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Three research trials were performed over two spring seasons to assess the performance of bell pepper (Capsicum annuum) cultivars in southern Georgia, USA. Studies were conducted in commercial production fields in Echols and Lowndes counties, which are major bell pepper production areas in Georgia, USA. Eleven cultivars were evaluated, and 10 newer bell pepper cultivars with improved disease resistances for bacterial leaf spot races 1 to 10, tomato spotted wilt virus, and/or phytophthora were compared with ‘Aristotle’, which historically was the most widely grown bell pepper hybrid in the southeastern United States. Our goal was to determine how the new and improved cultivars compared with ‘Aristotle’ for total yield, fruit size distribution, and unmarketable defects in the absence of disease pressure. There were significant differences in fruit size, marketable fruit, and culls between cultivars contrasting their adaptability for production in southern Georgia. The top performers for yield and preferred fruit size for fresh market bell pepper spring production in southern Georgia were ‘Autry’, ‘Standout’, and ‘Antebellum’. Yields and fruit size distribution of these cultivars were similar to ‘Aristotle’ but may be preferred in certain situations due to additional disease resistance. The cultivar, SV 3255 also had higher yields and low defects but had smaller average fruit sizes.

Burkholderia gladioli pv. alliicola, B. cepacia, and B. orbicola are common bacterial pathogens of onion. Onions produce organosulfur thiosulfinate defensive compounds after cellular decompartmentalization. Using whole-genome sequencing and in silico analysis, we identified putative thiosulfinate tolerance gene (TTG) clusters in multiple onion-associated Burkholderia species similar to those characterized in other Allium-associated bacterial endophytes and pathogens. Sequence analysis revealed the presence of three Burkholderia TTG cluster types, with both Type A and Type B being broadly distributed in B. gladioli, B. cepacia, and B. orbicola in both the chromosome and plasmids. Based on isolate natural variation and generation of isogenic strains, we determined the in vitro and in vivo contribution of TTG clusters in B. gladioli, B. cepacia, and B. orbicola. The Burkholderia TTG clusters contributed to enhanced allicin tolerance and improved growth in filtered onion extracts by all three species. TTG clusters also made clear contributions to B. gladioli foliar necrosis symptoms and bacterial populations. Surprisingly, the TTG cluster did not contribute to bacterial populations in onion bulb scales by these three species. Based on our findings, we hypothesize onion-associated Burkholderia may evade or inhibit the production of thiosulfinates in onion bulb tissues. [Graphic: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Abstract 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.

Phytophthora capsici (PCap), which causes Phytophthora root rot, is the most destructive soilborne pathogen for bell pepper (Capsicum annuum L.) production in Georgia, USA. Extensive host range, persistence of inoculum in the soil, and lack of effective chemical control methods make this disease particularly difficult to manage. Resistant cultivars offer a practical solution to manage PCap in affected bell pepper fields. However, most commercial cultivars resistant to PCap are predominantly grown in the northeastern United States. This research aimed to screen commercial bell pepper cultivars for resistance to PCap, marketable yield, fruit size distribution, and overall adaptability for production within the largest growing areas in southern Georgia. Field trials were conducted during Spring 2022 and 2023 in commercial growers’ fields with a known history of PCap infestation to evaluate PCap-resistant bell pepper cultivars across four trials conducted in the three counties (Colquitt, Echols, and Lowndes) responsible for ∼78% of bell pepper production in Georgia. Eleven cultivars were evaluated: nine that claimed PCap resistance and two widely grown PCap susceptible cultivars included for comparison. Phytophthora root rot incidence was very low in these research trials. However, there were significant differences in total yields, marketable yields, fruit size, and unmarketable yields. On the basis of marketable yields for jumbo, extra large, and large-sized fruit, we recommend the PCap-resistant cultivars PS 0994-1819, Paladin, and Mercer for fresh market bell pepper growers. Furthermore, ‘Revolution’ is recommended for processor growers due to higher jumbo-sized fruit yields and lower quality and higher pancaking for the spring season in southern Georgia, USA. Newer cultivars Tarpon and Nitro have a more desirable disease-resistance package; however, ‘Nitro’ had small-sized fruit, and ‘Tarpon’ tended to have lower total yields than current commercial standards.

The resistance (R) gene family in plants is a vital component of the plant defense system, enabling host resistance against pathogens through interactions with pathogen effector proteins. These R genes often encode nucleotide-binding (NB-ARC or N) and leucine-rich-repeat (LRR or L) domains, collectively forming the NLR protein family. The NLR proteins have been widely explored in crops from Poaceae and Brassicaceae, but limited studies are available for crops in other families, including Fabaceae. To address this gap, we conducted a comprehensive genome-wide analysis of putative NLR proteins in nine Fabaceae crops, including Glycine max, Lupinus angustifolius, Medicago truncatula, Pisum sativum, Phaseolus vulgaris, Trifolium pratense, Vigna angularis, Vigna radiata, and Vigna unguiculata. Our study revealed a substantial variation in the number of NLR proteins, independent of genome size. Notably, the NB-ARC domain exhibited a preferential co-occurrence with a specific LRR domain (IPR001611) in Fabaceae. Furthermore, through protein signature analysis, we identified both species-specific and shared domains across the nine crops. By classifying the identified proteins into seven distinct classes (N, L, CN, TN, NL, CNL, and TNL), we observed species-specific clustering within the CN, TN, and CNL classes, reflecting the diversification of species within Fabaceae. This genome-wide study enhances our understanding of the NLR protein repertoire and comprehensive protein signatures in nine Fabaceae species and provides valuable insights into plant defense mechanisms.

Organic vegetable producers in Georgia, USA, utilize a range of amendments to supply nitrogen (N) for crop production. However, differences in soil type, fertilizers and environmental conditions can result in variability in N mineralization rates among commonly utilized organic fertilizers in the region. In this study, the effects of temperature on N mineralization from three commercial organic fertilizers [feather meal (FM), pelleted poultry litter (PPL) and a mixed organic fertilizer (MIX)] in two soil types from Georgia, USA (Cecil sandy clay loam and Tifton loamy sand) were evaluated for 120 d. Net N mineralization (Net Nmin) varied with soil type, fertilizer and temperature. After 120 d, Net Nmin from the FM fertilizer ranged between 41% and 77% of total organic N applied, the MIX fertilizer ranged between 26% and 59% and the PPL fertilizer ranged between 0% and 22% across all soil types and temperatures. Incubation at higher temperatures (20 °C and 30 °C) impacted Net Nmin of FM fertilizer in the Tifton series soil. Temperature and soil type had a relatively minor impact on the potentially mineralizable N of the PPL and MIX fertilizers after 120 d of incubation; however, both factors impacted the rate of fertilizer release shortly after application, which could impact the synchronicity of N availability and plant uptake. Temperature-related differences in the mineralization of organic fertilizers may not be large enough to influence a grower’s decisions regarding N fertilizer inputs for vegetable crop production in the two soils. However, organic fertilizer source will likely play a significant role in N availability during the cropping season.

Pantoea ananatis is an unusual bacterial pathogen that lacks typical virulence determinants yet causes extensive necrosis in onion foliage and bulb tissues. The onion necrosis phenotype is dependent on the expression of the phosphonate toxin, pantaphos, which is synthesized by putative enzymes encoded by the HiVir (high virulence) gene cluster. The genetic contributions of individual hvr genes in HiVir-mediated onion necrosis remain largely unknown, except for the first gene, hvrA (phosphoenolpyruvate mutase, pepM), whose deletion resulted in the loss of onion pathogenicity. In this study, using gene-deletion mutation and complementation, we report that, of the ten remaining genes, hvrB to hvrF are also strictly required for the HiVir-mediated onion necrosis and in-planta bacterial growth, whereas hvrG to hvrJ partially contributed to these phenotypes. As the HiVir gene cluster is a common genetic feature shared among the onion-pathogenic P. ananatis strains that could serve as a useful diagnostic marker of onion pathogenicity, we sought to understand the genetic basis of HiVir-positive yet phenotypically deviant (non-pathogenic) strains. We identified and genetically characterized inactivating single nucleotide polymorphisms in the essential hvr genes of six phenotypically deviant P. ananatis strains. Finally, inoculation of cell-free spent medium of the isopropylthio-β-galactoside (IPTG)-inducible promoter (Ptac)-driven HiVir strain caused P. ananatis–characteristic red onion scale necrosis as well as cell death symptoms in tobacco. Co-inoculation of the spent medium with essential hvr mutant strains restored in-planta populations of the strains to the wild-type level, suggesting that necrotic tissues are important for the proliferation of P. ananatis in onion. [Graphic: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Onions (Allium cepa) are typically planted late fall and harvested in spring in the Vidalia, GA, USA, region. Onions grown here are renowned for their for sweetness and are marketed to consumers as Vidalia onions. High rainfall during the relatively long growing season (4 to 5 months) may result in nitrogen (N) leaching during production. Therefore, fertilizer applications are usually aligned with stages of crop development to ensure nutrient availability for the entire season. Although the impacts of N application rate have been previously investigated for Vidalia onion production, the optimal timing for the final N application of the season has not been determined. The objectives of this study were to determine the optimal timing of the last fertilizer N application (at bulb initiation, during bulb growth, or during bulb maturation) in conjunction with the impact of three N application rates (75, 105, and 135 lb/acre N) on yield and quality in Vidalia onion. Soil N levels were affected by N rate, year, and onion growth stage. In 2020, up to 135 lb/acre N was required to maximize onion yields, and in 2021, onion yields were unchanged among N fertilizer treatments. Final N applications at bulb initiation resulted in greater yields than applications made during bulb growth or bulb maturation. In addition, as the N rate increased and the time of final application occurred later in bulb development, pungency values increased. Incidence of sour skin (Burkholderia cepacia) and center rot (Pantoea sp.) diseases were greater in 2020 compared with 2021 and seemed to be affected by environmental conditions more than N fertilization.

Disease outbreaks of bacterial leaf spot and blight of pepper and tomato often occur in both transplant- and field-production systems worldwide. In some cases, the outbreaks are caused by novel bacterial species. Characterization of these novel bacterial species are critical in developing diagnostic assays and identifying management options for pathogen monitoring and sustainable production, respectively. We characterized strains belonging to novel Pseudomonas species that are responsible for outbreaks in pepper and tomato both in transplant-houses and in production fields in Georgia, USA. Phylogenomic analyses and whole genome sequence indices demonstrated that the pepper and tomato strains belonged to P. capsici. The whole-genome comparison revealed that 13 Pseudomonas strains from diverse isolation sources that were curated in NCBI were indeed P. capsici indicating a potential wide-host range for this bacterial species. Our greenhouse-based host-range assay also indicated that P. capsici strains were pathogenic on pepper, tomato, eggplant, cabbage, lettuce, and watermelon corroborating a wide-host-range. A phylogenetic tree inferred from the whole genome sequence data showed that the P. capsici strains from Georgia (pepper and tomato) were genetically diverse, and were closely related to tomato P. capsici strains from Florida. Genomic presence of traditional bacterial virulence factors in P. capsici strains was also determined. Pseudomonascapsici strains encode one set of type I secretion system, two sets of type II secretion systems, one set of type III secretion system, two sets of type V secretion systems, three sets of type VI secretion systems, and various secondary metabolite gene clusters including lipopeptides. In in-vitro assays, it was demonstrated that six out of seven P. capsici strains (pepper and tomato strains from Georgia) were not sensitive to 0.8 mM CuSO4. When the genomes of copper-tolerant strains were compared with the copper-sensitive strains, it was observed that the former strains encode a cluster of genes related to copper tolerance, which were absent in the genomes of copper-sensitive strains. Considering the ability of P. capsici strains to infect a range of vegetable hosts and possession of a wide range of bacterial virulence factors, secondary metabolites, and copper-tolerance genes, we envision that the management of this pathogen might potentially be a challenge.

Onion center rot is caused by at least four species of genus Pantoea (P. ananatis, P. agglomerans, P. allii, and P. stewartii subsp. indologenes). Critical onion pathogenicity determinants for P. ananatis were recently described, but whether those determinants are common among other onion-pathogenic Pantoea species remains unknown. In this work, we report onion pathogenicity determinants in P. stewartii subsp. indologenes and P. allii. We identified two distinct secondary metabolite biosynthetic gene clusters present separately in different strains of onion-pathogenic P. stewartii subsp. indologenes. One cluster is similar to the previously described HiVir phosphonate biosynthetic cluster identified in P. ananatis and another is a novel putative phosphonate biosynthetic gene cluster, which we named Halophos. The Halophos gene cluster was also identified in P. allii strains. Both clusters are predicted to be phosphonate biosynthetic clusters based on the presence of a characteristic phosphoenolpyruvate phosphomutase (pepM) gene. The deletion of the pepM gene from either HiVir or Halophos clusters in P. stewartii subsp. indologenes caused loss of necrosis on onion leaves and red onion scales and resulted in significantly lower bacterial populations compared with the corresponding wild-type and complemented strains. Seven (halB to halH) of 11 genes (halA to halK) in the Halophos gene cluster are required for onion necrosis phenotypes. The onion nonpathogenic strain PNA15-2 (P. stewartii subsp. indologenes) gained the capacity to cause foliar necrosis on onion via exogenous expression of a minimal seven-gene Halophos cluster (genes halB to halH). Furthermore, cell-free culture filtrates of PNA14-12 expressing the intact Halophos gene cluster caused necrosis on onion leaves consistent with the presence of a secreted toxin. Based on the similarity of proteins to those with experimentally determined functions, we are able to predict most of the steps in Halophos biosynthesis. Together, these observations indicate that production of the toxin phosphonate seems sufficient to account for virulence of a variety of different Pantoea strains, although strains differ in possessing a single but distinct phosphonate biosynthetic cluster. Overall, this is the first report of onion pathogenicity determinants in P. stewartii subsp. indologenes and P. allii. [Graphic: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

A range of organic fertilizers are available for vegetable crops; however, there is a lack of information regarding the performance and rates of organic fertilizers commonly used in the production of Vidalia onion (Allium cepa). Two commercial organic fertilizers, a mixed source organic fertilizer [MIX (10N–0.9P–6.6K)] and a pelleted poultry litter [PPL (5N–1.8P–2.5K)], were evaluated in two soil types at application rates of 0, 100, 150, 200, 250, and 300 lb/acre nitrogen (N) to determine their impact in the production of Vidalia onions in Georgia, USA, with the objective of determining an optimal fertilizer source and application rate. Field trials were conducted in the 2019–20 and 2020–21 growing seasons in Watkinsville, GA, USA (Cecil series sandy clay loam soil) and Tifton, GA, USA (Tifton series loamy sand soil) on certified organic land. There were significant interactions among location, year, and fertilizer application rate for total marketable yield. In Watkinsville, total marketable yields of onions at different N rates ranged between 1320 and 4565 lb/acre in 2019–20, and between 9951 and 28,749 lb/acre in 2020–21. In Tifton, total marketable yields ranged from 3776 to 9264 lb/acre and 7094 to 14,066 lb/acre in the 2019–20 and 2020–21 seasons, respectively. Aboveground onion N accumulation at harvest was affected by an interaction among location, study year, and fertilizer rate. The largest plant N accumulation was in Watkinsville in 2020–21, ranging from 26 to 50.8 lb/acre N in the 0- and 300-lb/acre N treatments, respectively. In 2020, there were no differences in soil inorganic N at harvest between plots receiving the MIX (9 lb/acre N) or PPL (9.8 lb/acre N) in either location. In 2021, soil inorganic N was greater in plots receiving the MIX fertilizer (14.8 lb/acre N) compared with the PPL fertilizer (11.2 lb/acre N). Yields increased linearly with additional fertilizer; therefore, an optimal application rate for organic fertilizers was not determined.

The challenges that sweet potato whitefly (Bemisia tabaci) creates for vegetable production have increased in the southeastern U.S. Growers must use intensive insecticide spray programs to suppress extremely high populations during the fall growing season. Thus, the objective of this study was to evaluate the use of a reflective plastic mulch and an insect row cover as alternative methods to the current grower practices to manage whiteflies in zucchini (Cucurbita pepo) production. Field experiments were conducted with a two-level factorial experimental design of cover and plastic mulch treatments arranged in a randomized complete block design, with four replications in Georgia in 2020 and 2021, and in Alabama in 2021. Cover treatments consisted of an insect row cover installed on zucchini beds at transplanting and removed at flowering and a no-cover treatment, while plastic mulch treatments consisted of reflective silver plastic mulching and white plastic mulching. During all growing seasons, weather conditions were monitored, whitefly populations were sampled weekly, zucchini biomass accumulation was measured at five stages of crop development, and fruit yield was determined at harvesting. Warm and dry weather conditions early in the growing season resulted in increased whitefly populations, regardless of location and year. In general, the reflective silver plastic mulching reduced whitefly populations compared to the conventional white plastic by 87% in Georgia in 2020, 33% in Georgia in 2021, and 30% in Alabama in 2021. The insect row cover treatment reduced whitefly populations to zero until its removal. Consequently, zucchini plants grown with the insect row cover and reflective silver plastic mulching had an increased rate of biomass accumulation due to the lower insect pressure in all locations. Zucchini grown using silver reflective plastic mulch and row covers had an overall increase of 17% and 14% in total yield compared to white plastic mulch and no-cover treatments, respectively. Significant differences in yield among locations were likely due to severe whitefly pressure early in the fall season, and total yields in Georgia in 2020 (11,451 kg ha−1) were 25% lower than in Georgia in 2021 (15,177 kg ha−1) and in Alabama in 2021 (15,248 kg ha−1). In conclusion, silver plastic mulching and row covers reduced the whitefly population and increased biomass accumulation and total yield. These treatments can be considered ready-to-use integrated pest management practices for growers.

Summer squash (Cucurbita pepo L.) is a major vegetable crop produced in Georgia and Florida during the fall season. This production is vulnerable to whitefly (Bemisisia tabaci Genn.)-transmitted viruses that lead to severe yield losses. Over the past several years, whitefly populations have increased during the fall, thus leading to an increase in whitefly-transmitted viruses such as Cucurbit leaf crumple virus (CuLCrV) and Cucurbit yellow stunting disorder virus (CYSDV). Whitefly management for summer squash relies on the use of insecticides and can be costly without providing adequate management of the viruses. Deployment of host resistance to whiteflies and their transmitted viruses (CuLCrV and CYSDV) is the best strategy for mitigating yield loss of summer squash; however, no resistant cultivars are commercially available. In the current study, resistance or tolerance to whiteflies, CuLCrV, and CYSDV was determined for squash germplasm from the U.S. Department of Agriculture (USDA) Germplasm Resources Information Network (GRIN), university breeding programs, and commercial companies in Georgia and Florida across 2 years. In both locations and years, visual virus symptom severity scores were collected and a quantitative polymerase chain reaction (qPCR) was used to determine the CuLCrV viral load and CYSDV presence in Georgia. Whitefly-induced feeding damage was evaluated by directly assessing the intensity of silverleaf symptoms and visual counts of whitefly adults on the foliage in the field or in photographs. Virus symptom severity was lower in C. moschata Duchesne ex Poir. genotypes, namely, PI 550689, PI 550692, PI 550694, PI 653064, and Squash Betternut 900, than in other evaluated genotypes. Two C. pepo accessions were common between both locations for viral severity (PI 442294) or viral severity and viral load (PI 171625). Lower CuLCrV loads were identified in C. ecuadorensis Cutler & Whitaker (PI 540895), and C. okeechobeensis (Small) L.H.Bailey (PI 540900) than other evaluated genotypes. Four genotypes tested negative for CYSDV during both years: C. pepo (PI 507882), C. moschata (PI 483345), C. ecuadorensis (PI 390455), and C. okeechobeensis (PI 540900); they are potential sources of resistance. Six C. moschata accessions (PI 211999, PI 550690, PI 550692, PI 550694, PI 634982, and PI 653064) showed high tolerance to silverleaf disorder and had the lowest adult whitefly counts. Collectively, the accessions identified in the current study are potential sources of resistance or tolerance to whitefly and whitefly-transmitted viruses (CuLCrV and CYSDV).

Pantoea ananatis is a member of a Pantoea species complex that causes center rot of bulb onions (A. cepa) and also infects other Allium crops like leeks (Allium porrum), chives (Allium schoenoprasum), bunching onion or Welsh onion (Allium fistulosum), and garlic (Allium sativum). This pathogen relies on a chromosomal phosphonate biosynthetic gene cluster (HiVir) and a plasmid-borne thiosulfinate tolerance cluster (alt) for onion pathogenicity and virulence, respectively. However, pathogenicity and virulence factors associated with other Allium species remain unknown. We used phenotype-dependent genome-wide association (GWAS) and phenotype-independent gene-pair coincidence (GPC) analyses on a panel of diverse 92 P. ananatis strains, which were inoculated on A. porrum and A. fistulosum × A. cepa under greenhouse conditions. Phenotypic assays showed that, in general, these strains were more aggressive on A. fistulosum × A. cepa as opposed to A. porrum. Of the 92 strains, only six showed highly aggressive foliar lesions on A. porrum compared to A. fistulosum × A. cepa. Conversely, nine strains showed highly aggressive foliar lesions on A. fistulosum × A. cepa compared to A. porrum. These results indicate that there are underlying genetic components in P. ananatis that may drive pathogenicity in these two Allium spp. Based on GWAS for foliar pathogenicity, 835 genes were associated with P. ananatis’ pathogenicity on A. fistulosum × A. cepa whereas 243 genes were associated with bacterial pathogenicity on A. porrum. The Hivir as well as the alt gene clusters were identified among these genes. Besides the ‘HiVir’ and the alt gene clusters that are known to contribute to pathogenicity and virulence from previous studies, genes annotated with functions related to stress responses, a potential toxin-antitoxin system, flagellar-motility, quorum sensing, and a previously described phosphonoglycan biosynthesis (pgb) cluster were identified. The GPC analysis resulted in the identification of 165 individual genes sorted into 39 significant gene-pair association components and 255 genes sorted into 50 significant gene-pair dissociation components. Within the coincident gene clusters, several genes that occurred on the GWAS outputs were associated with each other but dissociated with genes that did not appear in their respective GWAS output. To focus on candidate genes that could explain the difference in virulence between hosts, a comparative genomics analysis was performed on five P. ananatis strains that were differentially pathogenic on A. porrum or A. fistulosum × A. cepa. Here, we found a putative type III secretion system, and several other genes that occurred on both GWAS outputs of both Allium hosts. Further, we also demonstrated utilizing mutational analysis that the pepM gene in the HiVir cluster is important than the pepM gene in the pgb cluster for P. ananatis pathogenicity in A. fistulosum × A. cepa and A. porrum. Overall, our results support that P. ananatis may utilize a common set of genes or gene clusters to induce symptoms on A. fistulosum × A. cepa foliar tissue as well as A. cepa but implicates additional genes for infection on A. porrum.

Onion bulb rot can be caused by multiple plant pathogens including bacterial pathogens. During our routine survey of commercial onion farms in 2014, 2020, and 2021, seven putative Rouxiella spp. strains were isolated from symptomatic onion samples in Georgia, United States. Upon fulfilling Koch’s postulates on onion, a genome analysis was conducted. Whole-genome indices (ANI and dDDH) showed that the strains belonged to Rouxiella badensis. Although the seven R. badensis strains were not pathogenic on onion foliage, the strains were able to cause bulb rot and could also produce necrotic lesions in a red onion scale assay. R. badensis populations increased significantly and to a level comparable to P. ananatis PNA 97-1R in a red onion scale infection assay. The core-genome analysis grouped all onion R. badensis strains from Georgia together, and distinct from R. badensis strains isolated from other sources and locations. Based on the genome analysis of strains (from the current study and available genomes in the repository), type I, III (Ssa-Esc and Inv-Mxi-Spa types), and V secretion systems are present in R. badensis genomes, while type II, IV, and VI secretion systems are absent. However, various secondary metabolite gene clusters were identified from R. badensis genomes, and a thiol/redox-associated enzyme gene cluster similar to the Pantoea alt cluster mediating thiosulfinate tolerance was also present in onion strains of R. badensis. This is the first report of R. badensis as a plant pathogen.

Whitefly, Bemisia tabaci Gennadius (B cryptic species), transmits cucurbit leaf crumple virus (CuLCrV) in a persistent fashion. CuLCrV affects several crops such as squash and snap bean in the southeastern United States. CuLCrV is often found as a mixed infection with whitefly transmitted criniviruses, such as cucurbit yellow stunting disorder virus (CYSDV) in hosts such as squash, or as a single infection in hosts such as snap bean. The implications of different host plants (inoculum sources) with varying infection status on CuLCrV transmission/epidemics is not clear. This study conducted a series of whitefly mediated CuLCrV transmission experiments. In the first experiment, three plants species: squash, snap bean, and tobacco were inoculated by whiteflies feeding on field-collected mixed-infected squash plants. In the second experiment, three plant species, namely squash, snap bean, and tobacco with varying infection status (squash infected with CuLCrV and CYSDV and snap bean and tobacco infected with CuLCrV), were used as inoculum sources. In the third experiment, squash plants with differential CuLCrV accumulation levels and infection status (either singly infected with CuLCrV or mixed infected with CuLCrV and CYSDV) were used as inoculum sources. Irrespective of plant species and its infection status, CuLCrV accumulation in whiteflies was dependent upon the CuLCrV accumulation in the inoculum source plants. Furthermore, differential CuLCrV accumulation in whiteflies resulted in differential transmission, CuLCrV accumulation, and disease phenotype in the recipient squash plants. Overall, results demonstrate that whitefly mediated CuLCrV transmission between host plants follows a virus density dependent phenomenon with implications for epidemics.

Large populations of sweetpotato whiteflies (Bemisia tabaci) have become more regular occurrences during the fall months in parts of the southeastern United States. Large populations of sweetpotato whiteflies have resulted in a significant increase in the incidence of sweetpotato whitefly-transmitted viruses, particularly the cucurbit leaf crumple virus (CuLCrV), which has the potential to cause complete yield loss of fall-planted yellow squash and zucchini (Cucurbita pepo). This study evaluated commercial cultivars of yellow squash and zucchini for resistance against CuLCrV and estimated the yield and fruit quality under environmental conditions during the fall growing season in the southeastern United States. A factorial experimental design was conducted with nine yellow squash and 11 zucchini cultivars during Fall 2017, Fall 2018, and Fall 2019 in Tifton, GA. In situ weather stations monitored the weather conditions during growing seasons, and yellow pest monitor cards monitored the sweetpotato whitefly populations in 2018 and 2019. During all growing seasons, yellow squash and zucchini plants were rated weekly for the severity of CuLCrV. Harvests were conducted 12 times during each season, and fruit were graded as fancy, medium, and culls. Rainfall distribution directly affected the sweetpotato whitefly populations during the production year. In 2018, frequent rainfall events created field conditions that reduced the sweetpotato whitefly populations compared with those during 2017 and 2019. The severity of CuLCrV negatively impacted both the yield and quality of fruit of yellow squash and zucchini, and no resistant commercial cultivars of yellow squash or zucchini were identified. Nonetheless, the yellow squash cultivars Lioness, Gold Prize, and Grand Prize, and the zucchini cultivars SV6009YG and SV0914YG had the highest yields when subjected to the highest sweetpotato whitefly populations during the study.

A two-year study was conducted in 2019 and 2020 to characterize the spatial distribution of whitefly-transmitted virus complex (WTVC) in experimental and commercial yellow squash (Cucurbita pepo) fields in southern Georgia, USA. Field trials planted in Tifton, Georgia, USA were comprised of 30 rows of squash (variety: Gentry) divided into 300 quadrats. Six commercial fields in Tift, Worth, and Colquitt Counties in Georgia, USA were surveyed. In each field, 10 rows of plants per field were partitioned into 70 quadrats so that their approximate location could be mapped and disease incidence tracked in space over time. Plants in each quadrat were visually assessed for TVC symptoms at seven-day intervals (experimental fields) or at fourteen-day intervals (commercial fields). The spatial distribution of WTVC in a one-dimensional space was determined through ordinary runs analysis and in two-dimensional space through spatial autocorrelation analysis and spatial analysis by distance indices (SADIE). Ordinary runs analysis down and between rows suggested that disease was more likely to be transmitted from infected plants within rows than across rows. WTVC incidence in space was positively correlated with adult whiteflies on squash leaves taken one or two weeks prior to the assessment of virus incidence coinciding with latent period of Geminiviruses. SADIE generated disease severity maps indicated a shift from sparse and randomly distributed disease early in the infection cycle to denser, more aggregated patterns later in time in both experimental and commercial fields. Contour plots of interpolated indices from SADIE analysis suggested an edge effect on the spatial distribution of WTVC in experimental fields as well as in two of the commercial fields. An understanding of the shift from random to aggregated distribution and the edge effect on WTVC incidence are considerations for refining current management strategies.

The use of resistant cultivars against Xanthomonas campestris pv. campestris (Xcc) is considered a critical management practice for black rot (BR) management in cabbage (Brassica oleracea var. capitata). Although most studies that have evaluated resistance to BR were conducted in greenhouses without accounting for yield, there is a clear need to investigate cultivar performance under field conditions. The objectives of this study were to evaluate commercial cabbage cultivars for resistance to BR and determine yield and head quality under field conditions. Field experiments with eight cultivars (Acclaim, Bravo, Capture, Celebrate, Cheers, Melissa, Monterey, and TCA-549) were conducted in two cabbage growing seasons, Fall 2018 and Spring 2019. Fields were spray-inoculated with Xcc (3 × 105 cfu/mL) at 35 and 49 days after transplanting in Fall 2018 and Spring 2019, respectively. Cabbage BR severity was evaluated at weekly intervals starting from 7 days postinoculation (DPI) until harvest. Marketable and unmarketable yields and cabbage head quality were measured at harvest. Cabbage BR symptoms were detected in all tested cultivars for both growing seasons with initial symptoms observed as early as 28 and 21 DPI in Fall 2018 and Spring 2019, respectively. Severity of cabbage BR at harvest was significantly greater in Fall 2018 compared with Spring 2019, whereas marketable yield was significantly higher in Spring 2019 (45,169 lb/acre) compared with Fall 2018 (26,370 lb/acre). In both growing seasons, ‘TCA-549’ had the lowest severity of BR and ‘Melissa’ had the highest severity of BR. Area under the disease progress curve (AUDPC) was 175 and 13 for ‘TCA-549’ in Fall 2018 and Spring 2019, respectively. The AUDPC for ‘Melissa’ in Fall 2018 and Spring 2019 were 2376 and 905, respectively. Regardless of growing season, cabbage marketable yield was higher for ‘Acclaim’ (51,760 lb/acre) compared with all other cultivars; however, there was no significant difference between Acclaim and TCA-549 (42,934 lb/acre) for cabbage marketable yield. Cabbage marketable yield was the lowest for cultivars Melissa (18,275 lb/acre) and Capture (24,236 lb/acre). Overall, there was a significant correlation between BR disease severity and cabbage marketable and unmarketable yields. Increasing the BR severity decreased cabbage marketability due to an increase in unmarketable yields. Continued development of cultivar resistance to BR is important for cabbage production in the southeastern United States, given the favorable conditions for disease development. The use of cultivars with low susceptibility, pathogen-free seeds, crop rotation, and proper spraying programs should be considered the best management practice for BR disease.

Sida golden mosaic virus (SiGMV) was first detected from snap bean (Phaseolus vulgaris L.) in Florida in 2006 and recently in Georgia in 2018. Since 2018, it has caused significant economic losses to snap bean growers in Georgia. This study, using a SiGMV isolate field-collected from prickly sida (Sida spinosa L.), examined the putative host range, vector-mediated transmission, and SiGMV-modulated effects on host–vector interactions. In addition, this study analyzed the phylogenetic relationships of SiGMV with other begomoviruses reported from Sida spp. Host range studies confirmed that SiGMV can infect seasonal crops and perennial weed species such as snap bean, hollyhock (Alcea rosea L.), marsh mallow (Althaea officinalis L.), okra (Abelmoschus esculentus (L.) Moench), country mallow (Sida cordifolia L.), prickly sida (S. spinosa), and tobacco (Nicotiana tabacum L.). The incidence of infection ranged from 70 to 100%. SiGMV-induced symptoms and virus accumulation varied between hosts. The vector, Bemisia tabaci Gennadius, was able to complete its life cycle on all plant species, irrespective of SiGMV infection status. However, SiGMV infection in prickly sida and country mallow positively increased the fitness of whiteflies, whereas SiGMV infection in okra negatively influenced whitefly fitness. Whiteflies efficiently back-transmitted SiGMV from infected prickly sida, hollyhock, marsh mallow, and okra to snap bean, and the incidence of infection ranged from 27 to 80%. Complete DNA-A sequence from this study shared 97% identity with SiGMV sequences reported from Florida and it was determined to be closely related with sida viruses reported from the New World. These results suggest that SiGMV, a New World begomovirus, has a broad host range that would allow its establishment in the farmscapes/landscapes of the southeastern United States and is an emerging threat to snap bean and possibly other crops.

Pantoea ananatis, a gram negative and facultative anaerobic bacterium is a member of a Pantoea spp. complex that causes center rot of onion, which significantly affects onion yield and quality. This pathogen does not have typical virulence factors like type II or type III secretion systems but appears to require a biosynthetic gene-cluster, HiVir/PASVIL (located chromosomally comprised of 14 genes), for a phosphonate secondary metabolite, and the ‘alt’ gene cluster (located in plasmid and comprised of 11 genes) that aids in bacterial colonization in onion bulbs by imparting tolerance to thiosulfinates. We conducted a deep pan-genome-wide association study (pan-GWAS) to predict additional genes associated with pathogenicity in P. ananatis using a panel of diverse strains (n = 81). We utilized a red-onion scale necrosis assay as an indicator of pathogenicity. Based on this assay, we differentiated pathogenic (n = 51)- vs. non-pathogenic (n = 30)-strains phenotypically. Pan-genome analysis revealed a large core genome of 3,153 genes and a flexible accessory genome. Pan-GWAS using the presence and absence variants (PAVs) predicted 42 genes, including 14 from the previously identified HiVir/PASVIL cluster associated with pathogenicity, and 28 novel genes that were not previously associated with pathogenicity in onion. Of the 28 novel genes identified, eight have annotated functions of site-specific tyrosine kinase, N-acetylmuramoyl-L-alanine amidase, conjugal transfer, and HTH-type transcriptional regulator. The remaining 20 genes are currently hypothetical. Further, a core-genome SNPs-based phylogeny and horizontal gene transfer (HGT) studies were also conducted to assess the extent of lateral gene transfer among diverse P. ananatis strains. Phylogenetic analysis based on PAVs and whole genome multi locus sequence typing (wgMLST) rather than core-genome SNPs distinguished red-scale necrosis inducing (pathogenic) strains from non-scale necrosis inducing (non-pathogenic) strains of P. ananatis. A total of 1182 HGT events including the HiVir/PASVIL and alt cluster genes were identified. These events could be regarded as a major contributing factor to the diversification, niche-adaptation and potential acquisition of pathogenicity/virulence genes in P. ananatis.

The soil-borne pathogens, particularly Fusarium oxysporum f. sp. niveum (FON) and southern root-knot nematode (RKN, Meloidogyne incognita) are the major threats to watermelon production in the southeastern United States. The role of soil micronutrients on induced resistance (IR) to plant diseases is well-documented in soil-based media. However, soil-based media do not allow us to determine the contribution of individual micronutrients in the induction of IR. In this manuscript, we utilized hydroponics-medium to assess the effect of controlled application of micronutrients, including iron (Fe), manganese (Mn), and zinc (Zn) on the expression of important IR genes (PR1, PR5, and NPR1 from salicylic acid (SA) pathway, and VSP, PDF, and LOX genes from jasmonic acid (JA) pathway) in watermelon seedlings upon inoculation with either FON or RKN or both. A subset of micronutrient-treated plants was inoculated (on the eighth day of micronutrient application) with FON and RKN (single or mixed inoculation). The expression of the IR genes in treated and control samples was evaluated using qRT-PCR. Although, significant phenotypic differences were not observed with respect to the severity of wilt symptoms or RKN galling with any of the micronutrient treatments within the 30-day experimental period, differences in the induction of IR genes were considerably noticeable. However, the level of gene expression varied with sampling period, type and concentration of micronutrients applied, and pathogen inoculation. In the absence of pathogens, micronutrient applications on the seventh day, in general, downregulated the expression of the majority of the IR genes. However, pathogen inoculation preferentially either up- or down-regulated the expression levels of the IR genes at three days post-inoculation depending on the type and concentration of micronutrients. The results demonstrated here indicate that micronutrients in watermelon may potentially make watermelon plants susceptible to infection by FON and RKN. However, upon infection the IR genes are significantly up-regulated that they may potentially aid the prevention of further infection via SA- and JA-pathways. This is the first demonstration of the impact of micronutrients affecting IR in watermelon against FON and RKN infection.

Pantoea ananatis is a gram-negative bacterium and the primary causal agent of center rot of onions in Georgia. Previous genomic studies identified two virulence gene clusters, HiVir and alt, associated with center rot. The HiVir gene cluster is required to induce necrosis on onion tissues via synthesis of pantaphos, (2-hydroxy[phosphono-methyl)maleate), a phosphonate phytotoxin. The alt gene cluster aids in tolerance to thiosulfinates generated during onion tissue damage. Whole genome sequencing of other Pantoea species suggests that these gene clusters are present outside of P. ananatis. To assess the distribution of these gene clusters, two PCR primer sets were designed to detect the presence of HiVir and alt. Two hundred fifty-two strains of Pantoea spp. were phenotyped using the red onion scale necrosis (RSN) assay and were genotyped using PCR for the presence of these virulence genes. A diverse panel of strains from three distinct culture collections comprised of 24 Pantoea species, 41 isolation sources, and 23 countries, collected from 1946–2019, was tested. There is a significant association between the alt PCR assay and Pantoea strains recovered from symptomatic onion (P < 0.001). There is also a significant association of a positive HiVir PCR and RSN assay among P. ananatis strains but not among Pantoea spp., congeners. This may indicate a divergent HiVir cluster or different pathogenicity and virulence mechanisms. Last, we describe natural alt positive [RSN+/HiVir+/alt+] P. ananatis strains, which cause extensive bulb necrosis in a neck-to-bulb infection assay compared to alt negative [RSN+/HiVir+/alt–] P. ananatis strains. A combination of assays that include PCR of virulence genes [HiVir and alt] and an RSN assay can potentially aid in identification of onion-bulb-rotting pathogenic P. ananatis strains.

Begomoviruses are transmitted by several cryptic species of the sweetpotato whitefly, Bemisia tabaci (Gennadius), in a persistent and circulative manner. Upon virus acquisition and circulative translocation within the whitefly, a multitude of molecular interactions occur. This study investigated the differentially expressed transcript profiles associated with the acquisition of the Old World monopartite begomovirus, tomato yellow leaf curl virus (TYLCV), and two New World bipartite begomoviruses, sida golden mosaic virus (SiGMV) and cucurbit leaf crumple virus (CuLCrV), in two invasive B. tabaci cryptic species, Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED). A total of 881 and 559 genes were differentially expressed in viruliferous MEAM1 and MED whiteflies, respectively, compared with their non-viruliferous counterparts, of which 146 genes were common between the two cryptic species. For both cryptic species, the number of differentially expressed genes (DEGs) associated with TYLCV and SiGMV acquisition were higher compared with DEGs associated with CuLCrV acquisition. Pathway analysis indicated that the acquisition of begomoviruses induced differential changes in pathways associated with metabolism and organismal systems. Contrasting expression patterns of major genes associated with virus infection and immune systems were observed. These genes were generally overexpressed and underexpressed in B. tabaci MEAM1 and MED adults, respectively. Further, no specific expression pattern was observed among genes associated with fitness (egg production, spermatogenesis, and aging) in viruliferous whiteflies. The weighted gene correlation network analysis of viruliferous B. tabaci MEAM1 and MED adults identified different hub genes potentially implicated in the vector competence and circulative tropism of viruses. Taken together, the results indicate that both vector cryptic species and the acquired virus species could differentially affect gene expression.

Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED) are two of the most invasive members of the sweetpotato whitefly, Bemisia tabaci, cryptic species complexes and are efficient vectors of begomoviruses. Bemisia tabaci MEAM1 is the predominant vector of begomoviruses in open-field vegetable crops in the southeastern United States. However, recently B. tabaci MED also has been detected in the landscape outside of greenhouses in Florida and Georgia. This study compared the transmission efficiency of one Old-World (OW) and two New-World (NW) begomoviruses prevalent in the southeastern United States, viz., tomato yellow leaf curl virus (TYLCV), cucurbit leaf crumple virus (CuLCrV), and sida golden mosaic virus (SiGMV) between B. tabaci MEAM1 and B. tabaci MED. Bemisia tabaci MEAM1 efficiently transmitted TYLCV, CuLCrV, or SiGMV, whereas B. tabaci MED only transmitted TYLCV. Percent acquisition and retention of OW TYLCV following a 72 h acquisition access period was significantly higher for B. tabaci MED than B. tabaci MEAM1. In contrast, B. tabaci MEAM1 acquired and retained significantly more NW bipartite begomoviruses, CuLCrV or SiGMV, than B. tabaci MED. Quantitative analysis (qPCR) of virus DNA in whitefly internal tissues revealed reduced accumulation of CuLCrV or SiGMV in B. tabaci MED than in B. tabaci MEAM1. Fluorescent in situ hybridization (FISH) showed localization of CuLCrV or SiGMV in the midgut of B. tabaci MED and B. tabaci MEAM1. However, localization of CuLCrV or SiGMV was only observed in the primary salivary glands of B. tabaci MEAM1 and not B. tabaci MED. TYLCV localization was observed in all internal tissues of B. tabaci MEAM1 and B. tabaci MED. Overall, results demonstrate that both B. tabaci MEAM1 and B. tabaci MED are efficient vectors of OW TYLCV. However, for the NW begomoviruses, CuLCrV and SiGMV, B. tabaci MEAM1 seems to a better vector.

The sweetpotato whitefly, Bemisia tabaci MEAM1 Gennadius (Hemiptera: Aleyrodidae), and the complex of viruses it transmits are major limiting factors to squash production in the southeastern United States. At this time, insecticides are extensively relied upon for the management of whiteflies and, indirectly, whitefly-transmitted viruses. The development of a multi-faceted, integrated pest management (IPM) program is needed to increase the sustainability and profitability of squash production. Experiments in 2018 and 2019 evaluated the effects of insect exclusion netting (IEN) in combination with selected pesticides on whitefly population dynamics and virus incidence in greenhouse-grown squash seedlings. Field experiments from 2018 to 2021 evaluated the effects of mulch type (UV-reflective mulch, live mulch, and white plastic mulch), row covers, and insecticides on whitefly population dynamics, silver leaf disorder (SSL) intensity, virus symptom severity, and marketable yield. IEN significantly reduced whiteflies and virus incidence on squash seedlings in the greenhouse study. In the field mulch study, lower whitefly abundance and SSL intensity, as well as reduced virus symptom severity, were observed in plots with reflective mulch compared with white plastic or live mulch. In the insecticide/row cover study, whitefly abundance, SSL intensity, and virus symptom severity were lowest in the row cover and cyantraniliprole- and flupyradifurone-treated plots. Field plots with row covers and those with UV-reflective mulch consistently produced the greatest marketable yields. These findings demonstrate that growers can reduce whitefly and virus pressure and preserve yields in squash production in the southeastern United States by combining cultural and chemical tactics, including row covers, UV-reflective mulch, and select insecticides.

This review provides a synopsis of transcriptional responses pertaining to interactions between plant viruses and the insect vectors that transmit them in diverse modes. In the process, it attempts to catalog differential gene expression pertinent to virus–vector interactions in vectors such as virus reception, virus cell entry, virus tissue tropism, virus multiplication, and vector immune responses. Whiteflies, leafhoppers, planthoppers, and thrips are the main insect groups reviewed, along with aphids and leaf beetles. Much of the focus on gene expression pertinent to vector–virus interactions has centered around whole-body RNA extraction, whereas data on virus-induced tissue-specific gene expression in vectors is limited. This review compares transcriptional responses in different insect groups following the acquisition of non-persistent, semi-persistent, and persistent (non-propagative and propagative) plant viruses and identifies parallels and divergences in gene expression patterns. Understanding virus-induced changes in vectors at a transcriptional level can aid in the identification of candidate genes for targeting with RNAi and/or CRISPR editing in insect vectors for management approaches.

Pantoea stewartii subsp. indologenes (Psi) is a causative agent of leafspot on foxtail millet and pearl millet; however, novel strains were recently identified that are pathogenic on onions. Our recent host range evaluation study identified two pathovars; P. stewartii subsp. indologenes pv. cepacicola pv. nov. and P. stewartii subsp. indologenes pv. setariae pv. nov. that are pathogenic on onions and millets or on millets only, respectively. In the current study, we developed a pan-genome using the whole genome sequencing of newly identified/classified Psi strains from both pathovars [pv. cepacicola (n = 4) and pv. setariae (n = 13)]. The full spectrum of the pan-genome contained 7030 genes. Among these, 3546 (present in genomes of all 17 strains) were the core genes that were a subset of 3682 soft-core genes (present in ≥16 strains). The accessory genome included 1308 shell genes and 2040 cloud genes (present in ≤2 strains). The pan-genome showed a clear linear progression with >6000 genes, suggesting that the pan-genome of Psi is open. Comparative phylogenetic analysis showed differences in phylogenetic clustering of Pantoea spp. using PAVs/wgMLST approach in comparison with core genome SNPs-based phylogeny. Further, we conducted a horizontal gene transfer (HGT) study using Psi strains from both pathovars along with strains from other Pantoea species, namely, P. stewartii subsp. stewartii LMG 2715T, P. ananatis LMG 2665T, P. agglomerans LMG L15, and P. allii LMG 24248T. A total of 317 HGT events among four Pantoea species were identified with most gene transfer events occurring between Psi pv. cepacicola and Psi pv. setariae. Pan-GWAS analysis predicted a total of 154 genes, including seven gene-clusters, which were associated with the pathogenicity phenotype (necrosis on seedling) on onions. One of the gene-clusters contained 11 genes with known functions and was found to be chromosomally located.

Zinc (Zn) accumulation and deficiency affect plant response to pests and diseases differently in varying pathosystems. The concentrations of Zn in plants aid in priming defense signaling pathways and help in enhanced structural defenses against plant pathogens. Studies are lacking on how concentrations of Zn in watermelon plants influence defense against two important soil-borne pathogens: Fusarium oxysporum f. sp. niveum (FON) and southern root-knot nematode (RKN, Meloidogyne incognita). In this study a comparative transcriptomics evaluation of watermelon plants in response to high (1.2 ppm) and low (0.2 ppm) levels of Zn were determined. Differential transcript-level responses differed in watermelon plants when infected with FON or RKN or both under high- and low-Zn treatment regimes in a controlled hydroponics system. Higher numbers of differentially expressed genes (DEGs) were observed in high-Zn-treated than in low-Zn-treated non-inoculated plants, in plants inoculated with FON alone and in plants inoculated with RKN alone. However, in the co-inoculated system, low-Zn treatment had higher DEGs as compared to high-Zn treatment. In addition, most DEGs were significantly enriched in hormone signal transduction and MAPK signaling pathway, suggesting an induction of systemic resistance with high-Zn concentrations. Taken together, this study substantially expands transcriptome data resources and suggests a molecular potential framework for watermelon-Zn interaction in FON and RKN.

The production and quality of Phaseolusvulgaris (snap bean) have been negatively impacted by leaf crumple disease caused by two whitefly-transmitted begomoviruses: cucurbit leaf crumple virus (CuLCrV) and sida golden mosaic Florida virus (SiGMFV), which often appear as a mixed infection in Georgia. Host resistance is the most economical management strategy against whitefly-transmitted viruses. Currently, information is not available with respect to resistance to these two viruses in commercial cultivars. In two field seasons (2018 and 2019), we screened Phaseolus spp. genotypes (n = 84 in 2018; n = 80 in 2019; most of the genotypes were common in both years with a few exceptions) for resistance against CuLCrV and/or SiGMFV. We also included two commonly grown Lima bean (Phaseolus lunatus) varieties in our field screening. Twenty Phaseolus spp. genotypes with high to moderate-levels of resistance (disease severity ranging from 5%–50%) to CuLCrV and/or SiGMFV were identified. Twenty-one Phaseolus spp. genotypes were found to be highly susceptible with a disease severity of ≥66%. Furthermore, based on the greenhouse evaluation with two genotypes-each (two susceptible and two resistant; identified in field screen) exposed to viruliferous whiteflies infected with CuLCrV and SiGMFV, we observed that the susceptible genotypes accumulated higher copy numbers of both viruses and displayed severe crumple severity compared to the resistant genotypes, indicating that resistance might potentially be against the virus complex rather than against the whiteflies. Adult whitefly counts differed significantly among Phaseolus genotypes in both years. The whole genome of these Phaseolus spp. [snap bean (n = 82); Lima bean (n = 2)] genotypes was sequenced and genetic variability among them was identified. Over 900 giga-base (Gb) of filtered data were generated and >88% of the resulting data were mapped to the reference genome, and SNP and Indel variants in Phaseolus spp. genotypes were obtained. A total of 645,729 SNPs and 68,713 Indels, including 30,169 insertions and 38,543 deletions, were identified, which were distributed in 11 chromosomes with chromosome 02 harboring the maximum number of variants. This phenotypic and genotypic information will be helpful in genome-wide association studies that will aid in identifying the genetic basis of resistance to these begomoviruses in Phaseolus spp.

Bemisia tabaci is a whitefly species complex comprising important phloem feeding insect pests and plant virus vectors of many agricultural crops. Middle East–Asia Minor 1 (MEAM1) and Mediterranean (MED) are the two most invasive members of the B. tabaci species complex worldwide. The diversity of agroecosystems invaded by B. tabaci could potentially influence their population structure, but this has not been assessed at a farmscape level. A farmscape in this study is defined as heterogenous habitat with crop and non-crop areas spanning ~8 square kilometers. In this study, mitochondrial COI gene (mtCOI) sequences and six microsatellite markers were used to examine the population structure of B. tabaci MEAM1 colonizing different plant species at a farmscape level in Georgia, United States. Thirty-five populations of adult whiteflies on row and vegetable crops and weeds across major agricultural regions of Georgia were collected from fifteen farmscapes. Based on morphological features and mtCOI sequences, five species/cryptic species of whiteflies (B. tabaci MEAM1, B. tabaci MED, Dialeurodes citri, Trialeurodes abutiloneus, T. vaporariorum) were found. Analysis of 102 mtCOI sequences revealed the presence of a single B. tabaci MEAM1 haplotype across farmscapes in Georgia. Population genetics analyses (AMOVA, PCA and STRUCTURE) of B. tabaci MEAM1 (microsatellite data) revealed only minimal genetic differences among collected populations within and among farmscapes. Overall, our results suggest that there is a high level of gene flow among B. tabaci MEAM1 populations among farmscapes in Georgia. Frequent whitefly population explosions driven by a single or a few major whitefly-suitable hosts planted on a wide spatial scale may be the key factor behind the persistence of a single panmictic population over Georgia’s farmscapes. These population structuring effects are useful for delineating the spatial scale at which whiteflies must be managed and predicting the speed at which alleles associated with insecticide resistance might spread.

Mixed infection of plant viruses is ubiquitous in nature and can affect virus–plant–vector interactions differently than single virus infection. While several studies have examined virus–virus interactions involving mixed virus infection, relatively few have examined effects of mixed virus infection on vector preference and fitness, especially when multiple vectors are involved. This study explored how single and mixed viral infection of a non-persistently transmitted cucumber mosaic virus (CMV) and propagative and persistently-transmitted tomato spotted wilt orthotospovirus (TSWV) in pepper, Capsicum annum L., influenced the preference and fitness of their vectors, the green peach aphid, Myzus persicae (Sulzer), and the tobacco thrips, Frankliniella fusca (Hinds), respectively. In general, mixed infected plants exhibited severe symptoms compared with individually infected plants. An antagonistic interaction between the two viruses was observed when CMV titer was reduced following mixed infection with TSWV in comparison with the single infection. TSWV titer did not differ between single and mixed infection. Myzus persicae settling preference and median developmental were not significantly different between CMV and/or TSWV-infected and non-infected plants. Moreover, M. persicae fecundity did not differ between CMV-infected and non-infected pepper plants. However, M. persicae fecundity was substantially greater on TSWV-infected plants than non-infected plants. Myzus persicae fecundity on mixed-infected plants was significantly lower than on singly-infected and non-infected plants. Frankliniella fusca fecundity was higher on CMV and/or TSWV-infected pepper plants than non-infected pepper plants. Furthermore, F. fusca-induced feeding damage was higher on TSWV-infected than on CMV-infected, mixed-infected, or non-infected pepper plants. Overall, our results indicate that the effects of mixed virus infection on vectors were not different from those observed following single virus infection. Virus-induced host phenotype-modulated effects were realized on both specific and non-specific vectors, suggesting crosstalk involving all vectors and viruses in this pathosystem. The driving forces of these interactions need to be further examined. The effects of interactions between two viruses and two vectors towards epidemics of one or both viruses also need to be examined.

Pantoea ananatis is a member of the family Enterobacteriaceae and an enigmatic plant pathogen with a broad host range. Although P. ananatis strains can be aggressive on onion causing foliar necrosis and onion center rot, previous genomic analysis has shown that P. ananatis lacks the primary virulence secretion systems associated with other plant pathogens. We assessed a collection of fifty P. ananatis strains collected from Georgia over three decades to determine genetic factors that correlated with onion pathogenic potential. Previous genetic analysis studies have compared strains isolated from different hosts with varying diseases potential and isolation sources. Strains varied greatly in their pathogenic potential and aggressiveness on different cultivated Allium species like onion, leek, shallot, and chive. Using multi-locus sequence analysis (MLSA) and repetitive extragenic palindrome repeat (rep)-PCR techniques, we did not observe any correlation between onion pathogenic potential and genetic diversity among strains. Whole genome sequencing and pan-genomic analysis of a sub-set of 10 strains aided in the identification of a novel series of genetic regions, likely plasmid borne, and correlating with onion pathogenicity observed on single contigs of the genetic assemblies. We named these loci Onion Virulence Regions (OVR) A-D. The OVR loci contain genes involved in redox regulation as well as pectate lyase and rhamnogalacturonase genes. Previous studies have not identified distinct genetic loci or plasmids correlating with onion foliar pathogenicity or pathogenicity on a single host pathosystem. The lack of focus on a single host system for this phytopathgenic disease necessitates the pan-genomic analysis performed in this study.

We propose Pseudomonas coronafaciens sp. nov. as a new species in genus Pseudomonas, which is diverse from P. syringae. We also classified strains from onions which are responsible for yellow bud (YB) disease as P. coronafaciens. Sequencing of 16S rRNA gene and multi-locus sequence analysis (MLSA) of housekeeping genes (gyrB, rpoD, gltA and gap1 genes) for the P. syringae pv. coronafaciens strains along with other strains of P. syringae pathovars resulted in a distinct cluster separate from other P. syringae pathovars. Based on DNA-DNA relatedness, pathotype strain of P. syringae pv. coronafaciens (CFBP 2216PT) exhibited ≤35.5% similarity with the pathotype strains of P. syringae pv. syringae (CFBP 1392PT, 4702T) but exhibited ≥90.6% with the YB strains (YB 12-1, YB 12-4, YB 09-1). Also, the YB strains (YB 12-1, YB 12-4, YB 09-1) were able to infect only onion but not oat, rye and Italian ryegrass (common hosts for P. syrinage pv. coronafaciens). Contrastingly, P. syringae pv. coronafaciens strains (NCPPB 600PT, ATCC 19608, Pcf 83-300) produced typical halo blight symptoms on oat, rye and Italian rye grass but did not produce any symptoms on onion. These results provide evidence that P. syringae pv. coronafaciens should be elevated to a species level and the new YB strains may potentially be a novel pathovar of hereto proposed P. coronafaciens species.

The ability of seed-borne bacterial pathogens (Acidovorax citrulli, Clavibacter michiganensis subsp. michiganensis, Pseudomonas syringae pv. tomato, Xanthomonas euvesicatoria, and Pseudomonas syringae pv. glycinea) to infest seeds of host and non-host plants (watermelon, tomato, pepper, and soybean) and subsequent pathogen transmission to seedlings was investigated. A non-pathogenic, pigmented strain of Serratia marcescens was also included to assess a null-interacting situation with the same plant species. Flowers of host and non-host plants were inoculated with 1 × 10(6) colony forming units (CFUs)/flower for each bacterial species and allowed to develop into fruits or umbels (in case of onion). Seeds harvested from each host/non-host bacterial species combination were assayed for respective bacteria by plating on semi-selective media. Additionally, seedlots for each host/non-host bacterial species combination were also assayed for pathogen transmission by seedling grow-out (SGO) assays under greenhouse conditions. The mean percentage of seedlots infested with compatible and incompatible pathogens was 31.7 and 30.9% (by plating), respectively and they were not significantly different (P = 0.67). The percentage of seedlots infested with null-interacting bacterial species was 16.8% (by plating) and it was significantly lower than the infested lots generated with compatible and incompatible bacterial pathogens (P = 0.03). None of the seedlots with incompatible/null-interacting bacteria developed symptoms on seedlings; however, when seedlings were assayed for epiphytic bacterial presence, 19.5 and 9.4% of the lots were positive, respectively. These results indicate that the seeds of non-host plants can become infested with incompatible and null-interacting bacterial species through flower colonization and they can be transmitted via epiphytic colonization of seedlings. In addition, it was also observed that flowers and seeds of non-host plants can be colonized by compatible/incompatible/null-interacting bacteria to higher populations; however, the level of colonization differed significantly depending on the type of bacterial species used.