cake and neem cake (5:1 v/v), surface sterilized sesame seeds and sterilized spore sieving of F. dimorphicus from a pot culture in a 10% solution of a polysaccharide gum obtained from the seeds of Strychnos potatorum L. Entire mix was moulded into 2.5 cm cubes (ca. 5g) containing approximately 25–30 seeds and 200–300 spores cube^?1 and shade dried before application. The cubes were broadcast @ 600 kg ha^?1 in inoculated treatments. In uninoculated treatments, the oil cake cubes devoid of the fungal component was used. Harvested root samples from the inoculated treatments showed a high frequency (%F) and intensity (%M) of colonization by AM fungi as well as frequency of vesicles (%V) and arbuscules (%A) compared to uninoculated control. The growth (root length, shoot length and leaf area) and yield characters (pod number, seed number, seed weight and oil content) of sesame plants were significantly (p=0.05) improved under the present method of AM propagation indicating its viability under field condition.     

Degradation and polymerization of monolignols by <Emphasis Type="Italic">Abortiporus biennis</Emphasis>, and induction of its degradation with a reducing agent

This study was carried out to better understand the characteristic modification mechanisms of monolignols by enzyme system of Abortiporus biennis and to induce the degradation of monolignols. Degradation and polymerization of monolignols were simultaneously induced by A. biennis. Whole cells of A. biennis degraded coniferyl alcohol to vanillin and coniferyl aldehyde, and degraded sinapyl alcohol to 2,6-dimethoxybenzene- 1,4-diol, with the production of dimers. The molecular weight of monolignols treated with A. biennis increased drastically. The activities of lignin degrading enzymes were monitored for 24 h to determine whether there was any correlation between monolignol biomodification and ligninolytic enzymes. We concluded that complex enzyme systems were involved in the degradation and polymerization of monolignols. To degrade monolignols, ascorbic acid was added to the culture medium as a reducing agent. In the presence of ascorbic acid, the molecular weight was less increased in the case of coniferyl alcohol, while that of sinapyl alcohol was similar to that of the control. Furthermore, the addition of ascorbic acid led to the production of various degraded compounds: syringaldehyde and acid compounds. Accordingly, these results demonstrated that ascorbic acid prevented the rapid polymerization of monolignols, thus stabilizing radicals generated by enzymes of A. biennis. Thereafter, A. biennis catalyzed the oxidation of stable monolignols. As a result, ascorbic acid facilitated predominantly monolignols degradation by A. biennis through the stabilization of radicals. These findings showed outstanding ability of A. biennis to modify the lignin compounds rapidly and usefully.     

Identification of the di-<Emphasis Type="Italic">n</Emphasis>-butyl phthalate-biodegrading strains and the biodegradation pathway in strain LMB-1

DNA isolated from a greenhouse soil (Nanjing, Jiangsu Province, China) was suitable for PCR amplification of gene segment coding for the 16S rRNA. Diverse PCR products were characterized by cloning and sequencing, and analysis of bacterial colonies showed the presence over 26 phyla. The most bacteria belonged to Proteobacteria, Actinobacteria, Gemmatimonadetes, Acidobacteria and Planctomycetes. Furthermore, after the enrichment procedure of DBP-degrading microorganisms, 4 strains were isolated from the soil sample with di-n-butyl phthalate (DBP) biodegradability, and they were identified to be Rhizobium sp., Streptomyces sp., Pseudomonas sp. and Acinetobacter sp. Analysis of the degradation products by LC-MS led to identification of metabolites of DBP in strain LMB-1 (identified as Rhizobium sp.) which suggests that DBP was degraded through β-oxidation, demethylation, de-esterification and cleavage of aromatic ring.