Biological Control of F. Oxysporum F. Sp. Lycopersici Causing Wilt of Tomato by Pseudomonas Fluorescens

Abstract- Pseudomonas fluorescens is one of the major fungal biocontrol agents found in the soil and the rhizosphere of various crop systems. Ten isolates of P.fluorescens were isolated from rhizosphere soil samples collected from various tomato-growing fields and evaluated for their efficacy in increasing seed quality variables of tomato and in inhibiting the mycelial growth of Fusarium oxysporum. Pseudomonas isolate 2 produced effective results and was selected and mass multiplied. Talc and sodium alginate formulations of mass multiplied using different agents were prepared and evaluated for their effects against fusarium wilt under greenhouse conditions. Fresh cultures of Pf2 isolate was found to increase seedling emergence and reduce fusarium wilt disease incidence when compared to the control and the formulations

Establishing inoculum threshold levels for Bean common mosaic virus strain blackeye cowpea mosaic infection in cowpea seed

Bean common mosaic virus strain blackeye cowpea mosaic (BCMV-BlCM) is an important seed-borne virus infecting cowpea and is transmitted both by seeds and aphids. Infected cowpea seeds can act as primary source of inoculum for disease epidemics. Four field experiments were conducted during 2003 - 2006 to assess the role of different amounts of seed-borne inoculum in the dissemination of BCMVBlCM virus in cowpea under field conditions. The identity of BCMV-BlCM was confirmed by ELISA and IC-RT-PCR. Plants infected at an early growth stage appeared to serve as the primary source for subsequent virus spread by aphids. The mean disease incidence during four field experiments reached88-93% in plots sown with 10% infected seed. The disease incidence in plots sown with 5% infected seed recorded 46-63% while for plants raised from 3 and 2% BCMV-BlCM seed infection, disease incidence reached 32-49% and 17-23%, respectively. Mean yield losses in terms of seed yield per plant from four field experiments were 74 and 54% for initial seed infection of 10 and 5%, respectively. Seed infection of 2% BCMV-BlCM incidence resulted in an average of 24% mean seed yield loss/plant-1. The infection appeared to decrease the seed yield in terms of number and size. The BCMV incidence in harvested seed ranged from 0.3 - 19% for the different levels of initial seed infection. The field experiments demonstrated that sowing > 1% BCMV-BlCM infected seed can lead to significant losses in grain yield, while the spread of BCMV-BlCM infection resulting from sowing 1% infected seed did not significantly decrease seed yield. The role of establishing damage or inoculum thresholds from BCMVBlCM seed-borne infections is discussed in the present study.Keywords: Cowpea, potyvirus, seed-borne virus, thresholds, yield los

Lactic Acid Bacteria Mediated Induction of Defense Enzymes To Enhance the Resistance in Tomato against Ralstonia Solanacearum Causing Bacterial Wilt

The biocontrol agent Lactic acid bacterium (LAB) was used against the bacterial wilt caused by Ralstonia solanacearum. The present investigation focuses on the role of defense related enzymes in imparting resistance to tomato plants against R. solanacearum. The LAB isolate was tested for its ability to induce the production of defense-related enzymes in treated tomato seedlings. Tomato seedlings were raised from LAB pretreated seeds, were challenge inoculated with R. solanacearum, harvested at different time intervals (0–72 h) and assayed for defense enzyme activity. The LAB treated seeds showed increase in germination percentage (6%) and seedling vigour index (259) compared with control. Treatment of tomato seedlings with LAB isolate induced a significant amount of Peroxidase (POX), Polyphenol oxidase (PPO), Phenylalanine ammonialyase (PAL), total phenolics and β-1,3-glucanase activities. The activities of PAL, POX, PPO and β-1,3-glucanase reached maximum at 24 h, 24 h, 32 h and 24 h respectively after challenge inoculation. Increased accumulation of phenolics was noticed in plants pre-treated with LAB. Native PAGE analyses of POX and PPO were carried out for the time course of enzyme activities and the isoforms of POX and PPO were examined. In field study, ten isolates of R. solanacearum treated plots yielded an average of 32.4–50 kg/m2 and LAB treated plots an average of 153.5 kg/m2. As compared to the control, LAB increased the yield by 15.3% (8.2 kg/m2) and the pathogen infected plants and pre-treated with LAB gave an average of 55% (28.3 kg/m2 compared to the infected plots). Field experiment results indicated that LAB exhibited 61.1% of disease reduction of bacterial wilt in tomato

Biochemical characterization of Fusarium oxysporum f. sp. cubense isolates from India

The Fusarium wilt caused by Fusarium oxyspoum f. sp. cubense (Foc) is a major biotic constraint for banana production. The characteristics of F. oxyspoum f. sp. cubense isolates were investigated using electrophoretic studies of isozyme and whole-cell protein. The morphological characteristics of the isolates were very similar to each other. All the Foc isolates were pathogenic to banana cultivar 'Nanjangud Rasabale' but they did not induce any disease symptoms on cultivar 'Cavendish'. F. oxyspoum (Isolate 6) did not induce wilt symptoms on either 'Nanjangud' or 'Cavendish' cultivar. Isozyme banding patterns showed 46 scoreable markers and cluster analysis with UPGMA using genetic distance showed that the isolates belonged to three main groups. Group 1 contained isolates 1, 2, 4, 5, 7 and isolate 3 and 6 were placed in group 2 and 3. Results indicated that the estimated intra-specific variation may be more pronounced with isozyme analysis than with protein markers. The level of isozyme variability detected within F. oxysporum f.sp. cubense suggested that it is reliable, efficient and effective in determining genetic relationships among Foc isolates

Immunity elicitors for induced resistance against the downy mildew pathogen in pearl millet

Pearl millet (Pennisetum glaucum (L.) R. Br.) is a globally important cereal whose production is severely constrained by downy mildew caused by Sclerospora graminicola (Sacc.). In this study, immunity eliciting properties of 3,5-dichloroanthranilic acid (DCA), Cell Wall Glucan (CWG), Lipopolysaccharide (LPS), and Glycinebetaine (GB) was deciphered through enzymatic and protein studies based on elicitor treatment activated defense mechanisms. Glycinebetaine, LPS, CWS and DCA elicited enzyme activities and gene expression of the defense enzymes, such as β-1,3-glucanase, phenylalanine ammonia lyase (PAL), peroxidase (POX), polyphenol oxidase (PPO), lipoxygenase (LOX) and defense protein hydroxyproline-rich glycoproteins (HRGPs). However, the speed and the extent of elicitation differed. High levels of enzyme activities and gene expression in elicitor-treated P. glaucum positively correlated with the increased downy mildew resistance. A very rapid and large changes in elicitor-treated seedlings, in contrast to the delayed, smaller changes in the untreated susceptible control seedlings suggests that the rate and magnitude of defense gene expression are important for effective manifestation of defense against pathogen. As compared to other elicitors and control, GB promoted increase in enzyme activities and gene expression, implicating that GB is a promising elicitor of downy mildew resistance in P. glaucum

Fusarium oxysporum f. sp. lycopersici causal agent of vascular wilt disease of tomato: Biology to diversity– A review

Tomato (Lycopersicon esculentum) is one of the widely grown vegetables worldwide. Fusarium oxysporum f. sp. lycopersici (FOL) is the significant contributory pathogen of tomato vascular wilt. The initial symptoms of the disease appear in the lower leaves gradually, trail by wilting of the plants. It has been reported that FOL penetrates the tomato plant, colonizing and leaving the vascular tissue dark brown, and this discoloration extends to the apex, leading to the plants wilting, collapsing and dying. Therefore, it has been widely accepted that wilting caused by this fungus is the result of a combination of various physiological activities, including the accumulation of fungal mycelia in and around xylem, mycotoxin production, inactivation of host defense, and the production of tyloses; however, wilting symptoms are variable. Therefore, the selection of molecular markers may be a more effective means of screening tomato races. Several studies on the detection of FOL have been carried out and have suggested the potency of the technique for diagnosing FOL. This review focuses on biology and variability of FOL, understanding and presenting a holistic picture of the vascular wilt disease of tomato in relation to disease model, biology, virulence. We conclude that genomic and proteomic approachesare greater tools for identification of informative candidates involved in pathogenicity, which can be considered as one of the approaches in managing the disease

Molecular diversity of seed-borne Fusarium species associated with maize in India

A total of 106 maize seed samples were collected from different agro-climatic regions ofIndia. Sixty-two Fusarium isolates were recovered, 90% of which were identified as Fusarium verticillioidesbased on morphological and molecular characters. Use of the tef-1alpha gene corrected/refinedthe morphological species identifications of 11 isolates, and confirmed those of the remaining isolates. Genetic diversityamong the Fusarium isolates involved multilocus fingerprinting profiles by Inter Simple Sequence Repeats (ISSR) UPGMAand tef-1 alpha gene phenetic analyses; for which, we observed no significant differences among the isolates based ongeographic origin or fumonisin production; most of the subdivision related to species. Genotyping was performed on theF. verticillioides isolates, using 12 primer sets from the fumonisin pathway, to elucidate the molecular basis of fumonisinproduction or non-production. One fumonisin-negative isolate, UOMMF-16, was unable to amplify nine of the 12 fumonisincluster genes tested. We also used the CD-ELISA method to confirm fumonisin production for our 62 Fusariumisolates. Only 15 isolates were found to be fumonisin-negative. Interestingly, genotypic characterization revealed six isolateswith various gene deletion patterns that also tested positive for the production of fumonisins via CD-ELISA. Ourfindings confirm the importance of molecular studies for species delimitation, and for observing genetic and phenotypicdiversity, among the Fusaria.</p

Draft genome sequence of Sclerospora graminicola, the pearl millet downy mildew pathogen:Genome sequence of pearl millet downy mildew pathogen

Sclerospora graminicola pathogen is one of the most important biotic production constraints of pearl millet worldwide. We report a de novo whole genome assembly and analysis of pathotype 1. The draft genome assembly contained 299,901,251 bp with 65,404 genes. Pearl millet [Pennisetum glaucum (L.) R. Br.], is an important crop of the semi-arid and arid regions of the world. It is capable of growing in harsh and marginal environments with highest degree of tolerance to drought and heat among cereals (1). Downy mildew is the most devastating disease of pearl millet caused by Sclerospora graminicola (sacc. Schroet), particularly on genetically uniform hybrids. Estimated annual grain yield loss due to downy mildew is approximately 10?80 % (2-7). Pathotype 1 has been reported to be the highly virulent pathotype of Sclerospora graminicola in India (8). We report a de novo whole genome assembly and analysis of Sclerospora graminicola pathotype 1 from India. A susceptible pearl millet genotype Tift 23D2B1P1-P5 was used for obtaining single-zoospore isolates from the original oosporic sample. The library for whole genome sequencing was prepared according to the instructions by NEB ultra DNA library kit for Illumina (New England Biolabs, USA). The libraries were normalised, pooled and sequenced on Illumina HiSeq 2500 (Illumina Inc., San Diego, CA, USA) platform at 2 x100 bp length. Mate pair (MP) libraries were prepared using the Nextera mate pair library preparation kit (Illumina Inc., USA). 1 ?g of Genomic DNA was subject to tagmentation and was followed by strand displacement. Size selection tagmented/strand displaced DNA was carried out using AmpureXP beads. The libraries were validated using an Agilent Bioanalyser using DNA HS chip. The libraries were normalised, pooled and sequenced on Illumina MiSeq (Illumina Inc., USA) platform at 2 x300 bp length. The whole genome sequencing was performed by sequencing of 7.38 Gb with 73,889,924 paired end reads from paired end library, and 1.15 Gb with 3,851,788 reads from mate pair library generated from Illumina HiSeq2500 and Illumina MiSeq, respectively. The sequences were assembled using various assemblers like ABySS, MaSuRCA, Velvet, SOAPdenovo2, and ALLPATHS-LG. The assembly generated by MaSuRCA (9) algorithm was observed superior over other algorithms and hence used for scaffolding using SSPACE. Assembled draft genome sequence of S. graminicola pathotype 1 was 299,901,251 bp long, with a 47.2 % GC content consisting of 26,786 scaffolds with N50 of 17,909 bp with longest scaffold size of 238,843 bp. The overall coverage was 40X. The draft genome sequence was used for gene prediction using AUGUSTUS. The completeness of the assembly was investigated using CEGMA and revealed 92.74% proteins completely present and 95.56% proteins partially present, while BUSCO fungal dataset indicated 64.9% complete, 12.4% fragmented, 22.7% missing out of 290 BUSCO groups. A total of 52,285 predicted genes were annotated using BLASTX and 38,120 genes were observed with significant BLASTX match. Repetitive element analysis in the assembly revealed 8,196 simple repeats, 1,058 low complexity repeats and 5,562 dinucleotide to hexanucleotide microsatellite repeats.publishersversionPeer reviewe

Resultative Compound Verb in Modern Chinese : A Comment on Imai(1985) and Lu(1986)

<p>A. API and DMO suppresses NF-κB DNA binding ability in HCT116 cells. HCT116 cells were treated with DMO and API at indicated doses, nuclear extracts were prepared, and 20 μg of the nuclear extract protein was used for the ELISA-based DNA-binding assay *p<0.05; **p<0.005). B & C. NF-κB responsive elements linked to a luciferase reporter gene were transfected with wild-type or dominant-negative IκB and transfected cancer cells were treated at indicated doses for 6 h and luciferase activity was measured as described in Materials and Methods section. All luciferase experiments were done in triplicate and repeated twice (*p<0.05; **p<0.005). D. API abrogates constitutive IκBα phosphorylation in dose-dependent manner in HCT116 cells. HCT116 cells were treated with different concentrations of API (0, 5, 10 and 20 μM) for 6 h and cytoplasmic extract was prepared. Lysates were resolved on SDS gel and electrotransferred to a nitrocellulose membrane and probed with anti-phospho-IκBα/IκBα. The blot was washed, exposed to HRP-conjugated secondary antibodies for 1 h, and finally examined by chemiluminescence. GAPDH was used as loading control.</p