Aspergillus flavus pangenome (AflaPan) uncovers novel aflatoxin and secondary metabolite associated gene clusters.

BACKGROUND: Aspergillus flavus is an important agricultural and food safety threat due to its production of carcinogenic aflatoxins. It has high level of genetic diversity that is adapted to various environments. Recently, we reported two reference genomes of A. flavus isolates, AF13 (MAT1-2 and highly aflatoxigenic isolate) and NRRL3357 (MAT1-1 and moderate aflatoxin producer). Where, an insertion of 310 kb in AF13 included an aflatoxin producing gene bZIP transcription factor, named atfC. Observations of significant genomic variants between these isolates of contrasting phenotypes prompted an investigation into variation among other agricultural isolates of A. flavus with the goal of discovering novel genes potentially associated with aflatoxin production regulation. Present study was designed with three main objectives: (1) collection of large number of A. flavus isolates from diverse sources including maize plants and field soils; (2) whole genome sequencing of collected isolates and development of a pangenome; and (3) pangenome-wide association study (Pan-GWAS) to identify novel secondary metabolite cluster genes. RESULTS: Pangenome analysis of 346 A. flavus isolates identified a total of 17,855 unique orthologous gene clusters, with mere 41% (7,315) core genes and 59% (10,540) accessory genes indicating accumulation of high genomic diversity during domestication. 5,994 orthologous gene clusters in accessory genome not annotated in either the A. flavus AF13 or NRRL3357 reference genomes. Pan-genome wide association analysis of the genomic variations identified 391 significant associated pan-genes associated with aflatoxin production. Interestingly, most of the significantly associated pan-genes (94%; 369 associations) belonged to accessory genome indicating that genome expansion has resulted in the incorporation of new genes associated with aflatoxin and other secondary metabolites. CONCLUSION: In summary, this study provides complete pangenome framework for the species of Aspergillus flavus along with associated genes for pathogen survival and aflatoxin production. The large accessory genome indicated large genome diversity in the species A. flavus, however AflaPan is a closed pangenome represents optimum diversity of species A. flavus. Most importantly, the newly identified aflatoxin producing gene clusters will be a new source for seeking aflatoxin mitigation strategies and needs new attention in research.

Assessment of Prickly Sida as a Potential Inoculum Source for Sida Golden Mosaic Virus in Commercial Snap Bean Farms in Georgia, United States.

Sida golden mosaic virus (SiGMV), an obligate pathogen that infects snap beans (Phaseolus vulgaris), is known to infect prickly sida (Sida spinosa L.), which is a common weed in agricultural farms in Georgia. Prickly sida has also been reported as a suitable host of sweetpotato whitefly (Bemisia tabaci), the vector of SiGMV. Despite being a host for both SiGMV and its vector, the role of prickly sida as a reservoir and inoculum source for SiGMV in snap bean farms has not been evaluated. This study was conducted to document the occurrence of SiGMV-infected prickly sida plants and to assess its potential role as a source of SiGMV inoculum in snap bean farms. A survey of 17 commercial snap bean farms conducted in spring 2021 confirmed the presence of SiGMV-infected prickly sida in southern Georgia. In fall 2021 and 2022, on-farm field trials were conducted in four commercial farms where SiGMV-infected prickly sida plants were documented earlier as a part of survey in spring 2021. The spatial distribution and temporal patterns of adult whiteflies and SiGMV on snap bean were compared between macroplots (13.7 × 30.5 m) "with prickly sida" or "without prickly sida" that were at least 232 m apart from each other. We did not observe any consistent differences in counts of adult whiteflies between macroplots with or without prickly sida in the four commercial farms. SiGMV infection was detected earlier and with higher incidences in snap bean macroplots "with prickly sida" compared with macroplots "without prickly sida." An apparent disease gradient was observed in two of the four farms assessed. Higher SiGMV incidences were observed on the edges of macroplots "with prickly sida." These findings indicate prickly sida as a potential natural reservoir and a source for SiGMV spread in snap bean farms in southern Georgia.

Meta-Analysis of the Field Efficacy of Seed- and Soil-Applied Nematicides on Meloidogyne incognita and Rotylenchulus reniformis Across the U.S. Cotton Belt.

Meta-analysis was used to compare yield protection and nematode suppression provided by two seed-applied and two soil-applied nematicides against Meloidogyne incognita and Rotylenchulus reniformis on cotton across 3 years and several trial locations in the U.S. Cotton Belt. Nematicides consisted of thiodicarb- and fluopyram-treated seed, aldicarb and fluopyram applied in furrow, and combinations of the seed treatments and soil-applied fluopyram. The nematicides had no effect on nematode reproduction or root infection but had a significant impact on seed cotton yield response ([Formula: see text]), with an average increase of 176 and 197 kg/ha relative to the nontreated control in M. incognita and R. reniformis infested fields, respectively. However, because of significant variation in yield protection and nematode suppression by nematicides, five or six moderator variables (cultivar resistance [M. incognita only], nematode infestation level, nematicide treatment, application method, trial location, and growing season) were used depending on nematode species. In M. incognita-infested fields, greater yield protection was observed with nematicides applied in furrow and with seed-applied + in-furrow than with solo seed-applied nematicide applications. Most notable of these in-furrow nematicides were aldicarb and fluopyram (>131 g/ha) with or without a seed-applied nematicide compared with thiodicarb. In R. reniformis-infested fields, moderator variables provided no further explanation of the variation in yield response produced by nematicides. Furthermore, moderator variables provided little explanation of the variation in nematode suppression by nematicides in M. incognita- and R. reniformis-infested fields. The limited explanation by the moderator variables on the field efficacy of nematicides in M. incognita- and R. reniformis-infested fields demonstrates the difficulty of managing these pathogens with nonfumigant nematicides across the U.S. Cotton Belt.

Patterns of Seed-to-Seedling Transmission of Xanthomonas citri pv. malvacearum, the Causal Agent of Cotton Bacterial Blight.

Cotton bacterial blight (CBB), caused by Xanthomonas citri pv. malvacearum, was a major disease of cotton in the United States in the early part of the twentieth century. The reemergence of CBB revealed many gaps in our understanding of this important disease. In this study, we employed a wild-type (WT) field isolate of X. citri pv. malvacearum from Georgia (U.S.A.) to generate a nonpathogenic hrcV mutant lacking a functional type-III secretion system (T3SS-). We tagged the WT and T3SS- strains with an auto-bioluminescent Tn7 reporter and compared colonization patterns of CBB-susceptible and CBB-resistant cotton seedlings using macroscopic image analysis and bacterial load enumeration. WT and T3SS- X. citri pv. malvacearum strains colonized cotton cotyledons of CBB-resistant and CBB-susceptible cotton cultivars. However, X. citri pv. malvacearum populations were significantly higher in CBB-susceptible seedlings inoculated with the WT strain. Additionally, WT and T3SS- X. citri pv. malvacearum strains systemically colonized true leaves, although at different rates. Finally, we observed that seed-to-seedling transmission of X. citri pv. malvacearum may involve systemic spread through the vascular tissue of cotton plants. These findings yield novel insights into potential X. citri pv. malvacearum reservoirs for CBB outbreaks.

Effect of In-Furrow Application of Fluopyram on Leaf Spot Diseases of Peanut.

In peanut (Arachis hypogaea) production, in-furrow applications of the premix combination of the succinate-dehydrogenase-inhibitor (SDHI) fungicide and nematicide fluopyram and the insecticide imidacloprid are used primarily for management of nematode pests and for preventing feeding damage on foliage caused by tobacco thrips (Frankliniella fusca). Fluopyram is also active against many fungal pathogens. However, the effect of in-furrow applications of fluopyram on early leaf spot (Passalora arachidicola) or late leaf spot (Nothopassalora personata) has not been characterized. The purpose of this study was to determine the effects of in-furrow applications of fluopyram + imidacloprid or fluopyram alone on leaf spot epidemics. Field experiments were conducted in Tifton, GA in 2015, 2016, and 2018 to 2020. In all experiments, in-furrow applications of fluopyram + imidacloprid provided extended suppression of early leaf spot and late leaf spot epidemics compared with the nontreated control. In 2020, there was no difference between the effects of fluopyram + imidacloprid and fluopyram alone on leaf spot epidemics. Results indicated that fluopyram could complement early-season leaf spot management programs. Use of in-furrow applications of fluopyram should be considered as an SDHI fungicide application for resistance management purposes.

Assessment of Quinone Outside Inhibitor Sensitivity and Frogeye Leaf Spot Race of Cercospora sojina in Georgia Soybean.

Frogeye leaf spot (FLS), caused by the fungal pathogen Cercospora sojina K. Hara, is a foliar disease of soybean (Glycine max L. [Merr.]) responsible for yield reductions throughout the major soybean-producing regions of the world. In the United States, management of FLS relies heavily on the use of resistant cultivars and in-season fungicide applications, specifically within the class of quinone outside inhibitors (QoIs), which has resulted in the development of fungicide resistance in many states. In 2018 and 2019, 80 isolates of C. sojina were collected from six counties in Georgia and screened for QoI fungicide resistance using molecular and in vitro assays, with resistant isolates being confirmed from three counties. Additionally, 50 isolates, including a "baseline isolate" with no prior fungicide exposure, were used to determine the percent reduction of mycelial growth to two fungicides, azoxystrobin and pyraclostrobin, at six concentrations: 0.0001, 0.001, 0.01, 0.1, 1, and 10 µg ml-1. Mycelial growth observed for resistant isolates varied significantly from both sensitive isolates and baseline isolate for azoxystrobin concentrations of 10, 1, 0.1, and 0.01 µg ml-1 and for pyraclostrobin concentrations of 10, 1, 0.1, 0.01, and 0.001 µg ml-1. Moreover, 40 isolates were used to evaluate pathogen race on six soybean differential cultivars by assessing susceptible or resistant reactions. Isolate reactions suggested 12 races of C. sojina present in Georgia, 4 of which have not been previously described. Species richness indicators (rarefaction and abundance-based coverage estimators) indicated that within-county C. sojina race numbers were undersampled in this study, suggesting the potential for the presence of either additional undescribed races or known but unaccounted for races in Georgia. However, no isolates were pathogenic on 'Davis', a differential cultivar carrying the Rcs3 resistance allele, suggesting that the gene is still an effective source of resistance in Georgia.

Adding Epidemiologically Important Meteorological Data to Peanut Rx, the Risk Assessment Framework for Spotted Wilt of Peanut.

Management of disease affecting peanut in the southeastern United States has benefited from extensive field research identifying disease-associated risk factors since the 1990s. An assessment of risk factors associated with tomato spotted wilt (TSW), caused by tomato spotted wilt virus and spread exclusively by thrips, is available to growers through Peanut Rx, a tool developed to inform peanut management decisions. Peanut Rx provides an assessment of relative TSW risk as an index. The assessment provides information about the relative degree to which a field characterized by a specified suite of practices is at risk of crop loss caused by TSW. Loss results when infection occurs, and infection rates are determined, in part, by factors outside a grower's control, primarily the abundance of dispersing, viruliferous thrips. In this study, we incorporated meteorological variables useful for predicting thrips dispersal, increasing the robustness of the Peanut Rx framework in relation to variation in the weather. We used data from field experiments and a large grower survey to estimate the relationships between weather and TSW risk mediated by thrips vectors, and developed an addition to Peanut Rx that proved informative and easy to implement. The expected temporal occurrence of major thrips flights, as a function of heat and precipitation, was translated into the existing risk-point system of Peanut Rx. Results from the grower survey further demonstrated the validity of Peanut Rx for guiding growers' decisions to minimize risk of TSW.

Carbohydrate, glutathione, and polyamine metabolism are central to Aspergillus flavus oxidative stress responses over time.

BACKGROUND: The primary and secondary metabolites of fungi are critical for adaptation to environmental stresses, host pathogenicity, competition with other microbes, and reproductive fitness. Drought-derived reactive oxygen species (ROS) have been shown to stimulate aflatoxin production and regulate in Aspergillus flavus, and may function in signaling with host plants. Here, we have performed global, untargeted metabolomics to better understand the role of aflatoxin production in oxidative stress responses, and also explore isolate-specific oxidative stress responses over time. RESULTS: Two field isolates of A. flavus, AF13 and NRRL3357, possessing high and moderate aflatoxin production, respectively, were cultured in medium with and without supplementation with 15 mM H2O2, and mycelia were collected following 4 and 7 days in culture for global metabolomics. Overall, 389 compounds were described in the analysis which encompassed 9 biological super-pathways and 47 sub-pathways. These metabolites were examined for differential accumulation. Significant differences were observed in both isolates in response to oxidative stress and when comparing sampling time points. CONCLUSIONS: The moderately high aflatoxin-producing isolate, NRRL3357, showed extensive stimulation of antioxidant mechanisms and pathways including polyamines metabolism, glutathione metabolism, TCA cycle, and lipid metabolism while the highly aflatoxigenic isolate, AF13, showed a less vigorous response to stress. Carbohydrate pathway levels also imply that carbohydrate repression and starvation may influence metabolite accumulation at the later timepoint. Higher conidial oxidative stress tolerance and antioxidant capacity in AF13 compared to NRRL3357, inferred from their metabolomic profiles and growth curves over time, may be connected to aflatoxin production capability and aflatoxin-related antioxidant accumulation. The coincidence of several of the detected metabolites in H2O2-stressed A. flavus and drought-stressed hosts also suggests their potential role in the interaction between these organisms and their use as markers/targets to enhance host resistance through biomarker selection or genetic engineering.

Effect of Planting Date and Peanut Cultivar on Epidemics of Late Leaf Spot in Georgia.

Field trials were conducted in 2015 and 2016 in Tifton, GA to determine the effects of planting dates (24 and 27 April, 4, 11, 19, and 26 May 2015; and 11, 18, and 25 April and 2, 9, and 16 May 2016), peanut (Arachis hypogaea) cultivar (Georgia-06G and Georgia-12Y), and seed treatment (nontreated and treated with azoxystrobin, fludioxonil, and mefenoxam) on epidemics of late leaf spot (Nothopassalora personata), plant populations, and peanut yield. Final severity and AUDPC of late leaf spot increased with later planting dates in both years. For most planting dates in 2015 and the final planting date in 2016, final leaf spot severity and AUDPC were lower for Georgia-12Y than for Georgia-06G. Seed treatment increased plant populations for the 27 April and 4 May planting dates in 2015 and across all other treatments in 2016. Yields were higher for Georgia-12Y than for Georgia-06G in both years. In 2015, yields of both cultivars decreased according to linear functions of day of year of planting date, but there was no effect of planting date on yield in 2016. The combination of early planting with Georgia-12Y shows potential utility for management of leaf spot in situations such as organic production where fungicide use is minimal.

Relating Peanut Rx Risk Factors to Epidemics of Early and Late Leaf Spot of Peanut.

Previous research has demonstrated the efficacy of prescription fungicide programs, based upon Peanut Rx, to reduce combined effects of early leaf spot (ELS), caused by Passalora arachidicola (Cercospora arachidicola), and late leaf spot (LLS), caused by Nothopassalora personata (syn. Cercosporidium personatum), but the potential of Peanut Rx to predict each disease has never been formally evaluated. From 2010 to 2016, non-fungicide-treated peanut plots in Georgia and Florida were sampled to monitor the development of ELS and LLS. This resulted in 168 cases (unique combinations of Peanut Rx risk factors) with associated total leaf spot risk points ranging from 40 to 100. Defoliation ranged from 13.9 to 100%, and increased significantly with increasing total risk points (conditional R2 = 0.56; P < 0.001). Leaf spot onset (time in days after planting [DAP] when either leaf spot reached 1% lesion incidence), ELS onset, and LLS onset ranged from 29 to 140, 29 to 142, and 50 to 143 DAP, respectively, and decreased significantly with increasing risk points. Standardized AUDPC of ELS was significantly affected by risk points (conditional R2 = 0.53, P < 0.001), but not for LLS. After removing redundant Peanut Rx factors, planting date, rotation, historical leaf spot prevalence, cultivar, and field history were used as fixed effects in mixed effect regression models to evaluate their contribution to leaf spot, ELS or LLS prediction. Results from mixed effects regression confirmed that the selected Peanut Rx risk factors contributed to the variability of at least one measurement of development of combined or separate epidemics of ELS and LLS, but not all factors affected ELS and LLS equally. Historical leaf spot prevalence, a new potential preplant risk factor, was a consistent predictor of the dominant disease(s) observed in the field. Results presented here demonstrate that Peanut Rx is a very effective tool for predicting leaf spot onset regardless of which leaf spot is predominant, but also suggest that associated risk does not reflect the same development for each disease. These data will be useful for refining thresholds for differentiating high, moderate, and low risk fields, and reevaluating the timing of fungicide applications in reduced input programs with respect to disease onset.

Fusarium wilt of cotton may commonly result from the interaction of Fusarium oxysporum f. sp. vasinfectum with Belonolaimus longicaudatus.

The interaction between Fusarium oxysporum f. sp. vasinfectum (Fov) and Meloidogyne incognita (root-knot nematode) resulting in Fusarium wilt (FW) of cotton is well-known. Although Belonolaimus longicaudatus (sting nematode) can also interact with Fov and cause FW, it has long been believed that virtually all of the FW in Georgia is caused by the interaction of Fov with M. incognita. In recent years, FW has been reported more frequently in Georgia, which suggests that something affecting the disease complex may have changed. In 2015 and 2016, a survey of 27 Georgia cotton fields in 10 counties was conducted. At least 10 soil and stem samples per field were collected from individual plants showing symptoms of FW to quantify plant-parasitic nematode levels and identify Fov races. Fov race 1 was identified in all samples in 2015, but one sample also had the LA110 genotype and another sample also had the LA108 genotype. In 2016, all Fov races and genotypes found in 2015 were present, however, MDS-12 and LA127/140 also were found. Meloidogyne incognita was present in 18% of fields in 2015 and 40% in 2016, whereas B. longicaudatus was present in all fields in 2015 and 75% of fields in 2016. Regardless of whether they occurred separately or together, M. incognita and B. longicaudatus were present, respectively, in 18% and 55% of individual samples in 2015 and 40% and 51% in 2016. However, M. incognita without B. longicaudatus was found in 7% of samples in 2015 and 34% in 2016, whereas B. longicaudatus without M. incognita was found in 45% of samples in 2015 and 44% in 2016. We conclude that Fov race 1 continues to be the dominant race in Georgia and many instances of FW in Georgia may be due to Fov interacting with B. longicaudatus and not M. incognita as previously believed.The interaction between Fusarium oxysporum f. sp. vasinfectum (Fov) and Meloidogyne incognita (root-knot nematode) resulting in Fusarium wilt (FW) of cotton is well-known. Although Belonolaimus longicaudatus (sting nematode) can also interact with Fov and cause FW, it has long been believed that virtually all of the FW in Georgia is caused by the interaction of Fov with M. incognita. In recent years, FW has been reported more frequently in Georgia, which suggests that something affecting the disease complex may have changed. In 2015 and 2016, a survey of 27 Georgia cotton fields in 10 counties was conducted. At least 10 soil and stem samples per field were collected from individual plants showing symptoms of FW to quantify plant-parasitic nematode levels and identify Fov races. Fov race 1 was identified in all samples in 2015, but one sample also had the LA110 genotype and another sample also had the LA108 genotype. In 2016, all Fov races and genotypes found in 2015 were present, however, MDS­12 and LA127/140 also were found. Meloidogyne incognita was present in 18% of fields in 2015 and 40% in 2016, whereas B. longicaudatus was present in all fields in 2015 and 75% of fields in 2016. Regardless of whether they occurred separately or together, M. incognita and B. longicaudatus were present, respectively, in 18% and 55% of individual samples in 2015 and 40% and 51% in 2016. However, M. incognita without B. longicaudatus was found in 7% of samples in 2015 and 34% in 2016, whereas B. longicaudatus without M. incognita was found in 45% of samples in 2015 and 44% in 2016. We conclude that Fov race 1 continues to be the dominant race in Georgia and many instances of FW in Georgia may be due to Fov interacting with B. longicaudatus and not M. incognita as previously believed.

Deciphering drought-induced metabolic responses and regulation in developing maize kernels.

Drought stress conditions decrease maize growth and yield, and aggravate preharvest aflatoxin contamination. While several studies have been performed on mature kernels responding to drought stress, the metabolic profiles of developing kernels are not as well characterized, particularly in germplasm with contrasting resistance to both drought and mycotoxin contamination. Here, following screening for drought tolerance, a drought-sensitive line, B73, and a drought-tolerant line, Lo964, were selected and stressed beginning at 14 days after pollination. Developing kernels were sampled 7 and 14 days after drought induction (DAI) from both stressed and irrigated plants. Comparative biochemical and metabolomic analyses profiled 409 differentially accumulated metabolites. Multivariate statistics and pathway analyses showed that drought stress induced an accumulation of simple sugars and polyunsaturated fatty acids and a decrease in amines, polyamines and dipeptides in B73. Conversely, sphingolipid, sterol, phenylpropanoid and dipeptide metabolites accumulated in Lo964 under drought stress. Drought stress also resulted in the greater accumulation of reactive oxygen species (ROS) and aflatoxin in kernels of B73 in comparison with Lo964 implying a correlation in their production. Overall, field drought treatments disordered a cascade of normal metabolic programming during development of maize kernels and subsequently caused oxidative stress. The glutathione and urea cycles along with the metabolism of carbohydrates and lipids for osmoprotection, membrane maintenance and antioxidant protection were central among the drought stress responses observed in developing kernels. These results also provide novel targets to enhance host drought tolerance and disease resistance through the use of biotechnologies such as transgenics and genome editing.

Phylogenetic Diversity and Host Specialization of Corynespora cassiicola Responsible for Emerging Target Spot Disease of Cotton and Other Crops in the Southeastern United States.

Corynespora cassiicola is a ubiquitous fungus causing emerging plant diseases worldwide, including target spot of cotton, soybean, and tomato, which have rapidly increased in incidence and severity throughout the southeastern United States. The objectives of this study were to understand the causes for the emerging target spot epidemics in the United States by comparing phylogenetic relationships of isolates from cotton, tomato, soybean, and other crop plants and ornamental hosts, and through the determination of the host range of isolates from emerging populations. Fifty-three isolates were sampled from plants in the southeastern United States and 1,380 nucleotides from four nuclear loci were sequenced. Additionally, sequences of the same loci from 23 isolates representing each of the distinct lineages of C. cassiicola described from previous studies were included. Isolates clustered based on host of origin, regardless of the geographic location of sampling. There was no genetic diversity detected among isolates from cotton, which were genetically distinct from isolates from other host species. Furthermore, pathogenicity and virulence assays of 40 isolates from various hosts onto cotton, soybean, tomato, and cucumber showed that isolates from cotton were more aggressive to cotton than those from other hosts. Soybean and tomato were most susceptible to isolates that originated from the same host, providing evidence of host specialization. These results suggest that emerging target spot epidemics in the United States are caused by either the introduction of host-specific isolates or the evolution of more aggressive strains on each host.

Late Leaf Spot Severity and Yield of New Peanut Breeding Lines and Cultivars Grown Without Fungicides.

Peanut (Arachis hypogaea) cultivars with resistance or tolerance to Cercospora arachidicola and/or Cercosporidium personatum, the causes of early and late leaf spot, respectively, are needed for organic production in the southeastern U.S. To determine the potential of new breeding lines for use in such production systems, field experiments were conducted in Tifton, GA, in 2014 and 2015 in which nine breeding lines and two cultivars, Georgia-06G and Georgia-12Y, were grown without foliar fungicide applications. In one set of trials, cultivar Georgia-12Y and most of the breeding lines evaluated had early season vigor ratings, early-season canopy width measurements, final plant populations, and pod yield that were greater than those of standard cultivar Georgia-06G. In those trials, final late leaf spot Florida scale ratings were lower and canopy reflectance measured as the normalized difference vegetation index (NDVI), was higher all the breeding lines than those of Georgia-06G. In another set of trials, two of those same breeding lines had final late leaf spot ratings similar to those of Georgia-12Y in 2014, whereas in 2015, six of those breeding lines had final leaf spot ratings that were lower than those of Georgia-12Y. Yields were similar for Georgia-12Y and all the breeding lines in the Gibbs Farm trials. Across years and breeding lines at the Lang Farm, the relationship between visual estimates of defoliation and NDVI was described by a two sector piecewise regression with NDVI decreasing more rapidly with increasing defoliation above approximately 89%. The utility of NDVI for spot comparisons among breeding lines appears to be limited to situations where there are differences in defoliation. Georgia-12Y and multiple breeding lines evaluated show potential for use in situations such as organic production where acceptable fungicides available for seed treatment and leaf spot control are limited.

Genetic Diversity, Virulence, and Meloidogyne incognita Interactions of Fusarium oxysporum Isolates Causing Cotton Wilt in Georgia.

Locally severe outbreaks of Fusarium wilt of cotton (Gossypium spp.) in South Georgia raised concerns about the genotypes of the causal pathogen, Fusarium oxysporum f. sp. vasinfectum. Vegetative complementation tests and DNA sequence analysis were used to determine genetic diversity among 492 F. oxysporum f. sp. vasinfectum isolates obtained from 107 wilted plants collected from seven fields in five counties. Eight vegetative complementation groups (VCG) were found, with VCG 01117B and VCG 01121 occurring in 66% of the infected plants. The newly recognized VCG 01121 was the major VCG in Berrien County, the center of the outbreaks. All eight VCG resulted in significant increases in the percentages of wilted leaves (27 to 53%) and significant reductions in leaf weight (40 to 67%) and shoot weight (33 to 60%) after being stem punctured into Gossypium hirsutum 'Rowden'. They caused little or no significant reductions in shoot weight and height or increases in foliar symptoms and vascular browning in a soil-infestation assay. Soil infestation with Meloidogyne incognita race 3 (root-knot nematode) alone also failed to cause significant disease. When coinoculated with M. incognita race 3, all VCG caused moderate to severe wilt. Therefore, the VCG identified in this study belong to the vascular-competent pathotype, and should pose similar threats to cotton cultivars in the presence of the root-knot nematode. Use of nematode-resistant cultivars, therefore, is probably the best approach to control the disease in Georgia.

Stress Sensitivity Is Associated with Differential Accumulation of Reactive Oxygen and Nitrogen Species in Maize Genotypes with Contrasting Levels of Drought Tolerance.

Drought stress decreases crop growth, yield, and can further exacerbate pre-harvest aflatoxin contamination. Tolerance and adaptation to drought stress is an important trait of agricultural crops like maize. However, maize genotypes with contrasting drought tolerances have been shown to possess both common and genotype-specific adaptations to cope with drought stress. In this research, the physiological and metabolic response patterns in the leaves of maize seedlings subjected to drought stress were investigated using six maize genotypes including: A638, B73, Grace-E5, Lo964, Lo1016, and Va35. During drought treatments, drought-sensitive maize seedlings displayed more severe symptoms such as chlorosis and wilting, exhibited significant decreases in photosynthetic parameters, and accumulated significantly more reactive oxygen species (ROS) and reactive nitrogen species (RNS) than tolerant genotypes. Sensitive genotypes also showed rapid increases in enzyme activities involved in ROS and RNS metabolism. However, the measured antioxidant enzyme activities were higher in the tolerant genotypes than in the sensitive genotypes in which increased rapidly following drought stress. The results suggest that drought stress causes differential responses to oxidative and nitrosative stress in maize genotypes with tolerant genotypes with slower reaction and less ROS and RNS production than sensitive ones. These differential patterns may be utilized as potential biological markers for use in marker assisted breeding.

Protein profiles reveal diverse responsive signaling pathways in kernels of two maize inbred lines with contrasting drought sensitivity.

Drought stress is a major factor that contributes to disease susceptibility and yield loss in agricultural crops. To identify drought responsive proteins and explore metabolic pathways involved in maize tolerance to drought stress, two maize lines (B73 and Lo964) with contrasting drought sensitivity were examined. The treatments of drought and well water were applied at 14 days after pollination (DAP), and protein profiles were investigated in developing kernels (35 DAP) using iTRAQ (isobaric tags for relative and absolute quantitation). Proteomic analysis showed that 70 and 36 proteins were significantly altered in their expression under drought treatments in B73 and Lo964, respectively. The numbers and levels of differentially expressed proteins were generally higher in the sensitive genotype, B73, implying an increased sensitivity to drought given the function of the observed differentially expressed proteins, such as redox homeostasis, cell rescue/defense, hormone regulation and protein biosynthesis and degradation. Lo964 possessed a more stable status with fewer differentially expressed proteins. However, B73 seems to rapidly initiate signaling pathways in response to drought through adjusting diverse defense pathways. These changes in protein expression allow for the production of a drought stress-responsive network in maize kernels.

Physiological Effects of Meloidogyne incognita Infection on Cotton Genotypes with Differing Levels of Resistance in the Greenhouse.

Greenhouse tests were conducted to evaluate (i) the effect of Meloidogyne incognita infection in cotton on plant growth and physiology including the height-to-node ratio, chlorophyll content, dark-adapted quantum yield of photosystem II, and leaf area; and (ii) the extent to which moderate or high levels of resistance to M. incognita influenced these effects. Cultivars FiberMax 960 BR (susceptible to M. incognita) and Stoneville 5599 BR (moderately resistant) were tested together in three trials, and PD94042 (germplasm, susceptible) and 120R1B1 (breeding line genetically similar to PD94042, but highly resistant) were paired in two additional trials. Inoculation with M. incognita generally resulted in increases in root gall ratings and egg counts per gram of root compared with the noninoculated control, as well as reductions in plant dry weight, root weight, leaf area, boll number, and boll dry weight, thereby confirming that growth of our greenhouse-grown plants was reduced in the same ways that would be expected in field-grown plants. In all trials, M. incognita caused reductions in height-to-node ratios. Nematode infection consistently reduced the area under the height-to-node ratio curves for all genotypes, and these reductions were similar for resistant and susceptible genotypes (no significant genotype × inoculation interaction). Our study is the first to show that infection by M. incognita is associated with reduced chlorophyll content in cotton leaves, and the reduction in the resistant genotypes was similar to that in the susceptible genotypes (no interaction). The susceptible PD94042 tended to have increased leaf temperature compared with the genetically similar but highly resistant 120R1B1 (P < 0.08), likely attributable to increased water stress associated with M. incognita infection.

Cotton leafroll dwarf disease: An enigmatic viral disease in cotton.

TAXONOMY: Cotton leafroll dwarf virus (CLRDV) is a member of the genus Polerovirus, family Solemoviridae. Geographical Distribution: CLRDV is present in most cotton-producing regions worldwide, prominently in North and South America. PHYSICAL PROPERTIES: The virion is a nonenveloped icosahedron with T = 3 icosahedral lattice symmetry that has a diameter of 26-34 nm and comprises 180 molecules of the capsid protein. The CsCl buoyant density of the virion is 1.39-1.42 g/cm3 and S20w is 115-127S. Genome: CLRDV shares genomic features with other poleroviruses; its genome consists of monopartite, single-stranded, positive-sense RNA, is approximately 5.7-5.8 kb in length, and is composed of seven open reading frames (ORFs) with an intergenic region between ORF2 and ORF3a. TRANSMISSION: CLRDV is transmitted efficiently by the cotton aphid (Aphis gossypii Glover) in a circulative and nonpropagative manner. Host: CLRDV has a limited host range. Cotton is the primary host, and it has also been detected in different weeds in and around commercial cotton fields in Georgia, USA. SYMPTOMS: Cotton plants infected early in the growth stage exhibit reddening or bronzing of foliage, maroon stems and petioles, and drooping. Plants infected in later growth stages exhibit intense green foliage with leaf rugosity, moderate to severe stunting, shortened internodes, and increased boll shedding/abortion, resulting in poor boll retention. These symptoms are variable and are probably influenced by the time of infection, plant growth stage, varieties, soil health, and geographical location. CLRDV is also often detected in symptomless plants. CONTROL: Vector management with the application of chemical insecticides is ineffective. Some host plant varieties grown in South America are resistant, but all varieties grown in the United States are susceptible. Integrated disease management strategies, including weed management and removal of volunteer stalks, could reduce the abundance of virus inoculum in the field.