Risk Evaluation of Benzovindiflupyr Resistance of Verticillium dahliae Population in Maine.

Verticillium dahliae causes Verticillium wilt, resulting in significant losses to potato production. Benzovindiflupyr, a succinate dehydrogenase inhibitor, effectively controls V. dahliae. However, frequent applications of the chemical may expedite the development of fungicide resistance in the pathogen population. To evaluate the risk of benzovindiflupyr resistance, 38 V. dahliae strains were obtained from diseased potatoes in Maine. The sensitivity of the field population was determined based on effective concentration for 50% inhibition (EC50), which ranged from 0.07 to 11.28 µg ml-1 with a median of 1.08. Segregated clusters of EC50 values indicated that Maine V. dahliae populations have developed benzovindiflupyr resistance. By exposing conidia of V. dahliae to a high concentration of benzovindiflupyr, 18 benzovindiflupyr-resistant mutants were obtained. To examine their fitness, the mutants were continuously subculture-transferred for up to 10 generations. Mycelial growth, conidial production, competitiveness, pathogenicity, and cross resistance of the 10th generation mutants were examined. Results showed that 50% of the resistant mutants retained an adaptive level in mycelial growth, and 60% maintained conidial production similar to their parents. Pathogenicity did not change for any of the mutants. No cross resistance was detected between benzovindiflupyr and either azoxystrobin, boscalid, fluopyram, or pyrimethanil. Thus, the resistance risk in V. dahliae to benzovindiflupyr should be considered in Maine potato production.

Species of Dickeya and Pectobacterium Isolated during an Outbreak of Blackleg and Soft Rot of Potato in Northeastern and North Central United States.

An outbreak of bacterial soft rot and blackleg of potato has occurred since 2014 with the epicenter being in the northeastern region of the United States. Multiple species of Pectobacterium and Dickeya are causal agents, resulting in losses to commercial and seed potato production over the past decade in the Northeastern and North Central United States. To clarify the pathogen present at the outset of the epidemic in 2015 and 2016, a phylogenetic study was made of 121 pectolytic soft rot bacteria isolated from symptomatic potato; also included were 27 type strains of Dickeya and Pectobacterium species, and 47 historic reference strains. Phylogenetic trees constructed based on multilocus sequence alignments of concatenated dnaJ, dnaX and gyrB fragments revealed the epidemic isolates to cluster with type strains of D. chrysanthemi, D. dianthicola, D. dadantii, P. atrosepticum, P. brasiliense, P. carotovorum, P. parmentieri, P. polaris, P. punjabense, and P. versatile. Genetic diversity within D. dianthicola strains was low, with one sequence type (ST1) identified in 17 of 19 strains. Pectobacterium parmentieri was more diverse, with ten sequence types detected among 37 of the 2015-2016 strains. This study can aid in monitoring future shifts in potato soft rot pathogens within the U.S. and inform strategies for disease management.

Pangenomic Analysis of Dickeya dianthicola Strains Related to the Outbreak of Blackleg and Soft Rot of Potato in the United States.

Dickeya dianthicola has caused an outbreak of blackleg and soft rot of potato in the eastern half of the United States since 2015. To investigate genetic diversity of the pathogen, a comparative analysis was conducted on genomes of D. dianthicola strains. Whole genomes of 16 strains from the United States outbreak were assembled and compared with 16 previously sequenced genomes of D. dianthicola isolated from potato or carnation. Among the 32 strains, eight distinct clades were distinguished based on phylogenomic analysis. The outbreak strains were grouped into three clades, with the majority of the strains in clade I. Clade I strains were unique and homogeneous, suggesting a recent incursion of this strain into potato production from alternative hosts or environmental sources. The pangenome of the 32 strains contained 6,693 genes, 3,377 of which were core genes. By screening primary protein subunits associated with virulence from all U.S. strains, we found that many virulence-related gene clusters, such as plant cell wall degrading enzyme genes, flagellar and chemotaxis related genes, two-component regulatory genes, and type I/II/III secretion system genes, were highly conserved but that type IV and type VI secretion system genes varied. The clade I strains encoded two clusters of type IV secretion systems, whereas the clade II and III strains encoded only one cluster. Clade I and II strains encoded one more VgrG/PAAR spike protein than did clade III. Thus, we predicted that the presence of additional virulence-related genes may have enabled the unique clade I strain to become predominant in the U.S. outbreak.

Interaction between Dickeya dianthicola and Pectobacterium parmentieri in Potato Infection under Field Conditions.

Dickeya and Pectobacterium spp. both cause blackleg and soft rot of potato, which can be a yield-reducing factor to potato production. The purpose of this study was to examine the interaction between these two bacterial genera causing potato infection, and subsequent disease development and yield responses under field conditions. Analysis of 883 potato samples collected in Northeastern USA using polymerase chain reaction determined that Dickeya dianthicola and P. parmentieri were found in 38.1% and 53.3% of all samples, respectively, and that 20.6% of samples contained both D. dianthicola and P. parmentieri. To further investigate the relationship between the two bacterial species and their interaction, field trials were established. Potato seed pieces of "Russet Burbank", "Lamoka", and "Atlantic" were inoculated with bacterial suspension of D. dianthicola at 107 colony-forming unite (CFU)/mL using a vacuum infiltration method, air dried, and then planted in the field. Two-year results showed that there was a high correlation (p < 0.01) between yield loss and percent of inoculated seed pieces. In a secondary field trial conducted in 2018 and 2019, seed pieces of potato "Shepody", "Lamoka" and "Atlantic" were inoculated with D. dianthicola, P. parmentieri, or mixture of both species, and then planted. In 2019, disease severity index, as measured by the most sensitive variety "Lamoka", was 16.2 with D. dianthicola inoculation, 10.4 with P. parmentieri, 25.4 with inoculation with both bacteria. Two-year data had a similar trend. Thus, D. dianthicola was more virulent than P. parmentieri, but the co-inoculation of the two species resulted in increased disease severity compared to single-species inoculation with either pathogen.

Genotyping Dickeya dianthicola Causing Potato Blackleg and Soft Rot Outbreak Associated With Inoculum Geography in the United States.

An outbreak of blackleg and soft rot of potato, caused primarily by the bacterial pathogen Dickeya dianthicola, has resulted in significant economic losses in the northeastern United States since 2015. The spread of this seedborne disease is highly associated with seed distribution; therefore, the pathogen likely spread with seed tubers. To describe the blackleg epidemic and track inoculum origins, a total of 1,183 potato samples were collected from 11 states associated with blackleg outbreak from 2015 to 2019. Of these samples, 39.8% tested positive for D. dianthicola. Seventeen isolates of D. dianthicola were recovered from these samples and the genetic diversity of these isolates was examined. Fingerprinting with BOX-A1R-based repetitive extragenic palindromic PCR and phylogenetic analysis based on sequences of the 16S rRNA and gapA genes indicated that D. dianthicola isolates were divided into three genotypes, denoted types I, II, and III. Ninety-five percent of samples from Maine were type I. Type II was found in Maine only in 2015 and 2018. Type II was present throughout the 5 years in some states at a lower percentage than type I. Type III was found in Pennsylvania, New Jersey, and Massachusetts, but not in Maine. Therefore, type I appears to be associated with Maine, but type II appeared to be distributed throughout the northeastern United States. The type II and rarer type III strains were closer to the D. dianthicola type strain isolated from the United Kingdom. This work provides evidence that the outbreak of blackleg of potato in the northeastern United States was caused by multiple strains of D. dianthicola. The geographic origins of these strains remain unknown.

Soil health paradigms and implications for disease management.

Soil health has been defined as the capacity of soil to function as a vital living system to sustain biological productivity, maintain environmental quality, and promote plant, animal, and human health. Building and maintaining soil health are essential to agricultural sustainability and ecosystem function. Management practices that promote soil health, including the use of crop rotations, cover crops and green manures, organic amendments, and conservation tillage, also have generally positive effects on the management of soilborne diseases through a number of potential mechanisms, including increasing soil microbial biomass, activity, and diversity, resulting in greater biological suppression of pathogens and diseases. However, there also may be particular disease issues associated with some soil health management practices. In this review, research and progress made over the past twenty years regarding soil health, sustainability, and soil health management practices, with an emphasis on their implications for and effects on plant disease and disease management strategies, are summarized.

Links among nitrification, nitrifier communities, and edaphic properties in contrasting soils receiving dairy slurry.

Soil biotic and abiotic factors strongly influence nitrogen (N) availability and increases in nitrification rates associated with the application of manure. In this study, we examine the effects of edaphic properties and a dairy (Bos taurus) slurry amendment on N availability, nitrification rates and nitrifier communities. Soils of variable texture and clay mineralogy were collected from six USDA-ARS research sites and incubated for 28 d with and without dairy slurry applied at a rate of ~300 kg N ha(-1). Periodically, subsamples were removed for analyses of 2 M KCl extractable N and nitrification potential, as well as gene copy numbers of ammonia-oxidizing bacteria (AOB) and archaea (AOA). Spearman coefficients for nitrification potentials and AOB copy number were positively correlated with total soil C, total soil N, cation exchange capacity, and clay mineralogy in treatments with and without slurry application. Our data show that the quantity and type of clay minerals present in a soil affect nitrifier populations, nitrification rates, and the release of inorganic N. Nitrogen mineralization, nitrification potentials, and edaphic properties were positively correlated with AOB gene copy numbers. On average, AOA gene copy numbers were an order of magnitude lower than those of AOB across the six soils and did not increase with slurry application. Our research suggests that the two nitrifier communities overlap but have different optimum environmental conditions for growth and activity that are partly determined by the interaction of manure-derived ammonium with soil properties.

Effects of different potato cropping system approaches and water management on soilborne diseases and soil microbial communities.

Four different potato cropping systems, designed to address specific management goals of soil conservation, soil improvement, disease suppression, and a status quo standard rotation control, were evaluated for their effects on soilborne diseases of potato and soil microbial community characteristics. The status quo system (SQ) consisted of barley underseeded with red clover followed by potato (2-year). The soil-conserving system (SC) featured an additional year of forage grass and reduced tillage (3-year, barley/timothy-timothy-potato). The soil-improving system (SI) added yearly compost amendments to the SC rotation, and the disease-suppressive system (DS) featured diverse crops with known disease-suppressive capability (3-year, mustard/rapeseed-sudangrass/rye-potato). Each system was also compared with a continuous potato control (PP) and evaluated under both irrigated and nonirrigated conditions. Data collected over three potato seasons following full rotation cycles demonstrated that all rotations reduced stem canker (10 to 50%) relative to PP. The SQ, SC, and DS systems reduced black scurf (18 to 58%) relative to PP; SI reduced scurf under nonirrigated but not irrigated conditions; and scurf was lower in DS than all other systems. The SQ, SC, and DS systems also reduced common scab (15 to 45%), and scab was lower in DS than all other systems. Irrigation increased black scurf and common scab but also resulted in higher yields for most rotations. SI produced the highest yields under nonirrigated conditions, and DS produced high yields and low disease under both irrigation regimes. Each cropping system resulted in distinctive changes in soil microbial community characteristics as represented by microbial populations, substrate utilization, and fatty acid methyl-ester (FAME) profiles. SI tended to increase soil moisture, microbial populations, and activity, as well result in higher proportions of monounsaturated FAMEs and the FAME biomarker for mycorrhizae (16:1 ω6c) relative to most other rotations. DS resulted in moderate microbial populations and activity but higher substrate richness and diversity in substrate utilization profiles. DS also resulted in relatively higher proportions of FAME biomarkers for fungi (18:2 ω6c), actinomycetes, and gram-positive bacteria than most other systems, whereas PP resulted in the lowest microbial populations and activity; substrate richness and diversity; proportions of monounsaturated and polyunsaturated FAME classes; and fungal, mycorrhizae, and actinomycete FAME biomarkers of all cropping systems. Overall, soil water, soil quality, and soilborne diseases were all important factors affecting productivity, and cropping systems addressing these constraints improved production. Cropping system approaches will need to balance these factors to achieve sustainable production and disease management.

Management of Verticillium Wilt of Potato with Disease-Suppressive Green Manures and as Affected by Previous Cropping History.

The ability of disease-suppressive rotation crops to reduce potato disease problems and increase crop productivity in a field with prior severe Verticillium wilt, as well as the potential influence of previous cropping history on disease suppression, was evaluated over three field seasons in Maine. Disease-suppressive rotations consisted of: (i) a high-glucosinolate mustard blend ('Caliente 119') as a mixture of white mustard (Sinapis alba) and oriental mustard (Brassica juncea) with known biofumigation potential and (ii) a sorghum-sudangrass hybrid. Each were grown as single-season green manures followed by a subsequent potato crop. These rotations were compared with a standard barley rotation and a barley rotation followed by chemical fumigation with metam sodium as controls. Both green manure rotations significantly reduced (average reductions of 25 and 18%, respectively) Verticillium wilt in the subsequent potato crop compared with the standard barley control but were not as effective as chemical fumigation (35% reduction). The mustard blend also reduced other soilborne diseases (black scurf and common scab) better than all other rotations. Mustard blend and chemical fumigation treatments increased tuber yield relative to the barley control by 12 and 18%, respectively. However, by the second rotation cycle, disease levels were high in all rotations, and only chemical fumigation resulted in substantial disease reduction (35%). Rotations also had significant effects on soil microbiology, including soil bacterial and fungal populations and microbial community characteristics based on fatty acid profiles. However, only chemical fumigation significantly reduced soil populations of Verticillium spp. and increased general soil microbial activity. Previous cropping history did not significantly affect disease reduction, tuber yield, or soil microbial communities. This research indicates the potential for using disease-suppressive rotations for managing Verticillium wilt and other soilborne diseases but also indicates that multiple years of disease-suppressive crops may be needed to substantially reduce disease in heavily infested fields.

Rotation and Cover Crop Effects on Soilborne Potato Diseases, Tuber Yield, and Soil Microbial Communities.

Seven different 2-year rotations, consisting of barley/clover, canola, green bean, millet/rapeseed, soybean, sweet corn, and potato, all followed by potato, were assessed over 10 years (1997-2006) in a long-term cropping system trial for their effects on the development of soilborne potato diseases, tuber yield, and soil microbial communities. These same rotations were also assessed with and without the addition of a fall cover crop of no-tilled winter rye (except for barley/clover, for which underseeded ryegrass was substituted for clover) over a 4-year period. Canola and rapeseed rotations consistently reduced the severity of Rhizoctonia canker, black scurf, and common scab (18 to 38% reduction), and canola rotations resulted in higher tuber yields than continuous potato or barley/clover (6.8 to 8.2% higher). Addition of the winter rye cover crop further reduced black scurf and common scab (average 12.5 and 7.2% reduction, respectively) across all rotations. The combined effect of a canola or rapeseed rotation and winter rye cover crop reduced disease severity by 35 to 41% for black scurf and 20 to 33% for common scab relative to continuous potato with no cover crop. Verticillium wilt became a prominent disease problem only after four full rotation cycles, with high disease levels in all plots; however, incidence was lowest in barley rotations. Barley/clover and rapeseed rotations resulted in the highest soil bacterial populations and microbial activity, and all rotations had distinct effects on soil microbial community characteristics. Addition of a cover crop also resulted in increases in bacterial populations and microbial activity and had significant effects on soil microbial characteristics, in addition to slightly improving tuber yield (4% increase). Thus, in addition to positive effects in reducing erosion and improving soil quality, effective crop rotations in conjunction with planting cover crops can provide improved control of soilborne diseases. However, this study also demonstrated limitations with 2-year rotations in general, because all rotations resulted in increasing levels of common scab and Verticillium wilt over time.

Effects of different 3-year cropping systems on soil microbial communities and rhizoctonia diseases of potato.

ABSTRACT Eight different 3-year cropping systems, consisting of soybean-canola, soybean-barley, sweet corn-canola, sweet corn-soybean, green bean-sweet corn, canola-sweet corn, barley-clover, and continuous potato (non-rotation control) followed by potato as the third crop in all systems, were established in replicated field plots with two rotation entry points in Presque Isle, ME, in 1998. Cropping system effects on soil microbial community characteristics based on culturable soil microbial populations, single carbon source substrate utilization (SU) profiles, and whole-soil fatty acid methyl ester (FAME) profiles were evaluated in association with the development of soilborne diseases of potato in the 2000 and 2001 field seasons. Soil populations of culturable bacteria and overall microbial activity were highest following barley, canola, and sweet corn crops, and lowest following continuous potato. The SU profiles derived from BIOLOG ECO plates indicated higher substrate richness and diversity and greater utilization of certain carbohydrates, carboxylic acids, and amino acids associated with barley, canola, and some sweet corn rotations, indicating distinct differences in functional attributes of microbial communities among cropping systems. Soil FAME profiles also demonstrated distinct differences among cropping systems in their relative composition of fatty acid types and classes, representing structural attributes of microbial communities. Fatty acids most responsible for differentiation among cropping systems included 12:0, 16:1 omega5c, 16:1 omega7c, 18:1 omega9c, and 18:2omega6c. Based on FAME biomarkers, barley rotations resulted in higher fungi-to-bacteria ratios, sweet corn resulted in greater mycorrhizae populations, and continuous potato produced the lowest amounts of these and other biomarker traits. Incidence and severity of stem and stolon canker and black scurf of potato, caused by Rhizoctonia solani, were reduced for most rotations relative to the continuous potato control. Potato crops following canola, barley, or sweet corn provided the lowest levels of Rhizoctonia disease and best tuber quality, whereas potato crops following clover or soybean resulted in disease problems in some years. Both rotation crop and cropping sequence were important in shaping the microbial characteristics, soilborne disease, and tuber qualities. Several microbial parameters, including microbial populations and SU and FAME profile characteristics, were correlated with potato disease or yield measurements in one or both harvest years. In this study, we have demonstrated distinctive effects of specific rotation crops and cropping sequences on microbial communities and have begun to relate the implications of these changes to crop health and productivity.

Identification and Characterization of Isolates of Phytophthora infestans Using Fatty Acid Methyl Ester (FAME) Profiles.

The utility of fatty acid profiles for characterization and differentiation of isolates of P. infestans was investigated. Two libraries of fatty acid methyl ester (FAME) profiles (one representing average genotype characteristics and one representing individual isolate characteristics) were established from at least eight replicate samples of each of 25 different isolates of P. infestans, including representative isolates of US-1, US-6, US-7, US-8, US-11, US-14, and US-17 genotypes. These libraries then were used to identify and characterize additional unknown isolates. Fatty acid profile characteristics also were compared with cultural and genetic characteristics of the isolates. FAME profiles for isolates of P. infestans were consistent over multiple extractions and distinctly different from profiles for isolates of other Phytophthora species, such as P. capsici and P. erythroseptica, as well as isolates of Pythium spp. and various other fungal groups. Overall, profiles from different isolates within the same genotype shared similar characteristics, although there was overlap among some genotypes. Incubation temperature, growth medium, and prolonged storage on agar media all significantly affected fatty acid profiles; however, when these conditions were kept constant, profiles were distinct, consistent, and reproducible over time. Isolate profiles were sufficiently specific that individual isolates could be distinctly identified by FAME profiles. In general, individual isolate characteristics were more determinant than genotype group characteristics, although genotype could be determined for most isolates tested. Results indicated that FAME profiles can be an additional tool useful for characterizing isolates and populations of P. infestans.

Effects of Varying Environmental Conditions on Biological Control of Fusarium Wilt of Tomato by Nonpathogenic Fusarium spp.

ABSTRACT The influence of varying environmental and cropping conditions including temperature, light, soil type, pathogen isolate and race, and cultivar of tomato on biological control of Fusarium wilt of tomato by isolates of nonpathogenic Fusarium oxysporum (CS-20 and CS-24) and F. solani (CS-1) was evaluated in greenhouse and growth chamber experiments. Liquid spore suspensions (10(6)/ml) of the biocontrol isolates were applied to soilless potting mix at the time of tomato seeding, and the seedlings were transplanted into pathogen-infested field soil 2 weeks later. Temperature regimes ranging from 22 to 32 degrees C significantly affected disease development and plant physiological parameters. Biocontrol isolate CS-20 significantly reduced disease at all temperature regimes tested, yielding reductions of disease incidence of 59 to 100% relative to pathogen control treatments. Isolates CS-24 and CS-1 reduced disease incidence in the greenhouse and at high temperatures, but were less effective at the optimum temperature for disease development (27 degrees C). Growing plants under shade (50% of full light) versus full light affected some plant growth parameters, but did not affect the efficacy of biocontrol of any of the three bio-control isolates. Isolate CS-20 effectively reduced disease incidence (56 to 79% reduction) in four different field soils varying in texture (sandy to clayey) and organic matter content (0 to 3.2%). Isolate CS-1 reduced disease in the sandy and loamy soils (49 to 66% reduction), but was not effective in a heavy clay soil. Both CS-1 and CS-20 were equally effective against all three races of the pathogen, as well as multiple isolates of each race (48 to 66% reduction in disease incidence). Both isolates, CS-1 and CS-20, were equally effective in reducing disease incidence (66 to 80% reduction) by pathogenic races 1, 2, and 3 on eight different tomato cultivars containing varying levels of inherent resistance to Fusarium wilt (susceptible, resistant to race 1, or resistant to races 1 and 2). These results demonstrate that both these Fusarium isolates, and particularly CS-20, can effectively reduce Fusarium wilt disease of tomato under a variety of environmental conditions and have potential for further development.

Efficacy of Various Fungal and Bacterial Biocontrol Organisms for Control of Fusarium Wilt of Tomato.

Numerous fungi and bacteria, including existing biocontrol strains with known activity against soilborne fungal pathogens as well as isolates collected from the roots and rhizosphere of tomato plants growing in the field, were tested for their efficacy in controlling Fusarium wilt of tomato. Tomato seedlings were treated with the potential biocontrol agents in the greenhouse and transplanted into pathogen-infested field soil. Organisms tested included nonpathogenic strains of Fusarium spp., Trichoderma spp., Gliocladium virens, Pseudomonas fluorescens, Burkholderia cepacia, and others. Specific nonpathogenic isolates of F. oxysporum and F. solani collected from a Fusarium wilt-suppressive soil were the most effective antagonists, providing significant and consistent disease control (50 to 80% reduction of disease incidence) in several repeated tests. These isolates also were equally effective in controlling Fusarium wilt diseases of other crops, including watermelon and muskmelon. Other organisms, including isolates of G. virens, T. hamatum, P. fluorescens, and B. cepacia, also significantly reduced Fusarium wilt compared to disease controls (30 to 65% reduction), but were not as consistently effective as the nonpathogenic Fusarium isolates. Commercially available biocontrol products containing G. virens and T. harzianum (SoilGard and RootShield, respectively) also effectively reduced disease (62 to 68% reduction) when granules were incorporated into potting medium at 0.2% (wt/vol). Several fungal and bacterial isolates collected from the roots and rhizosphere of tomato plants also significantly reduced Fusarium wilt of tomato, but were no more effective than other previously identified biocontrol strains. Combinations of antagonists, including multiple Fusarium isolates, Fusarium with bacteria, and Fusarium with other fungi, also reduced disease, but did not provide significantly better control than the nonpathogenic Fusarium antagonists alone.