Development of a Recombinase Polymerase Amplification Assay for Rapid Detection of the New Root-lesion Nematode Pratylenchus dakotaensis on Soybean.

Root-lesion nematodes, Pratylenchus spp. are reported to cause serious yield losses in various crops including soybean. A new root-lesion nematode species was recently detected in a soybean field in North Dakota (ND) and named Pratylenchus dakotaensis. Nematode detection and differentiation from other species are critical in management strategies. Thus, a recombinase polymerase amplification (RPA) assay was developed for rapid detection of this nematode from field soils under isothermal conditions. New primers and probes were designed from ITS-rDNA region of the nematode genome and tested for both specificity and sensitivity. The RPA assay was able to detect DNA from a single adult nematode at 39.5°C in 20 minutes using both Basic and Exo kits. The specificity of the primers was initially confirmed through in silico analyses, followed by laboratory tests. The assay successfully amplified DNA from the target species, while no amplification occurred for other Pratylenchus spp. and non-Pratylenchus control species. Sensitivity testing with real-time RPA revealed its ability to detect DNA in dilutions equivalent to 1/32 of a single nematode from DNA extracted from inoculated sterile soil. To further validate the assay, it was tested with 19 field soil samples collected in ND. This assay amplified soil DNA extracts of all P. dakotaensis-infested field samples confirmed through conventional PCR. It did not amplify DNA from 13 other field soils infested with other Pratylenchus spp. This is the first report of RPA develoment for detecting a root-lesion nematode species. The RPA assay developed can help in the rapid detection of this nematode species for effective nematode management.

Simultaneous determination of cucurbitacins B, D, and E in Cucurbita seed oils by HPLC-DAD.

Cucurbitacins are triterpene bioactive constituents of natural products, particularly in the Cucurbitaceae plant family. The presence of cucurbitacins in seeds of the Cucurbita genus (pumpkin) has been only little studied. In this work, the content of cucurbitacins B, D, and E in seed oils from three cucurbits (Cucurbita moschata Duch, Cucurbita pepo Linn, and Cucurbita maxima Linn) was studied. An analytical method based on HPLC-DAD for the detection and quantification of these three cucurbitacins in seed oils was developed and validated according to ICH guidelines. The method showed good linearity, accuracy, and precision for the simultaneous quantification of cucurbitacins B, D, and E using C.moschata seed oil as a reference. When applied to C.pepo and C.maxima seed oils, cucurbitacin B and D were quantified but to a lesser extent. This is the first report of a simple, repeatable, and reproducible analytical tool to identify cucurbitacins in oilseeds from Cucurbita spp.

Inter- and Intra-Pathogen Interactions Emanating from Coinfection with Different Fungal and Viral strains in Canola Cultivars with Differing Host Resistances.

Few recent investigations examine coinfection interactions between fungal and viral plant pathogens. Here, we investigated coinfections between Leptosphaeria maculans and turnip mosaic virus (TuMV) in canola (Brassica napus). Different combinations of L. maculans isolate P11 and resistance breaking isolates L. maculans UWA192 and TuMV 12.1, were inoculated to three cultivars with differing pathogen resistances/susceptibilities. They were inoculated first to entire or half cotyledons 10-12 days after emergence, and second to opposite entire or half cotyledons on the same day (day 0) or 3 or 7 days afterwards. The parameters measured were L. maculans cotyledon disease index (%CDI), and TuMV systemically infected leaf symptom intensity (SI) and virus concentration (VC). Except when both day 0 inoculations were with isolate UWA192, %CDI values were supressed strongly or only weakly when isolates P11 and/or UWA192 were inoculated to plants with L. maculans single gene resistance (SGR) or polygenic resistance, respectively. However, except when isolate P11 was inoculated first and UWA192 second, these values declined after inoculation day 0 when SGR was absent. TuMV infection suppressed %CDI values, although this decrease was usually smaller following day 0 half cotyledon inoculations. When TuMV temperature sensitive extreme resistance was present and both inoculations were with TuMV, SI and VC values diminished greatly. However, the extent of this decrease was reduced when second inoculations were with L. maculans. SI and VC values were also smaller when SGR was present and second inoculations were with L. maculans. When L. maculans resistance was lacking, SI and VC values were smaller when second inoculations to entire cotyledons were with L. maculans rather than TuMV. This also occurred after second half cotyledon inoculations with isolate P11 but not isolate UWA192. Therefore, diverse inter- or intra-pathogen interactions developed depending upon host resistance, isolate combination, cotyledon inoculation approach and second inoculation timing.

Genome-wide association study of bacterial blight resistance in soybean.

Bacterial blight caused by Pseudomonas syringae pv. glycines (Psg) is a widespread foliar disease. Although four Resistance to Pseudomonas syringae pv. glycinea (Rpg) 1 ~ 4 (Rpg1~4) genes that have been observed to segregate in a Mendelian pattern have been reported to confer resistance to Psg in soybean, the genetic basis of quantitative resistance to bacterial blight in soybean remains unclear. In the present study, the Psg resistance of two soybean association panels consisting of 573 and 213 lines, respectively, were phenotyped in multiple environments in 2014 - 2016. Genome-wide association study (GWAS) were performed using 2 models FarmCPU and BLINK to identify Psg resistance loci. A total of 40 soybean varieties with high level of Psg resistance were identified, and 14 quantitative trait loci (QTLs) were detected on 12 soybean chromosomes. These QTLs were identified for the first time. The majority of the QTLs were only detected in one or the other association panels, while qRPG-18-1 was detected in both association panels for at least one growing season. A total of 46 candidate Psg resistance genes were identified from the qRpg_13_1, qRPG-15-1, and qRPG-18-1 loci based on gene function annotation. In addition, we found the genomic region covering rpg1-b and rpg1-r harbored the synteny with a genomic region on chromosome 15, and identified 16 nucleotide binding site - leucine-rich repeat (NBS-LRR) genes as the candidate Psg resistance genes from the synteny blocks. This study provides new information for dissecting the genetic control of Psg resistance in soybean.

Mixing Elemental Sulfur or Chlorothalonil with QoI and DMI Fungicides for Management of Late Leaf Spot of Peanut.

Reduced sensitivity to demethylation inhibitor (DMI) and quinone outside inhibitor (QoI) fungicides in Nothopassalora personata, the cause of late leaf spot of peanut (Arachis hypogaea) complicates management of this disease in the southeastern U.S. Mixtures with protectant fungicides may help preserve the utility of members of both DMI and QoI fungicide groups for leaf spot management. Field experiments were conducted in Tifton, GA from 2019 to 2021 and in Plains, GA during 2019 and 2020. The primary objective was to determine the effects of mixtures of DMI fungicides, tebuconazole and mefentrifluconazole, and QoI fungicides, azoxystrobin and pyraclostrobin, with micronized elemental sulfur on late leaf spot in fields with populations of N. personata with suspected reduced sensitivity to DMI and QoI fungicides. In four of the experiments, the efficacies of elemental sulfur and chlorothalonil as mixing partners were also compared. In most cases, standardized area under the disease progress curve (sAUDPC) and final percent defoliation were less for all DMI and QoI fungicides mixed with sulfur or chlorothalonil than for the respective fungicides alone. In most cases, sAUDPC and final percent defoliation were similar for sulfur and chlorothalonil when mixed with the respective DMI or QoI fungicide. These results indicate that mixtures of DMI or QoI fungicides with either micronized sulfur or chlorothalonil can improve control of late leaf spot compared to the DMI or QoI fungicide alone. These results also indicate that elemental sulfur has potential as an alternative to chlorothalonil in tank mixes where that protectant fungicide is currently being used as a mixing partner to improve leaf spot control.

First Report of Xanthomonas euvesicatoria pv. sesami Causing Leaf Spot on Sesame (Sesamum indicum L.) in South Carolina, USA.

Sesame (Sesamum indicum L.) is an annual plant known as one of the first domesticated oilseed crops. It is cultivated worldwide, mostly in Asia, Africa, and the Americas (Singh, 2006). In August 2022 and September 2023, dark angular necrotic spots on leaves and stems (100% incidence), blights, and severe defoliation were observed in a 4-acre rainfed sesame field located in the Colleton County of South Carolina, USA (Fig. S1). Bacterial streaming from cut leaf lesions was observed from diseased plants in both years. Two plants were collected for pathogen isolation in 2023. Symptomatic leaves were surface sterilized with 70% ethanol for 1 min and dried in a laminar flow hood. For each isolate, four sterile toothpicks were used to poke lesion margins and stirred in 300 µl of sterile distilled water in a 2-ml sterile microcentrifuge tube and soaked at room temperature (c. 21 °C) for 10 min. Each bacterial suspension (10 µl) was streaked on nutrient agar (NA) in a Petri dish. Convex and mucoid yellow colonies formed after a 48-h incubation at 28°C in the dark. Two isolates (S813 and S814), one from each plant, were obtained by transferring single colonies to new NA plates. Both isolates were preliminarily identified as Xanthomonas [S813: X. campestris (P = 0.53); S814: X. campestris (P = 0.77)] using a Biolog Microbial Identification System (GEN III Microplate; Identification Database v.2.8.0.15G). PCR amplification of the atpD and dnaK genes was performed for both isolates using the conditions described in Félix-Gastélum et al. (2019). The sequences of both amplicons are 100% identical for each gene between the two isolates. PCR and sequencing of the gyrB gene was also done for S813 with the primers from Young et al. (2008). The atpD (S813/S814), dnaK (S813/S814), and gyrB (S813) sequences (GenBank accessions: PP507118 to PP507120) showed the best match with 100% identity to the corresponding gene sequences [GenBank accessions: KJ491167 (100% coverage), KJ491257 (99% coverage), EU285201 (100% coverage)] of the X. euvesicatoria pv. sesami (=X. campestris pv. sesami) type strain LMG865 (Constantin et al. 2015, Parkinson et al. 2009). A neighbor joining tree with the concatenated sequences of these three genes (2,210 nt) showed that S813 and LMG 865 had the closet relationship with X. euvesicatoria pv. alfalfae (CFBP3836, Fig. S2). To fulfill Koch's postulates, three healthy sesame plants (cultivar Shirogoma) were spray inoculated separately with each suspension of S813 and S814 in sterile tap water until runoff (approx. 5×108 CFU/ml). Two sesame plants were sprayed with sterile tap water and served as negative control. All plants were maintained in a greenhouse at approximately 28/20°C (day/night) with natural photoperiod. Dark leaf spots and leaf yellowing were observed on inoculated plants 7 to 14 days after inoculation. No disease symptom was observed on the control plants. Bacteria were reisolated from leaf spots of the inoculated plants and confirmed to be X. euvesicatoria pv. sesami based on atpD and dnaK sequences. The disease was first reported in Sudan (Sabet and Dowson, 1960), after which it was reported in USA (Isakeit et al., 2012) and Mexico (Félix-Gastélum et al. 2019). To the best of our knowledge, this is the first report of this disease in South Carolina, USA. Since the interest of sesame to the farmers is increasing in the southeastern USA, it is necessary to perform further research to examine the disease distribution and its economic impact.

Chia (Salvia hispanica L.), a functional 'superfood': new insights into its botanical, genetic and nutraceutical characteristics.

BACKGROUND: Chia (Salvia hispanica L.) seeds have become increasingly popular among health-conscious consumers due to their high content of ω-3 fatty acids, which provide various health benefits. Comprehensive chemical analyses of chia seeds' fatty acids and proteins have been conducted, revealing their functional properties. Recent studies have confirmed the high ω-3 content of chia seed oil and have hinted at additional functional characteristics. SCOPE: This review article aims to provide an overview of the botanical, morphological, and biochemical features of chia plants, seeds, and seed mucilage. Additionally, we discuss the recent developments in genetic and molecular research on chia, including the latest transcriptomic and functional studies that examine the genes responsible for chia fatty acid biosynthesis. In recent years, research on chia seeds has shifted its focus from studying the physicochemical characteristics and chemical composition of seeds to understanding the metabolic pathways and molecular mechanisms that contribute to their nutritional benefits. This has led to a growing interest in various pharmaceutical, nutraceutical, and agricultural applications of chia. In this context, we discuss the latest research on chia, as well as the questions that remain unanswered, and identify areas that require further exploration. CONCLUSIONS: Nutraceutical compounds associated with significant health benefits including ω-3 PUFAs, proteins, and phenolic compounds with antioxidant activity have been measured in high quantities in chia seeds. However, comprehensive investigations through both in vitro experiments and in vivo animal and controlled human trials are expected to provide greater clarity on the medicinal, antimicrobial, and antifungal effects of chia seeds. The recently published genome of chia and gene editing technologies, such as CRISPR, facilitate functional studies deciphering molecular mechanisms of biosynthesis and metabolic pathways in this crop. This necessitates development of stable transformation protocols and creation of a publicly available lipid database, mutant collection, and large-scale transcriptomic datasets for chia.

Evaluation of Compost and Manure Amendments for Suppressing Heterodera glycines Ichinohe, 1952.

Soybean cyst nematode is a major pest of soybean crops, causing significant yield losses and economic impact. Current management strategies primarily rely on resistant varieties, cover crops, and seed treatments. However, there is a growing interest in developing sustainable, ecologically based approaches to integrate SCN risk reduction into soybean production systems. This study aimed to evaluate the efficacy of various compost and manure amendments in suppressing SCN populations and promoting soybean productivity. An in vitro egg hatching assay was conducted to screen the inhibitory effects of different compost and manure extracts on SCN egg hatching. Results indicated that poultry manure, Layer Ash Blend®, and swine manure extracts significantly inhibited SCN hatching compared to other treatments across multiple time points. Greenhouse trials further validated the effectiveness of Layer Manure®, poultry manure, High Carbon Dairy Doo®, and Seed Starter 101® in suppressing SCN cysts, eggs, and juveniles. A field microplot trial confirmed the practical promise of Layer Ash Blend® and poultry manure in SCN management, with significant reductions in SCN populations and increased soybean yields. The study also investigated the impact of these amendments on promoting the population of bacterivorous and frugivorous nematodes, contributing to a biological diverse soil ecosystem. Overall, the results indicate that amending SCN-infested soil with specific compost or manure formulations can effectively suppress nematode populations while improving soybean productivity. These findings contribute to the development of sustainable strategies for SCN management in soybean production systems.

Survey of Heterodera glycines population abundance and virulence phenotypes during 2021-2022 in Heilongjiang Province.

Soybean cyst nematode (SCN), Heterodera glycines, poses a significant threat to global soybean production. Heilongjiang, the largest soybean-producing province in China, contributes over 40% to the country's total yield. This province has much longer history of SCN infestation. To assess the current situation in Heilongjiang, we conducted a survey to determine the SCN population density and virulence phenotypes during 2021-2022 and compared the data with a previous study in 2015. A total of 377 soil samples from 48 counties representing eleven major soybean-planting regions were collected. The prevalence of SCN increased from 55.4% in 2015 to 59% in the current survey. The population densities ranged from 80 to 26,700 eggs and juveniles per 100 cm3 of soil. Virulence phenotypes were evaluated for 60 representative SCN populations using the HG type test, revealing nine different HG types. The most common virulence phenotypes were HG types 7 and 0, accounting for 56.7% and 20% of all SCN populations, respectively. The prevalence of populations with a reproductive index (FI) greater than 10% on PI548316 increased from 64.5% in 2015 to 71.7%. However, the FI on the commonly used resistance sources PI 548402 (Peking) and PI 437654 remained low at 3.3%. These findings highlight the increasing prevalence and changing virulence phenotypes of SCN in Heilongjiang. They also emphasize the importance of rotating soybean varieties with different resistance sources and urgently identifying new sources of resistance to combat SCN.

First Report of Wilt Caused by Fusarium nanum (FIESC 25) on Mungbean (Vigna radiata) in Pakistan.

Mungbean, Vigna radia (L.) R. Wilczek, is ranked 2nd next to chickpea (Cicer arietinum) in total cultivation and production in Pakistan. In August of 2022 and 2023, mungbean plants (cv. PRI Mung-2018) were found wilting in a field at the Ayub Agricultural Research Institute, Faisalabad, Pakistan. Wilted leaves turned yellow, died, but remained attached to the stem. Vascular tissue at the base of the stem showed light to dark brown discoloration. Roots were stunted with purplish brown to black discoloration. Symptomatic mungbean plants were collected from fields at five different locations (20 samples/location). Disease incidence was similar among the five fields, ranging from 5 to 10% at each location depending upon type of germplasm and date of sowing. For fungal isolation and morphological identification, symptomatic stem and root tissues were cut into ~5 mm2 pieces with a sterilized blade. Tissues were surface-sterilized for one min in a 0.5% sodium hypochlorite solution, rinsed twice in sterilized water, air dried on sterilized filter paper, and aseptically placed on potato dextrose agar (PDA) containing 0.5 g/L-1 streptomycin sulphate. Plates were incubated for 3-4 days at 25 ± 2°C with a 12-h photoperiod. Single-spore cultures were used for morphological and molecular analyses. Isolates on PDA grew rapidly and produced abundant white aerial mycelium that turned off-white to beige with age. Macroconidia were hyaline, falcate, typically 3-to-6 septate with a pointed apical cell and a foot-shaped basal cell, measuring 24.5-49.5 x 2.7-4.7 µm (n = 40). Globose to obovate chlamydospores measuring 5.8 ± 0.5 µm (n = 40) were produced singly or in chains and were intercalary or terminal and possessed roughened walls. The morphological data indicated the isolates were members of the genus Fusarium (Leslie and Summerell 2006). To obtain a species-level identification, a portion of translation elongation factor 1-α (TEF1), the largest subunit of RNA polymerase (RPB1), and the second largest subunit of RNA polymerase (RPB2) region were PCR amplified and sequenced using EF1/EF2 (O'Donnell et al. 1998), Fa/G2R (Hofstetter et al. 2007), and 5f2/7cr (Liu et al. 1999) primers, respectively. DNA sequences of these genes were deposited in GenBank under accession numbers MW059021, MW059017 and MW059019, respectively. The partial TEF1, RPB1 and RPB2 sequences were queried against the Fusarium MLST database (https://fusarium.mycobank.org/page/Fusarium_identification), using the polyphasic identification tool. The BLASTn search revealed 99.9% identity of the isolate to F. nanum (Xia et al. 2019), formerly FIESC 25 of the F. incarnatum-equiseti species complex (MRC 2610, NRRL 54143; O'Donnell et al. 2018). To confirm pathogenicity, roots of 3-5 leaf stage mungbean seedlings were soaked in a 106 spores ml-1 conidial suspension of the fungus for 15 min and then planted in 10 cm pots containing sterilized soil. Mock-inoculated plants with sterile water served as a negative control. Twenty pots that were used for each inoculated and control treatment were maintained at 25 ± 2°C, 14:8 h photoperiod, and 80% relative humidity in a growth chamber. After 15 days, leaf yellowing, internal browning from the base of stems and root discoloration was observed in all the inoculated plants. The uninoculated negative control plants remained asymptomatic. Fusarium nanum was re-isolated from artificially inoculated plants and identified by colony growth, conidial characteristics on PDA and molecular analyses (TEF1). To our knowledge, this is the first report of wilt caused by F.nanum on mungbean in Pakistan. In Pakistan, mungbean cultivation in irrigated areas has increased in recent years. It has been introduced frequently in citrus orchards, crop rotation of maize and sesame, intercropping with sugarcane and as green manure. However, citrus, maize, sesame and sugarcane are also hosts of Fusarium spp. Therefore, this information warrants sustainable crop protection and may have an impact on further interaction of F. nanum with other wilt pathogens.

No Common Candidate Genes for Resistance to Fusarium graminearum, F. proliferatum, F. sporotrichioides, and F. subglutinans in Soybean Accessions from Maturity Groups 0 and I: Findings from Genome-wide Association Mapping.

Seedling diseases and root rot, caused by species of Fusarium, can limit soybean (Glycine max L.) production in the United States. Currently, there are few commercially available cultivars resistant to Fusarium. This study was conducted to assess the resistance of soybean maturity group (MG) accessions from 0 and I to Fusarium proliferatum, F. sporotrichioides, and F. subglutinans, as well as to identify common quantitative trait loci (QTLs) for resistance to these pathogens, in addition to F. graminearum, using a genome-wide association study (GWAS). A total of 155, 91, and 48 accessions from the United States Department of Agriculture (USDA) soybean germplasm collection from MG 0 and I were screened with a single isolate each of F. proliferatum, F. sporotrichioides, and F. subglutinans, respectively, using the inoculum layer inoculation method in the greenhouse. The disease severity was assessed 21 days postinoculation and analyzed using nonparametric statistics to determine the relative treatment effects (RTEs). Eleven and seven accessions showed significantly lower RTEs when inoculated with F. proliferatum and F. subglutinans, respectively, compared with the susceptible cultivar 'Williams 82'. One accession was significantly less susceptible to both F. proliferatum and F. subglutinans. The GWAS conducted with 41,985 single-nucleotide markers identified one QTL associated with resistance to both F. proliferatum and F. sporotrichioides, as well as another QTL for resistance to both F. subglutinans and F. graminearum. However, no common QTLs were identified for the four pathogens. The USDA accessions and QTLs identified in this study can be utilized to selectively breed resistance to multiple species of Fusarium.[Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

First report of Colletotrichum incanum causing leaf spots on common bean in Europe (Slovenia).

A Phaseolus vulgaris L. leaf showing necrotic spots was collected in an experimental bean field in central Slovenia in August 2021. The field contained diverse common bean lines sourced from genebank collections, with each line represented by 10 plants. While symptomatic leaves were seen across various lines, the reported species derived exclusively from a Huasca Huallaga Colorado plant (single-seed descent, USDA accession PI153714, doi: 10.18730/H7P9N), a Peruvian landrace. After incubating the leaf for 2 d at ambient temperature in a moist chamber, setose acervuli developed producing curved, distally tapering and proximately truncated conidia. Single-spore cultures developed equally-shaped conidia measuring 14.5-21.5 (avg. 18.5) × 3-4 (avg. 3.5) µm (n=60) on corn meal agar when mounted in lactic acid. Obtained morphological characters and sequences of the partial actin (GenBank accession, OR208162), beta-tubulin (OR208164), and histone 3 (OR208165) gene identified the isolate as Colletotrichum incanum H.-C. Yang, J.S. Haudenshield & G.L. Hartman. Sequences were identical to those from CBS 133485 (= NRRL 62592, IL6A), ex-type strain of C. incanum (KC110823, KC110814, and KC110796). Partial sequences of the chitin synthase (CHS) gene (OR208163), not available for the ex-type strain, was identical to sequences of other C. incanum strains reported from China (KP145539, ON189040, and OQ613679-OQ613686) or differed in two nucleotide positions (OL471268 and OL471269). The strain from Slovenia was deposited in the CBS biobanks of the Westerdijk Fungal Biodiversity Institute (Utrecht, The Netherlands) as CBS 150848. Pathogenicity of the strain was tested by spraying ca. 3×105 conidia as a watery spore suspension onto each leaf of 6 greenhouse-grown and 3 wk-old common bean plantlets (cv. KIS Amand). Nonsterile commercial substrate (Potgrond H, AGRO-FertiCrop) was used and natural light conditions at ambient temperatures (18-23°C) applied. Sterile water was sprayed on 6, equally grown negative control plants. Treated plants showed small brownish spots after 3 wks similar to those described by Yang et al. (2014) on soybean. Setose acervuli formed within 5 days after detached leaves were incubated in moist chambers. No acervuli formed on negative control plants. Conidia re-isolated from these acervuli and obtained cultures were morphologically identical to originally obtained conidia and cultures and those used for performing the pathogenicity test. Anthracnose is an important disease of common bean attributed to various races of C. lindemuthianum (Sacc. & Magnus) Briosi & Cavara (Nunes et al. 2021). Reporting an additional agent potentially able to cause diseases in common bean and so far not known to occur in Europe is of high relevance as the various genetic bean lines used in Europe may show alternative susceptibility levels to it. However, symptoms caused by C. incanum seem to be less severe as those caused by C. lindemuthianum and the species belongs to the C. spaethianum species complex, whose members have so far not been considered as pathogens of economic importance (Talhinhas & Baroncelli 2021). Yang et al. (2014) based C. incanum on isolates from soybean petioles (USA) and associated it with common bean by re-identifying strain ATCC 64682 obtained by Tu (1990) in Canada. Database queries revealed that it was encountered also on sugar beet (USA; Hanson et al. 2023) and on various crop hosts in China (e.g., chili; Diao et al. 2017), but not in Europe. The work was funded by the Ministry of Agriculture, Forestry and Food and conducted as part of research programs P4-0072 and P4-0431, financed by the Slovenian Research and Innovation Agency ARIS, and the Horizon 2020 project INCREASE funded by the European Union.

First report of Candidatus Phytoplasma australasia (16SrII- subgroup D) associated with virescence of Chia (Salvia hispanica L.) from India.

Chia (Salvia hispanica L., Lamiaceae) is an important commercial and medicinal crop recently popularized in India and widely cultivated in Karnataka (Joy et al., 2022). During the field survey of chia crop diseases, characteristic virescence like symptoms were observed at Main Agricultural Research Station, UAS, Raichur as well as at Mysuru and HD Kote region. The incidence was ranged from 2 - 4 per cent in an area of 30 hectares. Typical symptoms associated with chia are malformed shoot and/or inflorescence axis with reduced floral parts with greenish florets. The stem axis become thick, flattened, leaves are reduced towards terminal region. A total of five phytoplasma suspected samples and five suspected healthy samples were used for identification purpose. The Plant Genomic DNA Miniprep Kit (Sigma Aldrich, USA) was used to extract the DNA from five symptomatic and five asymptomatic samples and the DNA was used as template to amplify the phytoplasma-specific 16S rDNA gene using P1/P7 primers (Deng and Hiruki, 1991; Schneider et al., 1995) followed by nested PCR using R16F2n/R16R2 primers (Gundersen and Lee 1996). The expected 1.25-kb amplicon was detected from the suspected symptomatic samples. Nested PCR products were purified and sequenced from both the directions using ABIX370 Genetic Analyzer (Applied Biosystems, Waltham, MA). The analysis revealed that all five sequences shared 100 per cent identity with Candidatus Phytoplasma aurantifolia (OM649850, ON975012) and Tomato big bud phytoplasma (EF193359). The in-silico RFLP pattern of F2n/R2 primed region of 16S rDNA gene analyzed by using iPhyClassifier (Zhao et al. 2009) revealed that the sequence shared 98.72 per cent nucleotide sequence similarity with coefficient value of 1.00 to the reference strain RFLP pattern of 16Sr group II, subgroup D (witches'-broom disease of lime; U15442). Based on 16SrDNA sequences and in-silico RFLP analysis, the phytoplasma associated with the chia virescence was identified as a member of 16SrII-D group. Further, SecA gene was also amplified from the samples using SecAfor1/SecArev3 primer pair (Hodgetts et al., 2008). All samples produced ~400 bp products and sequenced as detailed above. Sequence analysis by nBLAST revealed 100 per cent similarity to Ca. P. australasia (MW020545) and Ca. P. aurantifolia isolate Idukki Kerala 1 (MK726369) both representing 16SrII-D group phytoplasma. The representative sequence (16Sr: PP359693, PP359694; secA:PP386558, PP386559) were deposited in GenBank. Chia virescence phytoplasma belonging to Ca. phytoplasma australasia has not been reported anywhere. The phytopathological studies associated with chia crop are very limited. Joy et al. (2022) reported the occurrence of foot rot disease caused by Athelia rolfsii. Several hosts are recorded to be associated with 16SrII D phytoplasma which includes china aster, eggplant and crotalaria (Mahadevakumar et al., 2017, Yadav et al., 2016a, b). Now the wide occurrence of the phytoplasma in the area might have transmitted by vectors. The occurrence of virescence is of great importance as it affects the overall yield which reduces the market value. To our knowledge, this is the first report of a group 16SrII-D phytoplasma associated with chia virescence in India.

First report of soybean root rot caused by Fusarium falciforme in the Republic of Korea.

Phytopathogenic Fusarium species causing root and stem rot diseases in susceptible soybean (Glycine max (L.) Merrill) are a major threat to soybean production worldwide. Several Fusarium species have been reported to infect soybean plants in the Republic of Korea, including F. solani, F. oxysporum, F. fujikuroi, and F. graminearum (Cho et al., 2004; Choi et al., 2019; Kang et al., 2020). During the nationwide survey of soybean diseases in 2015, soybean plants showing symptoms of leaf chlorosis, wilting, and shoot death were found in soybean fields in Seosan, Chungnam. Fusarium isolates were obtained from the margins of sterilized necrotic symptomatic and asymptomatic regions of the stem tissues of diseased samples by culturing on potato dextrose agar (PDA). To examine the morphological characteristics, isolates were cultured on PDA at 25°C in the darkness for 10 days. Colonies produced white aerial mycelia with apricot pigments in the medium. Macroconidia were hyaline, slightly curved in shape with 3 or 4 septa, and their average length and width were 34.6± 0.56 µm (31.4 to 37.8 µm) and 4.7±0.16 µm (4.1 to 5.8 µm), respectively (n = 20). Microconidia were elongated, oval with 0 or 1 septum, and their average length and width were 11.4±0.87 and 5.2±0.32 µm, respectively (n = 20). The colonies and conidia exhibited morphological similarities to those of F. falciforme (Xu et al., 2022). Using the primers described by O'Donnell et al. (2008), identity of a representative strain '15-110' was further confirmed by sequencing portions of two genes, the translation elongation factor 1-alpha (EF-1α) and the second largest subunit of RNA polymerase II (RPB2). The two sequences (GenBank accession No. OQ992718 and OR060664) of 15-110 were 99% similar to those of two F. falciforme strains, 21BeanYC6-14 (GenBank accession nos. ON375419 and ON331931), and 21BeanYC6-16 (GenBank accession nos. ON697187 and ON331933). To test the pathogenicity, a single-spore isolate was cultured on carnation leaf agar (CLA) at 25℃ for 10 days. Pathogenicity test was performed by root-cutting assays using 14-day-old soybean seedlings of 'Daewon' and 'Taekwang'. Ten-day-old mycelia of 15-110 were collected from the CLA plates by scraping with distilled water, and the spore suspension was filtered and diluted to 1 × 106 conidia/mL. The roots of the soybean seedlings were partially cut and inoculated by soaking in the diluted spore suspension for two hours. The seedlings were then transplanted into 12 cm plastic pots (11 cm in height) and grown in a growth chamber at 25°C, 14h light/10h dark for 2 weeks. The infected plants exhibited wilting, observed brown discoloration on the root, and eventually died within 2 weeks, whereas the control plants inoculated with sterile water remained healthy. F. falciforme 15-110 was reisolated from infected plants, but not from the uninoculated controls. The morphology of the re-isolated fungus on PDA and its target gene sequences were identical to those of the original colony. To the best of our knowledge, this is the first report of root rot in soybean caused by F. falciforme in the Republic of Korea. Fusarium spp. induce a range of diseases in soybean plants, including root rot, damping-off, and wilt. Given the variable aggressiveness and susceptibility to fungicides among different Fusarium species, it is imperative to identify the Fusarium species posing a threat to soybean production. This understanding is crucial for developing a targeted and tailored disease management strategy to control Fusarium diseases.

Biological Control of Rapeseed Clubroot (Plasmodiophora brassicae) Using the Endophytic Fungus Didymella macrostoma P2.

Didymella macrostoma P2 was isolated from rapeseed (Brassica napus), and it is an endophyte of rapeseed and an antagonist of three rapeseed pathogens, Botrytis cinerea, Leptosphaeria biglobosa, and Sclerotinia sclerotiorum. However, whether P2 has a suppressive effect on infection of rapeseed by the clubroot pathogen Plasmodiophora brassicae remains unknown. This study was conducted to detect production of antimicrobials by P2 and to determine the efficacy of the antimicrobials and P2 pycnidiospores in suppression of rapeseed clubroot. The results showed that cultural filtrates (CFs) of P2 in potato dextrose broth and the substances in pycnidiospore mucilages exuded from P2 pycnidia were inhibitory to P. brassicae. In the indoor experiment, seeds of the susceptible rapeseed cultivar Zhongshuang No. 9 treated with P2 CF and the P2 pycnidiospore suspension (P2 SS, 1 × 107 spores/ml) reduced clubroot severity by 31 to 70% on the 30-day-old seedlings compared with the control (seeds treated with water). P2 was reisolated from the roots of the seedlings in the treatment of P2 SS; the average isolation frequency in the healthy roots (26%) was much higher than that (5%) in the diseased roots. In the field experiment, seeds of another susceptible rapeseed cultivar, Huayouza 50 (HYZ50), treated with P2 CF, P2 CE (chloroform extract of P2 CF, 30 µg/ml), and P2 SS reduced clubroot severity by 29 to 48% on 60-day-old seedlings and by 28 to 59% on adult plants (220 days old) compared with the control treatment. The three P2 treatments on HYZ50 produced significantly (P < 0.05) higher seed yield than the control treatment on this rapeseed cultivar, and they even generated seed yield similar to that produced by the resistant rapeseed cultivar Shengguang 165R in one of the two seasons. These results suggest that D. macrostoma P2 is an effective biocontrol agent against rapeseed clubroot.

Multiple Forms of Resistance to the Phomopsis Stem Canker Pathogens Diaporthe helianthi and D. gulyae in Sunflower.

Phomopsis stem canker of cultivated sunflower (Helianthus annuus L.) can be caused by multiple necrotrophic fungi in the genus Diaporthe, with Diaporthe helianthi and D. gulyae being the most common causal agents in the United States. Infection begins at the leaf margins and proceeds primarily through the vasculature, progressing from the leaf through the petiole to the stem, resulting in formation of brown stem lesions centered around the petiole. Sunflower resistance to Phomopsis stem canker is quantitative and genetically complex. Due to the intricate disease process, resistance is possible at different stages of infection, and multiple forms of defense may contribute to the overall level of quantitative resistance. In this study, sunflower lines exhibiting field resistance to Phomopsis stem canker were evaluated for stem and leaf resistance to multiple isolates of D. helianthi and D. gulyae in greenhouse experiments, and responses to the two species were compared. Additionally, selected resistant and susceptible lines were evaluated for petiole transmission resistance to D. helianthi. Lines with distinct forms of resistance were identified, and results indicated that responses to stem inoculation were strongly correlated (Spearman's coefficient 0.598, P < 0.001) for the two fungal species, while leaf responses were not (Spearman's coefficient 0.396, P = 0.076). These results provide a basis for genetic dissection of distinct forms of sunflower resistance to Phomopsis stem canker and will facilitate combining different forms of resistance to potentially achieve durable control of this disease in sunflower hybrids.

Picarbutrazox Effectiveness Added to a Seed Treatment Mixture for Management of Oomycetes that Impact Soybean in Ohio.

None of the current oomycota fungicides are effective towards all species of Phytophthora, Phytopythium, Globisporangium, and Pythium that affect soybean seed and seedlings in Ohio. Picarbutrazox is a new oomyceticide with a novel mode of action towards oomycete pathogens. Our objectives were to evaluate picarbutrazox to determine (i) baseline sensitivity (EC50) to 189 isolates of 29 species, (ii) the efficacy with a base seed treatment with three cultivars with different levels of resistance in 14 field environments; and (iii) if the rhizosphere microbiome was affected by the addition of the seed treatment on a moderately susceptible cultivar. The mycelial growth of all isolates was inhibited beginning at 0.001 µg, and the EC50 ranged from 0.0013 to 0.0483 µg of active ingredient (a.i.)/ml. The effect of seed treatment was significantly different for plant population and yield in eight of 14 and six of 12 environments, respectively. The addition of picarbutrazox at 1 and 2.5 g of a.i./100 kg seed to the base seed treatment compared to the base alone was associated with higher plant populations and yield in three and one environments, respectively. There was limited impact of the seed treatment mefenoxam 7.5 g of a.i. plus picarbutrazox 1 g of a.i./100 kg seed on the oomycetes detected in the rhizosphere of soybean seedlings collected at the V1 growth stage. Picarbutrazox has efficacy towards a wider range of oomycetes that cause disease on soybean, and this will be another oomyceticide tool to combat early season damping-off in areas where environmental conditions highly favor disease development.

Use of insect pest thresholds in oilseed rape and cereals: is it worth it?

BACKGROUND: Integrated pest management (IPM) uses thresholds to minimize pesticide use, and field monitoring of damaging organisms is an important component to evaluate whether or not thresholds have been breached. However, monitoring requires time and knowledge which impacts costs and benefits. In this study, we evaluated the effects of using insect pest thresholds on time effort, frequency of insecticide treatment and economics in comparison with common farm practices (business as usual) in winter wheat (WW), winter barley (WB) and winter oilseed rape (OSR). This study was done over 2 years (2018 to 2020) on 24 conventionally managed farms in North Rhine-Westphalia (Germany). RESULTS: Farmers spent significantly more time (42 min ha-1/season) monitoring insect pests in OSR than in WW (16 min ha-1/season) and WB (19 min ha-1/season). The use of insecticides in OSR was significantly reduced by 42% in comparison to business as usual. In cereal crops, the use of insecticide treatment was reduced by 50% but this was not significantly different to business as usual. Yields were not significantly reduced when crops were managed in accordance with IPM, and negative differences were small and not significant. However, economically, the costs of monitoring can only be recovered when labor costs and commodity prices are low and insecticide cost is high. CONCLUSION: Insect pest thresholds can help link the policy and environmental goals of insecticide reduction and the agronomic goal of production security. In the future, the time and cost required for monitoring should be reduced through intelligent solutions and tools, increasing the economic viability of monitoring and IPM. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Sensor-Based Quantification of Peanut Disease Defoliation Using an Unmanned Aircraft System and Multispectral Imagery.

Early leaf spot (Passalora arachidicola) and late leaf spot (Nothopassalora personata) are two of the most economically important foliar fungal diseases of peanut, often requiring seven to eight fungicide applications to protect against defoliation and yield loss. Rust (Puccinia arachidis) may also cause significant defoliation depending on season and location. Sensor technologies are increasingly being utilized to objectively monitor plant disease epidemics for research and supporting integrated management decisions. This study aimed to develop an algorithm to quantify peanut disease defoliation using multispectral imagery captured by an unmanned aircraft system. The algorithm combined the Green Normalized Difference Vegetation Index and the Modified Soil-Adjusted Vegetation Index and included calibration to site-specific peak canopy growth. Beta regression was used to train a model for percent net defoliation with observed visual estimations of the variety 'GA-06G' (0 to 95%) as the target and imagery as the predictor (train: pseudo-R2 = 0.71, test k-fold cross-validation: R2 = 0.84 and RMSE = 4.0%). The model performed well on new data from two field trials not included in model training that compared 25 (R2 = 0.79, RMSE = 3.7%) and seven (R2 = 0.87, RMSE = 9.4%) fungicide programs. This objective method of assessing mid-to-late season disease severity can be used to assist growers with harvest decisions and researchers with reproducible assessment of field experiments. This model will be integrated into future work with proximal ground sensors for pathogen identification and early season disease detection.[Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.