Post-emergence Damping-off Research Articles

Root Rot Management in Common Bean (Phaseolus vulgaris L.) Through Integrated Biocontrol Strategies using Metabolites from Trichoderma harzianum, Serratia marcescens, and Vermicompost Tea

Common bean (Phaseolus vulgaris L.) is an essential food staple and source of income for small-holder farmers across Africa. However, yields are greatly threatened by fungal diseases like root rot induced by Rhizoctonia solani. This study aimed to evaluate an integrated approach utilizing vermicompost tea (VCT) and antagonistic microbes for effective and sustainable management of R. solani root rot in common beans. Fourteen fungal strains were first isolated from infected common bean plants collected across three Egyptian governorates, with R. solani being the most virulent isolate with 50% dominance. Subsequently, the antagonistic potential of vermicompost tea (VCT), Serratia sp., and Trichoderma sp. was assessed against this destructive pathogen. Combinations of 10% VCT and the biocontrol agent isolates displayed potent inhibition of R. solani growth in vitro, prompting in planta testing. Under greenhouse conditions, integrated applications of 5 or 10% VCT with Serratia marcescens, Trichoderma harzianum, or effective microorganisms (EM1) afforded up to 95% protection against pre- and post-emergence damping-off induced by R. solani in common bean cv. Giza 6. Similarly, under field conditions, combining VCT with EM1 (VCT + EM1) or Trichoderma harzianum (VCT + Trichoderma harzianum) substantially suppressed disease severity by 65.6% and 64.34%, respectively, relative to untreated plants. These treatments also elicited defense enzyme activity and distinctly improved growth parameters including 136.68% and 132.49% increases in pod weight per plant over control plants. GC–MS profiling of Trichoderma harzianum, Serratia marcescens, and vermicompost tea (VCT) extracts revealed unique compounds dominated by cyclic pregnane, fatty acid methyl esters, linoleic acid derivatives, and free fatty acids like oleic, palmitic, and stearic acids with confirmed biocontrol and plant growth-promoting activities. The results verify VCT-mediated delivery of synergistic microbial consortia as a sustainable platform for integrated management of debilitating soil-borne diseases, enhancing productivity and incomes for smallholder bean farmers through regeneration of soil health. Further large-scale validation can pave the adoption of this climate-resilient approach for securing food and nutrition security.

Interacting effects of botanicals, biocontrol agents, and potting media on Rhizoctonia solani led damping-off of okra seedlings

The interacting effects of botanicals, biocontrol agents, and potting media against the Rhizoctonia solani led damping-off in okra ( Abelmoschus esculentus L.) were investigated. The treatments comprised of different combinations of extracts of five botanicals:wood apple/Indian bael – Aegle marmelos L., Neem – Azadirachta indica A. Juss, Turmeric – Curcuma longa L., Ginger – Zingiber officinale Roscoe, and Onion – Allium cepa L.,and two biocontrol agents, Pseudomonas fluorescens and Trichoderma viride as first factors and two potting media; vermicompost with soil and farm yard manure with soil as the second factor in a completely randomized design with 42 treatments and 4 replications. The seeds were sown after pathogen inoculation. Okra seedling health was recorded daily for the first 16 days after sowing. Pseudomonas soil treatment and Pseudomonas+onion seed treatment showed the least pre-emergence damping off (35%). Almost all the combinations of wood apple extracts showed inhibitory effects on seed germination and growth of seedlings. The greatest number of days for the onset of post-emergence damping-off was recorded with Pseudomonas + Onion seed treatment (14.88 days) followed by Pseudomonas seed treatment (14.13 days). Germination Index (GI) was better for farm yard manure(0.32) than Vermicompost substrates (0.23). The GI was greatest in Pseudomonas soil treatment (0.53) followed by Pseudomonas + Onion seed treatment (0.51). Similarly, Dickson Quality Index (DQI) was greatest for Pseudomonas + Neem seed treatment (0.0015), with a greater value of DQI indicating better seedling quality. No significant interaction was observed between the substrates and biocontrol agents for any studied parameters. Pseudomonas + Onion seed treatment and Pseudomonas soil treatment were most effective for controlling R. solani -led damping-off in okra. • We studied the control of damping-off disease in okra seedlings. • Seed treatment with Pseudomonas fluorescens and onion blub extract were effective. • Trichoderma treatment in combination with other botanicals decreased the efficacy. • Hard wood apple extract had inhibitory effects on the germination of okra.

Evaluation of Damping-Off Tolerance in Spinach Cultivars in Field Soils and in a Standardized Lab Assay with Pythium ultimum

Spinach growers face increasing problems of damping-off in the production of fresh-market (baby-leaf) spinach due to increasing restrictions on chemical treatments. Damping-off-tolerant cultivars are increasingly important, requiring proper evaluation methods. From three locations in The Netherlands with a history of spinach cultivation and from one location in France, potential damping-off pathogens were isolated from the soil, identified to species or genus, and tested for their pathogenicity. Pythium ultimum was the most prevalent pathogen in those fields, causing spinach pre- and postemergence damping-off. Eight spinach cultivars with two or three seed lots each were evaluated at the same field locations and in a greenhouse with soil sampled from one of the Dutch field sites. Preemergence damping-off was more discriminating for differences among the cultivars than postemergence damping-off. Variation in levels of infection among trials, replicates, and seed lots of the same cultivars emphasized the need for a more standardized phenotyping assay. For such an assay, a cornmeal-sand-based inoculum with a pathogenic P. ultimum isolate was added to a substrate mixture of sand, perlite, and vermiculite and moistened until 50% water-holding capacity, in which spinach seeds were incubated for 10 days in a dark climate cabinet at 15°C, including a control treatment without P. ultimum inoculum. The assay showed reproducible results for discriminating differences in preemergence damping-off tolerance levels among seed lots. However, cultivar differences in preemergence damping-off tolerance levels could not be confirmed due to a large variation among seed lots that needs further investigation.

Variations in Pathogenicity of Three Different Forms of Rhizoctonia Inoculum and Assessment of Cultivar Resistance of Sugar Beet

Rhizoctonia solani causes pre-emergence and post-emergence damping-off, as well as crown and root rot of sugar beet (Beta vulgaris L.), which significantly affects the yield returns in the USA and Europe. The pathogen overwinters as sclerotia or melanized mycelium. Traditionally, the resistance of cultivars to R. solani is evaluated by scoring disease reactions at the crowns and roots of older seedlings, thus resistance is not evaluated during seed germination. Moreover, earlier studies evaluated cultivars resistance to R. solani using colonized whole barley or wheat grains which, unlike sclerotia, are artificial inocula of the pathogen that require time, space and technical know-how to produce. Moreover, colonized grains are prone to contamination with other pathogens, consumed by rodents/birds while applied in the field, and are often uneconomic. Considering those limitations, a study was undertaken (1) to develop in vitro methods to generate large-scale sclerotia, (2) to compare pathogenic potentials of sclerotia, mycelia, and colonized barley grains for optimization of dampingoff assays, and (3) to evaluate Rhizoctonia resistance of selected commercial cultivars during the seed germination phase. Comparing six different culture media, we found that R. solani had the highest radial growth (8.9 ± 0.04, cm³) at 8-days and the maximum number of sclerotia produced (203 ± 4.6) at 28-days in CV8 medium. We demonstrated significant differences in pathogenicity of the three different forms of R. solani inocula and susceptibility of cultivars to preand post-emergence damping-off. The highest pre-emergence damping-off and root rot were observed with sclerotia, and the highest post-emergence dampingoff was recorded with both sclerotial and colonized barley inocula. In addition, varietal differences in susceptibility to pre- and post-emergence damping-off were noted. The highest pre-emergence damping-off was recorded on cv Crystal 101RR and lowest in Maribo MA 504. The highest post-emergence damping-off was recorded on BTS 8500 and the lowest in Crystal 467. The maximum mean root rot was observed in BTS 8500, BTS 8606, and Crystal 101R. Our studies demonstrated that sclerotia serve as efficient natural inocula, reemphasized that host-pathogen interactions differ at the early vs. late stages of sugar beet growth, and highlighted the need to reevaluate commercial sugar beet cultivars for resistance at the seed germination stage.

First report of Pythium ultimum causing damping-off of Sugar beet (Beta vulgaris. L) in Montana, USA.

Sugar beet (Beta vulgaris L.) is a globally important crop for sugar. In May 2019, sugar beet seedlings were observed with wilting, lodging and a few were dead in Glendive (46.970170, -104.838204), Montana. Symptoms appeared near the soil line as the stem (hypocotyl) turned dark brown to black with characteristic thread-like infections which resembled Pythium damping-off. It affected approximately 10% of the growing seedlings. Diseased sugar beet root tissues were excised with a sterile scalpel and small pieces (10 mm²) were surface sterilized with 70 % ethanol for 30 seconds, rinsed twice with autoclaved water, air-dried and transferred to potato dextrose agar (PDA) media amended with pimaricin-vancomycin-PCNB (Conway, 1985). Four plates were incubated at 25° C in the dark (Masago et al., 1977) and two weeks later white, dense colony was observed (Zhang et al., 2018). The terminal smooth, globose oogonia (average 18.5 µm in diameter) and antheridia (average 14.5 × 9.5 µm) extended below the oogonium were observed via VWR N. A. 0.30 microscope. The morphological features of the four isolates were consistent with Pythium ultimum Trow (Watanabe, 2002). Genomic DNAs (NORGEN BIOTEK CORP, Fungi DNA Isolation Kit #26200) of four isolates were used for polymerase chain reaction (PCR) with the ITS6-ITS7 primers (Taheri et al., 2017). Subsequently, PCR products were flushed by E.Z.N.A ®Cycle Pure Kit, OMEGA and four samples were sent for Sanger sequencing to GenScript (GenScript, Piscataway, NJ). The sequences were identical and submitted to GenBank, NCBI (accession no. MN398593). The NCBI Blast analysis showed 100% sequence homology to Pythium ultimum with the following GenBank accessions; KF181451.1, KF181449.1 and AY598657.2. Pathogenicity test was done on sugar beet with the same isolates in the greenhouse. Two week old, pythium culture was mixed with vermiculite and perlite mixer (PRO-MIX FLX) in the plastic trays (24´´ x 15´´× 3˝), (22 °C, 75% Relaive Humidity). Sterile water (500 ml/each tray) was added in the mixer to provide sufficient moisture. Twenty seeds of cv. Hilleshog 4302 were sown in the tray, and the trays were replicated thrice with inoculated and mock treatments. Plants were watered as needed to maintain adequate soil moisture conducive for plant growth and disease development. Seven days after sowing, 50% and 100% germination was observed in the inoculated and control treatments, respectively. At the beginning of the second week, 30% post-emergence damping-off was observed in the inoculated treatments. Diseased seedlings were gently pulled out from the pots where similar symptoms were observed in the sugar beet seedlings as described previously. No incidence of disease was observed in mock-treated seedlings. Consistent reisolation of Pythium ultimum was morphologically and molecularly confirmed from the diseased seedlings, thus fulfilling Koch's postulates. Pythium spp identification is prerequisite to develop effective management of pre and post-emergence damping-off. Pythium ultimum was previously reported in Nebraska to cause sugar beet seed rot and pre-emergence damping-off (Harvenson 2006). To our knowledge, this is the first report of Pythium ultimum causing damping-off on sugar beet in the Sidney factory district in Montana.

Characterization of native plant growth promoting rhizobacteria and their anti-oomycete potential against Phytophthora capsici affecting chilli pepper (Capsicum annum L.)

Phytophthora capsici is a notorious fungus which infects many crop plants at their early and late growth stages. In the present study, twelve P. capsici isolates were morphologically characterized, and based on pathogenicity assays; two highly virulent isolates causing post-emergence damping-off on locally cultivated chilli pepper were screened. Two P. capsici isolates, HydPak1 (MF322868) and HydPk2 (MF322869) were identified based on internal transcribed spacer (ITS) sequence homology. Plant growth promoting rhizobacteria (PGPR) play a significant role in disease suppression and plant growth promotion in various crops. Out of fifteen bacterial strains recovered from chilli rhizosphere, eight were found potential antagonists to P. capsici in vitro. Bacterial strains with strong antifungal potential were subjected to biochemical and molecular analysis. All tested bacterial strains, were positive for hydrogen cyanide (HCN), catalase production and indole-3-acetic acid (IAA) production (ranging from 6.10 to 56.23 µg ml−1), while siderophore production varied between 12.5 and 33.5%. The 16S rRNA sequence analysis of tested bacterial strains showed 98–100% identity with Pseudomonas putida, P. libanensis, P. aeruginosa, Bacillus subtilis, B. megaterium, and B. cereus sequences available in the National Center for Biotechnology Information (NCBI) GenBank nucleotide database. All sequences of identified bacteria were submitted to GenBank for accessions numbers (MH796347-50, MH796355-56, MH801129 and MH801071). Greenhouse studies concluded that all tested bacterial strains significantly suppressed the P. capsici infections (52.3–63%) and enhanced the plant growth characters in chilli pepper. Efficacy of many of these tested rhizobacteria is being first time reported against P. capsici from Pakistan. Plant growth promoting rhizobacteria (PGPR) exhibiting multiple traits may be used in the development of new, eco-friendly, and effective bioformulations as an alternative to synthetic fungicides.

Effects of Soil Conditions on Root Rot of Soybean Caused by Fusarium graminearum.

Fusarium graminearum is an important soybean pathogen that causes seedling disease, root rot, and pre- and postemergence damping-off. However, effects of soil conditions on the disease are not well understood. The objective of this greenhouse study was to determine the impacts of soil texture, pH, and soil moisture on seedling root rot symptoms and detrimental effects on seedling development caused by F. graminearum. F. graminearum-infested millet was added (10%, vol/vol) to soil with four different textures (sand, loamy sand, sandy loam, and loam). Soil moisture was maintained at saturation, field capacity or permanent wilting point at soil pH levels of 6 or 8. Seedlings were evaluated 4 weeks after planting for root rot, root length, root and shoot dry weights, leaf area, and F. graminearum colonization (by qPCR). There was a significant interaction between soil moisture and soil texture for root rot assessed visually (P < 0.0001). Highest severity (67%) and amount of F. graminearum DNA were observed at pH 6 and permanent wilting point in sandy loam soils. Pot saturation resulted in the lowest levels of disease in sandy loam and loam soils (11.6 and 10.8%, respectively). Reductions in seedling growth parameters, including root length, foliar area, shoot and root dry weights, and root tips, relative to the noninfested control, were significantly greater in sandy loam soils. In contrast, there were no significant growth reductions in sand. This study showed that levels of root rot increased under moisture-limiting conditions, producing detrimental effects on plant development.

A Review and Perspective on Soybean (Glycine maxL.) Breeding for the Resistance toPhytophthora sojaein Korea

Phytophthora root and stem rot (PRR) of soybean is a severe disease that causes significant economic losses in soybean-growing countries. The causal agent, Phytophthora sojae, is a soil-borne oomycete that causes pre- and post-emergence damping-off of soybean in poorly drained soils. PRR has not been a severe problem in South Korea; however, as the cultivation of soybean in paddy fields recently increased, there is a growing interest because the incidence of PRR can be extensively expanded. R-gene mediated resistance is known as the main strategy for the management of this disease. On the other hand, partial resistance has also been known to provide more effective disease management than the Rps (Resistance to Phytophthora sojae) resistance. Responses of domestic soybean cultivars to a few P. sojae isolates were recently reported, while phenotypic distribution of numerous germplasms is mostly unknown. The objectives of this review are to summarize published studies conducted on PRR, to suggest future directions of genetic researches and breeding to the target pathogen. This review will discuss the followings; i) a brief introduction to PRR and the causal agent P. sojae, ii) types of genetic resistance to P. sojae and findings of genes/QTL conditioning to resistance, iii) changes in virulence pathotype of P. sojae populations, and iv) current status and perspective of genetic/genomic researches on the interaction of soybean and P. sojae in the South Korea.

Morphological Characterization and Pathogenicity of pythium sp. Infecting Cucumber Seedlings in Greenhouses

Greenhouse-grown cucumber seedlings exhibiting typical symptoms of Pythium damping-off and root rot were collected. The causal agents were isolate on Pythium selective medium; nystatin, ampicillin, rifampicin, and miconazole (NARM) in potato dextrose agar (PDA). Four different isolates was isolated and their pathogenicity was tested using cotyledon and young seedlings assays. Results of these isolates showed variation in their ability to infect cucumber seedling. Isolate GZ-11 was highly pathogenic, caused 100 % pre- and post-emergence damping-off. The isolate was identified to the species level based on morphological characters along with sequences of the internal transcribed spacer (ITS) of the ribosomal DNA and showed 99% similarity to Pythium spinosum (Synonym of Globisporangium spinosum). Aggressiveness of P. spinosum was investigated at two levels of inoculum concentrations (10 and 20 g/kg soil). Results reported that P. spinosum was highly aggressive and significantly reduced cucumber seed germination compared to the control.

Chemical management of damping-off (Pythium ultimum) disease of tomato

Damping-off (Pythium ultimum Trow) is an important disease of tomato in mid hills of Himachal Pradesh which is a major impediment in raising healthy seedlings. Keeping this in view, recently available and conventional fungicides were evaluated under in vitro and pot house conditions both as seed treatment and soil drenching for the management of this disease. In vitro evaluation revealed that Apron XL, Curzate M8, Metal, Ridomil Gold and Sectin were found most effective at all concentrations evaluated and caused complete mycelial inhibition and were statistically at par with each other. These fungicides were further tested as seed treatment and drench application against pre-emergence and post-emergence phase of the disease under pot conditions. Maximum seed germination and minimum pre-emergence damping-off were recorded in seeds treated with Metal, Ridomil Gold, Curzate M 8 followed by Sectin and Apron XL while, maximum reduction in post-emergence damping-off was observed in Apron XL, Curzate M 8, Metal, Ridomil Gold and Sectin.

Thermotherapy of Sunflower Seeds Controlling Charcoal-Rot Caused by Macrophomina phaseolina

Various fungi were isolated from sunflower seed samples collected from two fields in Ismailia and Beni-Suef governorates. Fifteen fungal species belonging to 10 genera were isolated from Beni-Suef fields and thirteen fungal species belonging to 8 genera were isolated from Ismailia field. Alternaria alternata recorded the highest frequency from sunflower seed samples collected from the two locations followed by Aspergillius flavus and A. niger. Macrophomina phaseolina was the main seed-borne pathogen isolated from sunflower seeds collected from the two locations followed by Fusarium oxysporum and Fusarium spp. Applying thermo-treatments through microwave radiation in five exposing periods (15, 30 sec, 1, 2 and 4 min.) and hot water treatment at five soaking periods (30 sec, 1, 2, 4 and 8 min.) gave significant impacts on the occurrence of seed-borne pathogen M. phaseolina in sunflower seeds as well as seed germination. Moreover, increasing the exposure time of microwave radiation or soaking in hot water led to enhancing their effects. Soaking in hot water negatively affected thefrequency of M. phaseolina than microwave radiation treatment. Generally, the obtained results clearly show that microwave radiations and hot water treatments significantly decreased the incidence of pre- and post-emergence damping-off. Moreover, sunflower seeds exposed to either treatment then planted under greenhouse conditions in infested and non-infested soil with M. phaseolina or under open field conditions, naturally infested, led to increase of sunflower seed yield. In this respect, the highest impact was obtained when sunflower seeds were soaked in hot water at 2 min followed by soaking in hotwater for 1 min and exposure to microwave radiation for 1 min. Data also proved that the efficiency of some thermal treatments was approximately close to efficiency of fungicide in reducing damping-off and charcoal rot diseases during 2017 and 2018 trail seasons, where soaking in hot water for 2 min. was the closest one.

Molecular characterization of genomic regions for resistance to Pythium ultimum var. ultimum in the soybean cultivar Magellan.

Two novel QTL for resistance to Pythium ultimum var. ultimum were identified in soybean using an Illumina SNP Chip and whole genome re-sequencing. Pythium ultimum var. ultimum is one of numerous Pythium spp. that causes severe pre- and post-emergence damping-off of seedlings and root rot of soybean [Glycine max (L.) Merr.]. The objective of this research was to identify quantitative trait loci (QTL) for resistance to P. ultimum var. ultimum in a recombinant inbred line population derived from a cross of 'Magellan' (moderately resistant) and PI 438489B (susceptible). Two different mapping approaches were utilized: the universal soybean linkage panel (USLP 1.0) and the bin map constructed from whole genome re-sequencing (WGRS) technology. Two genomic regions associated with variation in three disease-related parameters were detected using both approaches, with the bin map providing higher resolution. Using WGRS, the first QTL were mapped within a 350-kbp region on Chr. 6 and explained 7.5-13.5% of the phenotypic variance. The second QTL were positioned in a 260-kbp confidence interval on Chr. 8 and explained 6.3-16.8% of the phenotypic variation. Candidate genes potentially associated with disease resistance were proposed. High-resolution genetic linkage maps with a number of significant SNP markers could benefit marker-assisted breeding and dissection of the molecular mechanisms underlying soybean resistance to Pythium damping-off in 'Magellan.' Additionally, the outputs of this study may encourage more screening of diverse soybean germplasm and utilization of genome-wide association studies to understand the genetic basis of quantitative disease resistance.

Identifikasi dan Kuantifikasi Metabolit Bakteri Pelarut Fosfat dan Pengaruhnya terhadap Aktivitas Rhizoctonia solani pada Tanaman Kedelai

Phosphate solubilizing bacteria (PSB) metabolites are organic acids, phosphomonoesterase enzyme (alkaline phosphatase) and antibiotic, which is able to dissolve insoluble phosphate. Phosphate solubilizing bacteria used in this study was expected to suppress Rhizoctonia solani attacks. This experiment was aimed at (1) identifiying and quantifying PSB metabolites, and (2) examining their capability as biocontrol agent for Rhizoctonia solani in vitro and hydroponics soybean. This study was conducted in three stages. The first stage of this study was culturing two PSB isolates (Pseudomonas putida 27.4B and Pseudomonas diminuta) in the Pikovskaya medium to analyze their metabolites. The second and third stage of this study was testing the antagonist of two bacteria to suppressed R. solani activity, which was conducted in vitro, and in hydroponics medium soybean as indicator plant. The results showed that P. putida 27.4B and P. diminuta produced organic acids i.e.: citrate, formic, succinic, acetic, propionate, butyrate, and oxalate. The totals of organic acids from each bacterium were 70,3 mg.kg-1 and 61,9 mg.kg-1. Production of alkaline phosphatase enzyme in Pikovskaya medium of P. Putida27.4B was 11,71 μg pNP .mL-1.h-1 and P. diminuta was 24,04 μg pNP.mL-1.h-1. Concentration of this enzyme in soil medium was higher than that in Pikovskaya medium with 26,27 μg pNP.g-1.h-1 and 39,03 μg pNP.g-1.h-1 respectively. This study also showed that total concentration of antibiotics (tetracycline, oxitetracycline and penicillin) produced by the PSB, were 3,2 μg.mL-1 (P. putida 27.4B) and 10,96 μg.m1-1 (P. diminuta), respectively. The results from second stage of this study showed that by using in vitro, the reduced growth of R. solani was observed 58,35% with P. putida 27.4B and 41,96% with P. diminuta. In addition, inoculations of PSB in hydroponics medium reduced the fungal pathogenesis from 10,71% to 21,42% of pre and post emergence damping-off. Visually, the symptom of pathogen attack appeared within the period of 2 untill 14 days after infection.

Evaluation of fungicides, biocontrol agents and botanicals for management of damping-off in cabbage seedlings caused by Fursarium moniliforme sheld

The antagonistic effect of 10 fungicides, four biocontrol agents and 10 locally available botanicals in controlling the growth and sporulation of Fusarium moniliforme in vitro were tested. A pot culture experiment was also conducted to study the effect of seed treatment and soil treatment alone or in combination on germination and seedling vigour of cabbage. In vitro evaluation of ten fungicides by poisoned food technique showed that all the fungicides efficiently inhibited complete inhibition of the linear growth of the fungus was observed with Thiophenate methyl, Carbendazim, Mancozeb, Difenconazole, Propiconazole, Hexaconazole, Sixer and Tricyclazole. Sporulation was completely inhibited by all the fungicides evaluated. In dual culture technique of the fungus and biocontrol agents, of the four biocontrol agents evaluated the antagonists, T. hamatum completely overgrew the pathogen, T. harzianum colonized on half of the growth of the pathogen, T. viride remained locked with the fungus at the point of contact and P. citrinum formed inhibition zone with the fungus. Sporulation was completely inhibited by all the four antagonists. Among the ten locally available botanicals tested by poisoned food technique in vitro, garlic extract showed maximum fungitoxic propertyfollowed wild sage (Lantana camara), onion (Allium cepa), neem (Azadirachta indica), goat weed (Ageratum conyzoides) and marigold (Tagetes erecta) . In pot culture experiment, the least disease incidence was recorded in soil treatment with carbendazim (26%) followed by T. harzianum (29%) and P. glabrum (29.7%). However,considering the various drawbacks and hazardous effects resulting from use of chemicals, biocontrol agents and plant extracts can be recommended to use in raising healthy seedlings free from pre- and post-emergence damping-off.

Antifungal activity of some medicinal plants on Soybean seed-borne Macrophomina phaseolina

Soybean Glycine max (L.) Merr. seeds, pods and seedlings are susceptible to fungal attack due to its rich nutrient content. The most commonly isolated soybean (JS-335) seed-borne fungi were Fusarium spp., Macrophomina phaseolina., Pythium spp., Aspergillus spp., Phoma spp., and Phomopsis spp. Macrophomina phaseolina is an important plant pathogenic fungus that causes charcoal rot of soybean and infects more than 500 hosts. In humid climates, the fungus causes a post emergence damping-off of soybean seedlings leading to 50% of crop losses. The objective of this work was to study the efficacy of the botanicals on soybean seed-borne Macrophomina phaseolina. Among the 10 botanicals screened, Datura metel (L.) methanol leaf extract showed the most promising activity against Macrophomina phaseolina.

First Report of Root Rot Caused by Rhizoctonia solani AG-10 on Canola in Washington State.

Canola (Brassica napus L) production has gained renewed interest in Washington State over the past few years, primarily for the purpose of producing biofuel. Plants were observed to be showing symptoms of Rhizoctonia root rot and postemergence damping-off. In many cases, this was due to Rhizoctonia solani AG-2-1, which was previously documented (4). However, additional plants were occasionally observed that were stunted and had reduced vigor, but lacked the distinctive severe stem damage and postemergence damping-off, which are both symptoms of infection with R. solani AG-2-1. Isolates of R. solani AG-10 were collected from symptomatic plants or baited from root zone soil at various dryland production locations in eastern Washington, including sites near Colfax, Pullman, and Walla Walla. Initial identification was determined by quantitative (Q)-PCR using R. solani AG-10 specific primers (3). The identity was verified by sequencing random isolates identified by Q-PCR (GenBank Accessions Nos. JQ068147, JQ068148 and JQ068149). All sequenced isolates had 99% identity to previously reported isolates of R. solani AG-10. Six isolates were chosen to test pathogenicity on canola plants in the greenhouse. Sterilized oats were inoculated with each of six isolates of R. solani AG-10 and grown for 4 weeks. The soil was infested with ground oat inoculum (1% wt/wt) and spring canola cv. Sunrise was seeded into 3.8 × 21-cm containers. After 3 weeks of incubation at 15°C, plants were harvested and assessed. Emergence was reduced in the infested soil with 73 to 93% (average 81%) emergence compared with 100% emergence in the noninfested soil. There was no evidence of postemergence damping-off. However, all six isolates of R. solani AG-10 significantly reduced the plant height and top dry weights compared with the noninfested controls. The plant height in infested soil was 28 to 42% (average 34%) shorter and top dry weights were 37 to 70% (average 54%) lower than in noninfested soil. Roots of infected plants had a light brown discoloration along with reduced length and fewer lateral roots. Additional host plants were tested, including wheat (Triticum aestivum L.), barley (Hordeum vulgare L.), pea (Pisum sativum L.), chickpea (Cicer arietinum L.), and lentil (Lens culinaris Medik.). There was no significant reduction in plant height or plant dry weight for any of these hosts. R. solani AG-10 was previously found to be weakly virulent on canola and other cruciferous hosts in Australia (1,2). To our knowledge, this is the first report of R. solani AG-10 causing disease on canola in Washington State. Reference: (1) R. K. Khangura et al. Plant Dis. 83:714, 1999. (2) G. C. MacNish et al. Australas. Plant Pathol. 24:252, 1995. (3) P. A. Okubara et al. Phytopathology 98:837, 2008. (4) T. C. Paulitz et al. Plant Dis. 90:829, 2006.

Evaluation of Microbial, Botanical, and Organic Treatments for Control of Peanut Seedling Diseases.

Diseases affecting stand establishment are a major obstacle to organic production of peanut (Arachis hypogaea). Stand losses of 50% or more are possible with untreated seed. Biological, botanical, and organic seed treatments or soil amendments were tested for efficacy against pre- and postemergence damping-off of peanut in greenhouse, microplot, and field plot trials. Seed of the lines Perry, GP-NC 343, and Bailey (tested as N03081T) were used in all trials. Commercial formulations of Bacillus subtilis (Kodiak), B. pumilus (Yield Shield), Trichoderma harzianum (T-22 PB and Plantshield HC), Muscodor albus, and Coniothyrium minitans (Contans); activated charcoal; two separate soil amendments of dried herbage of Monarda didyma cultivars; a commercial fungicide control (Vitavax PC); and an untreated control were tested in natural soil in the greenhouse. Vitavax PC and Kodiak were the only treatments that resulted in higher percent emergence and survival than in untreated seed. A separate greenhouse experiment was conducted in natural soil or natural soil infested with field isolates of Aspergillus niger. Seed were treated with Kodiak, copper hydroxide (Champion), Plantshield HC, Kodiak + Plantshield HC, Streptomyces griseoviridis (Mycostop), hot water, Vitavax PC, or were left untreated. Seedling emergence and survival was much lower in infested versus uninfested soil. Seed treatment with Kodiak increased percent emergence and survival compared to untreated seed, but was not as effective as Vitavax PC. Field microplot studies in 2007 and 2008 at Clayton, NC, evaluated four seed treatments on the peanut lines following small grain cover crops, soil amendment with M. albus, or no cover. Cover crops did not affect emergence or interact with seed treatments. In field studies in 2007 and 2008 at Lewiston, NC, the peanut lines were planted with M. albus infurrow, with Kodiak or T. harzianum seed treatments, or were untreated. In the 2007 trial, none of the treatments improved stands compared to the untreated check. In 2008, the highest stand counts were produced by seed treated with Kodiak. In both years, Bailey produced the greatest stand counts. A. niger was strongly associated with postemergence damping-off in the field. Regardless of peanut line, in many trials, Kodiak seed treatment increased emergence and survival over untreated seed.

Field performance of bio-primed seeds to suppress Colletotrichum truncatum causing damping-off and seedling stand of soybean

The soybean is prone to be attacked by Colletotrichum truncatum at seed and seedling stages, resulting in pre- and post-emergence damping-off. The efficacy of bio-priming for the control of damping-off of soybean caused by C. truncatum and the final seedling stand using two fungal biocontrol agents (BCAs) Trichoderma harzianum and T. virens and one bacterial BCA Pseudomonas aeruginosa was evaluated under field conditions. Treatments consisted of chemo-primed with Benlate® as a positive control; bio-primed with P. aeruginosa; bio-primed with T. harzianum; bio-primed with T. virens; bio-primed with the mixture of T. virens and T. harzianum and the controls as hydro-primed and non-primed seeds. Trichoderma isolates used singly or as a mixture established on the seed surface with germinating hyphae, whereas the strain P. aeruginosa colonized profusely as determined by increased colony forming units (CFU) from 1.2 × 10 9 to 5.1 × 10 9 seed −1 after 12 h of bio-priming. All bio-priming treatments significantly reduced pre- and post-emergence damping-off relative to hydro- and non-primed seed controls. In general, bio-priming with P. aeruginosa was the most effective treatment for controlling pre and post-emergence damping-off, with reductions in disease incidence with increases ranging from 48.6% to 51.9% and 65.0% to 97.2%, respectively. Moreover, P. aeruginosa resulted in enhancement of seed germination and healthy seedling stand ranging from 32.4% to 60.0% and 56.0% to 73.9%, respectively. Bio-priming with T. harzianum reduced pre- and post-emergence damping-off by 42.8–46.8% and 35.0–85.1%, respectively. However, P. aeruginosa was generally comparable to T. harzianum and the fungicide Benlate®. The combination treatment of T. harzianum and T. virens produced compareable results to T. harzianum alone, and T. virens was the least effective of the bio-primed treatments. Bio-priming with P. aeruginosa or T. harzianum offered an effective biological seed treatment system and an alternative to the fungicide Benlate® for control of damping-off of soybean caused by C. truncatum of soybean.

Tratamento de sementes de algodoeiro com fungicidas no controle de patógenos causadores de tombamento de plântulas

Dentre as doenças que incidem sobre o algodoeiro, o "tombamento" é considerado uma das principais, sendo causado por um complexo de fungos de solo e da semente, os quais, ocorrendo separadamente ou em combinação, podem ocasionar o tombamento de pré e pós-emergência das plântulas. Os principais agentes etiológicos do tombamento são Rhizoctonia solani Khun, Colletotrichum gossypii South e Colletotrichum gossypii South var. cephalosporioides Costa. O objetivo deste trabalho foi avaliar o efeito de diversos fungicidas, utilizados em tratamento de sementes de algodão, no controle de patógenos associados às sementes e/ou presentes no solo. Sementes livres de patógenos, não-inoculadas e inoculadas com Colletotrichum gossypii var. cephalosporioides(Cgc)), foram tratadas com tolylfluanid + pencycuron + triadimenol, carboxin + thiram e fluazinam + tiofanato metílico. No teste de emergência em areia, o tratamento testemunha inoculada com Cgc apresentou o menor índice de emergência. O maior índice de emergência ocorreu em sementes sem inoculação e tratadas com tolylfluanid + pencycuron + triadimenol. O tratamento mais eficiente no controle do tombamento de pós-emergência do algodoeiro, em substratos contendo Rhizoctonia solani, foi a mistura tolylfluanid + pencycuron + triadimenol. No campo, a maior incidência e severidade da ramulose, causada por Cgc, ocorreu em plantas provenientes de sementes não tratadas com fungicidas e inoculadas com Cgc. Nenhum dos fungicidas testados foi fitotóxico ao algodão.

Efeito do tratamento de sementes de algodoeiro com fungicidas no controle do tombamento de plântulas causado por Rhizoctonia solani, sob condições de casa de vegetação

O objetivo deste trabalho foi avaliar a eficacia do tratamento de sementes de algodoeiro com fungicidas no controle do tombamento de plântulas causado por Rhizoctonia solani. O ensaio foi conduzido na casa de vegetacao da Embrapa Agropecuaria Oeste, em Dourados, Mato Grosso do Sul. Sementes nao tratadas e tratadas com os fungicidas foram semeadas em areia contida em bandejas plasticas, dispostas em orificios individuais, equidistantes e a 3 cm de profundidade. A inoculacao com R. solani AG-4 foi feita utilizando-se 5g do inoculo do fungo/bandeja, distribuidos de forma homogenea na superficie do substrato. O fungo foi cultivado por 35 dias em sementes de aveia preta autoclavadas e trituradas em moinho (1mm). Foi observado efeito significativo do tratamento fungicida na emergencia inicial e final de plântulas, bem como no controle do tombamento de pre e pos-emergencia do algodoeiro, com os melhores resultados sendo obtidos pelos tratamentos tolylfluanid+pencycuron+triadimenol e azoxystrobin+fludioxonil+mefenoxan, seguidos de carboxin+thiram, PCNB e pencycuron. O fungicida menos eficiente foi o carbendazim+thiram. Melhores resultados com relacao as variaveis estudadas foram obtidos com a utilizacao de misturas de fungicidas em comparacao ao uso isolado de um determinado produto. Nao foram observadas diferencas significativas com relacao ao indice de velocidade de emergencia, altura e peso fresco das plântulas.