Sensitivity of Meloidogyne incognita, Fusarium oxysporum f. sp. niveum, and Stagonosporopsis citrulli to Succinate Dehydrogenase Inhibitors Used for Control of Watermelon Diseases.

Watermelon is affected by diseases such as Fusarium wilt, gummy stem blight, and root-knot nematode (RKN). Succinate dehydrogenase inhibitors (SDHIs) with potential fungicide and nematicide activity provide the opportunity to control multiple diseases with one compound. In this study, we aimed to determine the sensitivity of Meloidogyne incognita race 4 (MI4), Fusarium oxysporum f. sp. niveum (FON), and Stagonosporopsis citrulli (SCIT) to existing SDHIs: benzovindiflupyr, fluopyram, cyclobutrifluram, and pydiflumetofen. All SDHIs had fungicidal activity against 19 SCIT isolates in mycelial growth assays, but isolates were most sensitive to pydiflumetofen (median EC50 = 0.41 µg/ml). Most of the 50 FON isolates tested were sensitive to cyclobutrifluram for mycelial growth (median EC50 = 4.04 µg/ml) and conidial germination (median EC50 = 0.2 µg/ml) assays but were not sensitive to fluopyram. MI4 was most sensitive to cyclobutrifluram for egg hatch (mean EC50 = 0.0019 µg/ml) and J2 motility (mean EC50 = 1.16 µg/ml) assays but was not sensitive to pydiflumetofen. Significant positive correlations between the sensitivity of SCIT (mycelial growth) and FON (mycelial growth and conidial germination) for cyclobutrifluram and benzovindiflupyr (SCIT r = 0.88; FON r = 0.7; P < 0.0001) and cyclobutrifluram and pydiflumetofen (SCIT r = 0.83; FON r = 0.67 and 0.77; P < 0.0001) indicate a potential for cross-resistance between these SDHIs for these fungal pathogens. Overall, results suggest that cyclobutrifluram may be used for managing RKN, whereas it should be used judiciously for Fusarium wilt of watermelon and gummy stem blight due to the existence of insensitive isolates to the fungicide.

Long-Read Sequencing Genome Assembly of Ceratocystis fimbriata Enables Development of Molecular Diagnostics for Sweetpotato Black Rot.

Ceratocystis fimbriata is a destructive fungal pathogen of sweetpotato (Ipomoea batatas) that leads to losses at all stages of sweetpotato production. Accurate detection of C. fimbriata would allow for more efficient deployment of management tactics in sweetpotato production. To develop a diagnostic assay, a hybrid genome assembly of C. fimbriata isolate AS236 was generated. The resulting 31.7-MB assembly was near-chromosome level, with 18 contigs, 6,481 predicted genes, and a BUSCO completion score of 98.4% when compared with the fungus-specific lineage database. Additional Illumina DNA reads from C. manginecans, C. platani, and a second C. fimbriata isolate (C1421) were then mapped to the assembled genome using BOWTIE2 and counted using HTSeq, which identified 148 genes present only within C. fimbriata as molecular diagnostic candidates; 6 single-copy and 35 highly multi-copy (>40 BLAST hits), as determined through a self-BLAST-P alignment. Primers for PCR were designed in the 200-bp flanking region of the first exon for each candidate, and the candidates were validated against a diverse DNA panel containing Ceratocystis species, sweetpotato pathogens, and plants. After validation, two diagnostic candidates amplified only C. fimbriata DNA and were considered to be highly specific to the species. These genetic markers will serve as valuable diagnostic tools with multiple applications including the detection of C. fimbriata in seed, soil, and wash water in sweetpotato production.

Phytophthora capsici: Recent Progress on Fundamental Biology and Disease Management 100 Years After Its Description.

Phytophthora capsici is a destructive oomycete pathogen of vegetable, ornamental, and tropical crops. First described by L.H. Leonian in 1922 as a pathogen of pepper in New Mexico, USA, P. capsici is now widespread in temperate and tropical countries alike. Phytophthora capsici is notorious for its capability to evade disease management strategies. High genetic diversity allows P. capsici populations to overcome fungicides and host resistance, the formation of oospores results in long-term persistence in soils, zoospore differentiation in the presence of water increases epidemic potential, and a broad host range maximizes economic losses and limits the effectiveness of crop rotation. The severity of disease caused by P. capsici and management challenges have led to numerous research efforts in the past 100 years. Here, we discuss recent findings regarding the biology, genetic diversity, disease management, fungicide resistance, host resistance, genomics, and effector biology of P. capsici.

Carboxylic Acid Amide but Not Quinone Outside Inhibitor Fungicide Resistance Mutations Show Clade-Specific Occurrence in Pseudoperonospora cubensis Causing Downy Mildew in Commercial and Wild Cucurbits.

Since its reemergence in 2004, Pseudoperonospora cubensis, the causal agent of cucurbit downy mildew (CDM), has experienced significant changes in fungicide sensitivity. Presently, frequent fungicide applications are required to control the disease in cucumber due to the loss of host resistance. Carboxylic acid amides (CAA) and quinone outside inhibitors (QoI) are two fungicide groups used to control foliar diseases in cucurbits, including CDM. Resistance to these fungicides is associated with single nucleotide polymorphism (SNP) mutations. In this study, we used population analyses to determine the occurrence of fungicide resistance mutations to CAA and QoI fungicides in host-adapted clade 1 and clade 2 P. cubensis isolates. Our results revealed that CAA-resistant genotypes occurred more prominently in clade 2 isolates, with more sensitive genotypes observed in clade 1 isolates, while QoI resistance was widespread across isolates from both clades. We also determined that wild cucurbits can serve as reservoirs for P. cubensis isolates containing fungicide resistance alleles. Finally, we report that the G1105W substitution associated with CAA resistance was more prominent within clade 2 P. cubensis isolates while the G1105V resistance substitution and sensitivity genotypes were more prominent in clade 1 isolates. Our findings of clade-specific occurrence of fungicide resistance mutations highlight the importance of understanding the population dynamics of P. cubensis clades by crop and region to design effective fungicide programs and establish accurate baseline sensitivity to active ingredients in P. cubensis populations.

Development, Validation, and Utility of Species-Specific Diagnostic Markers for Detection of Peronospora belbahrii.

Peronospora belbahrii is an oomycete and the cause of basil downy mildew, one of the most destructive diseases affecting basil production worldwide. Disease management is challenging due to wind-dispersed sporangia and contaminated seed; therefore, identifying P. belbahrii in seed lots before sale or planting or in the field before symptoms develop could allow for timely deployment of disease management strategies. In this study, a draft genome assembly and next-generation sequencing reads for P. belbahrii, as well as publicly available DNA-seq and RNA-seq reads of several other downy mildew pathogens, were incorporated into a bioinformatics pipeline to predict P. belbahrii-specific diagnostic markers. The specificity of each candidate marker was validated against a diverse DNA collection of P. belbahrii, host tissue, and related oomycetes using PCR. Two species-specific markers were identified and used as templates to develop a highly sensitive probe-based real-time quantitative PCR (qPCR) assay that could detect P. belbahrii in leaf tissue and seed samples. Both markers were capable of reliably detecting as low as 500 fg/µl of P. belbahrii genomic DNA and as few as 10 sporangia. The qPCR assay was then validated with seed samples collected from a basil cultivar experiment. In total, 48 seed samples were collected and tested; P. belbahrii was detected in samples of all cultivars at estimated concentrations of 600 fg/µl up to 250 pg/µl and at as few as 10 sporangia up to >1,000 sporangia. The markers and assays are valuable for diagnostics and identifying P. belbahrii-contaminated seed lots to mitigate the effects of future basil downy mildew epidemics.

Sweetpotato Root Development Influences Susceptibility to Black Rot Caused by the Fungal Pathogen Ceratocystis fimbriata.

Black rot of sweetpotato, caused by Ceratocystis fimbriata, is an important reemerging disease threatening sweetpotato production in the United States. This study assessed disease susceptibility of the storage root surface, storage root cambium, and slips (vine cuttings) of 48 sweetpotato cultivars, advanced breeding lines, and wild relative accessions. We also characterized the effect of storage root development on susceptibility to C. fimbriata. None of the cultivars examined at the storage root level were resistant, with most cultivars exhibiting similar levels of susceptibility. In storage roots, Jewel and Covington were the least susceptible and significantly different from White Bonita, the most susceptible cultivar. In the slip, significant differences in disease incidence were observed for above- and below-ground plant structures among cultivars, advanced breeding lines, and wild relative accessions. Burgundy and Ipomoea littoralis displayed less below-ground disease incidence compared with NASPOT 8, Sunnyside, and LSU-417, the most susceptible cultivars. Correlation of black rot susceptibility between storage roots and slips was not significant, suggesting that slip assays are not useful to predict resistance in storage roots. Immature, early-developing storage roots were comparatively more susceptible than older, fully developed storage roots. The high significant correlation between the storage root cross-section area and the cross-sectional lesion ratio suggests the presence of an unfavorable environment for C. fimbriata as the storage root develops. Incorporating applications of effective fungicides at transplanting and during early-storage root development when sweetpotato tissues are most susceptible to black rot infection may improve disease management efforts.

Clade-Specific Biosurveillance of Pseudoperonospora cubensis Using Spore Traps for Precision Disease Management of Cucurbit Downy Mildew.

Pseudoperonospora cubensis is an obligate oomycete and cause of cucurbit downy mildew (CDM), the most destructive foliar disease affecting cucurbit hosts. Annual epidemics develop throughout the United States as windborne sporangia travel great distances and survive prolonged exposure to solar radiation. Recent genomic evidence suggests that P. cubensis isolates display host adaptation based on their respective clade. Early detection is key for fungicide application timing, and identification of the host-adapted clade provides information on the risk of infection for specific cucurbit crops. In this study, a multiplex quantitative PCR assay was developed based on species- and clade-specific nuclear genomic markers. The assay detected as few as 10 sporangia or DNA at 100 fg/ml for both clades and was validated in the field by deploying rotorod spore samplers in cucurbit sentinel plots located at two research stations in North Carolina. Using this assay, sporangia DNA was detected in spore trap sampling rods before signs of P. cubensis or CDM symptoms were observed in the sentinel plots. Both clade 1 and clade 2 DNA were detected in late-season cucumber and watermelon plots but only clade 2 DNA was detected in the early-season cucumber plots. These results will significantly improve disease management of CDM by monitoring inoculum levels to determine the cucurbit crops at risk of infection throughout each growing season.

Population Analyses Reveal Two Host-Adapted Clades of Pseudoperonospora cubensis, the Causal Agent of Cucurbit Downy Mildew, on Commercial and Wild Cucurbits.

Pseudoperonospora cubensis, the causal agent of cucurbit downy mildew, is an airborne, obligate oomycete pathogen that re-emerged in 2004 and causes foliar disease and yield losses in all major cucurbit crops in the United States. Approximately 60 species in the family Cucurbitaceae have been reported as hosts of P. cubensis. Commercial hosts including cucumber, cantaloupe, pumpkin, squash, and watermelon are grown in North Carolina and many host species occur in the wild as weeds. Little is known about the contribution of wild cucurbits to the yearly epidemic; thus, this study aimed to determine the role of commercial and wild cucurbits in the structuring of P. cubensis populations in North Carolina, a region with high pathogen diversity. Ten microsatellite markers were used to analyze 385 isolates from six commercial and four wild cucurbits from three locations representing different growing regions across North Carolina. Population analyses revealed that wild and commercial cucurbits are hosts of P. cubensis in the United States, that host is the main factor structuring P. cubensis populations, and that P. cubensis has two distinct, host-adapted clades at the cucurbit species level, with clade 1 showing random mating and evidence of recombination and clade 2 showing nonrandom mating and no evidence of recombination. Our findings have implications for disease management because clade-specific factors such as host susceptibility and inoculum availability of each clade by region may influence P. cubensis outbreaks in different commercial cucurbits, timing of fungicide applications, and phenotyping for breeding efforts.

Assessing the Role of Temperature, Inoculum Density, and Wounding on Disease Progression of the Fungal Pathogen Ceratocystis fimbriata Causing Black Rot in Sweetpotato.

In 2014, Ceratocystis fimbriata, causal agent of black rot in sweetpotato, reemerged and inflicted large financial losses on growers in the United States. Black rot continues to damage sweetpotatoes and has become a priority to the industry since then. In contrast, little is known about the biology of C. fimbriata and the epidemiology of sweetpotato black rot. In this study, effects of environmental factors such as inoculum density, RH, and temperature on sweetpotato black rot were determined. Cured sweetpotatoes were wounded with a toothpick to simulate puncture wounds, inoculated with different spore suspensions (inoculum density) (104, 105, or 106 spores/ml), and incubated under different RH (85.53, 94.09, or 97.01%) and temperature (13, 18, 23, 29, or 35°C) for 21 days. In a separate experiment, five root wounding types (cuts, punctures, abrasions, end breaks, and macerating bruises) were compared. All wounded roots were subsequently soaked in a 103 spores/ml suspension and incubated at 100% RH and 23°C for 21 days. This study found 29 and 23°C to be the optimal temperature for black rot disease development and sporulation, respectively. No pathogen growth was observed at 13 and 35°C. Increased inoculum density significantly (P < 0.0001) increased disease incidence, but increasing RH had an effect only on sporulation area. All wound types resulted in increased disease incidence and sporulation as early as 7 days postinoculation. Our results highlight the importance of characterizing factors that affect disease development for achieving successful disease management strategies. Findings from this study will be used to improve disease management for sweetpotato black rot by suggesting tighter regulation of curing and storage conditions and better postharvest handling of sweetpotato roots to avoid unnecessary wounding.

Genome Sequencing and Transcriptome Analysis of the Hop Downy Mildew Pathogen Pseudoperonospora humuli Reveal Species-Specific Genes for Molecular Detection.

Pseudoperonospora humuli is an obligate oomycete pathogen of hop (Humulus lupulus) that causes downy mildew, an important disease in most production regions in the Northern Hemisphere. The pathogen can cause a systemic infection in hop, overwinter in the root system, and infect propagation material. Substantial yield loss may occur owing to P. humuli infection of strobiles (seed cones), shoots, and cone-bearing branches. Fungicide application and cultural practices are the primary methods to manage hop downy mildew. However, effective, sustainable, and cost-effective management of downy mildew can be improved by developing early detection systems to inform on disease risk and timely fungicide application. However, no species-specific diagnostic assays or genomic resources are available for P. humuli. The genome of the P. humuli OR502AA isolate was partially sequenced using Illumina technology and assembled with ABySS. The assembly had a minimum scaffold length of 500 bp and an N50 (median scaffold length of the assembled genome) of 19.2 kbp. A total number of 18,656 genes were identified using MAKER standard gene predictions. Additionally, transcriptome assemblies were generated using RNA-seq and Trinity for seven additional P. humuli isolates. Bioinformatics analyses of next generation sequencing reads of P. humuli and P. cubensis (a closely related sister species) identified 242 candidate species-specific P. humuli genes that could be used as diagnostic molecular markers. These candidate genes were validated using polymerase chain reaction against a diverse collection of isolates from P. humuli, P. cubensis, and other oomycetes. Overall, four diagnostic markers were found to be uniquely present in P. humuli. These candidate markers identified through comparative genomics can be used for pathogen diagnostics in propagation material, such as rhizomes and vegetative cuttings, or adapted for biosurveillance of airborne sporangia, an important source of inoculum in hop downy mildew epidemics.

Analysis of microsatellites from transcriptome sequences of Phytophthora capsici and applications for population studies.

Phytophthora capsici is a devastating oomycete that affects solanaceous, cucurbitaceous, fabaceous, and other crops in the United States (US) and worldwide. The release of the P. capsici genome allows for design of robust markers for genetic studies. We identified and characterized microsatellites in the P. capsici transcriptome. A subset of 50 microsatellites were assayed in a diverse set of P. capsici isolates and evaluated for polymorphism. Polymorphic microsatellites were confirmed by fragment analysis, and 12 were used for population characterization of 50 P. capsici isolates from different states, hosts, and mating types. Analysis of genetic relationship among isolates revealed significant geographic structure by state. Our findings highlight the usefulness of these 12 microsatellites to characterize the population structure of P. capsici and potential transferability to closely-related Phytophthora spp. since markers are located in coding regions. Our markers will facilitate genetic characterization and complement phenotypic studies of P. capsici populations, which may assist in deployment of disease management strategies.

Genetic Diversity, Fungicide Sensitivity, and Host Resistance to Ceratocystis fimbriata Infecting Sweetpotato in North Carolina.

Black rot of sweetpotato, caused by Ceratocystis fimbriata, has recently reemerged as a significant threat to sweetpotato production in North Carolina and other states across the United States. This disease has historically been controlled largely through cultural management strategies and, in some cases, fungicide application. The sudden and destructive reemergence of this disease in 2015 created the need for rapidly evaluating disease control strategies. Genetic diversity of current C. fimbriata isolates infecting sweetpotato in North Carolina was assessed using ITS, TEF, and MAT-2 sequences. All 50 tested isolates were confirmed to be of a single mating type, MAT-2, based on PCR amplification. Alignment of ITS, TEF, and MAT-2 sequences revealed all isolates were identical at each locus. Fourteen common sweetpotato cultivars and advanced breeding lines were screened for black rot resistance using two isolates. None of the cultivars were completely resistant to the disease and most were equally susceptible. 'Stokes Purple' and 'Covington' were the least susceptible, but significantly (P < 0.05) differed only from 'Bellevue', the most susceptible cultivar. Sensitivity of 50 C. fimbriata isolates to difenoconazole, fludioxonil, thiabendazole, dicloran, azoxystrobin, pyraclostrobin, fenamidone, and fluazinam was evaluated in vitro. Difenoconazole, thiabendazole, and fluazinam were most effective in reducing mycelia growth. Postharvest fungicide application on black rot-infected roots provided similar results. Low efficacy of dicloran, as well as a range of EC50 values among isolates, suggests potential resistance to this commonly applied fungicide. Results obtained in this study provide current and useful information so that improved recommendations can be made to reduce losses in sweetpotato to black rot.

Using Next-Generation Sequencing to Develop Molecular Diagnostics for Pseudoperonospora cubensis, the Cucurbit Downy Mildew Pathogen.

Advances in next-generation sequencing (NGS) allow for rapid development of genomics resources needed to generate molecular diagnostics assays for infectious agents. NGS approaches are particularly helpful for organisms that cannot be cultured, such as the downy mildew pathogens, a group of biotrophic obligate oomycetes that infect crops of economic importance. Unlike most downy mildew pathogens that are highly host-specific, Pseudoperonospora cubensis causes disease on a broad range of crops belonging to the family Cucurbitaceae. In this study, we identified candidate diagnostic markers for P. cubensis by comparing NGS data from a diverse panel of P. cubensis and P. humuli isolates, two very closely related oomycete species. P. cubensis isolates from diverse hosts and geographical regions in the United States were selected for sequencing to ensure that candidates were conserved in P. cubensis isolates infecting different cucurbit hosts. Genomic regions unique to and conserved in P. cubensis isolates were identified through bioinformatics. These candidate regions were then validated using PCR against a larger collection of isolates from P. cubensis, P. humuli, and other oomycetes. Overall seven diagnostic markers were found to be specific to P. cubensis. These markers could be used for pathogen diagnostics on infected tissue, or adapted for monitoring airborne inoculum with real-time PCR and spore traps.

Etiology and Epidemiological Conditions Promoting Fusarium Root Rot in Sweetpotato.

Sweetpotato production in the United States is limited by several postharvest diseases, and one of the most common is Fusarium root rot. Although Fusarium solani is believed to be the primary causal agent of disease, numerous other Fusarium spp. have been reported to infect sweetpotato. However, the diversity of Fusarium spp. infecting sweetpotato in North Carolina is unknown. In addition, the lack of labeled and effective fungicides for control of Fusarium root rot in sweetpotato creates the need for integrated strategies to control disease. Nonetheless, epidemiological factors that promote Fusarium root rot in sweetpotato remain unexplored. A survey of Fusarium spp. infecting sweetpotato in North Carolina identified six species contributing to disease, with F. solani as the primary causal agent. The effects of storage temperature (13, 18, 23, 29, and 35°C), relative humidity (80, 90, and 100%), and initial inoculum level (3-, 5-, and 7-mm-diameter mycelia plug) were examined for progression of Fusarium root rot caused by F. solani and F. proliferatum on 'Covington' sweetpotato. Fusarium root rot was significantly reduced (P < 0.05) at lower temperatures (13°C), low relative humidity levels (80%), and low initial inoculum levels for both pathogens. Sporulation of F. proliferatum was also reduced under the same conditions. Qualitative mycotoxin analysis of roots infected with one of five Fusarium spp. revealed the production of fumonisin B1 by F. proliferatum when infecting sweetpotato. This study is a step toward characterizing the etiology and epidemiology of Fusarium root rot in sweetpotato, which allows for improved disease management recommendations to limit postharvest losses to this disease.

Susceptibility of Maize to Stalk Rot Caused by Fusarium graminearum Deoxynivalenol and Zearalenone Mutants.

Fusarium graminearum is a destructive pathogen of cereals that can cause stalk rot in maize. Stalk rot results in yield losses due to impaired grain filling, premature senescence, and lodging, which limits production and harvesting of ears. In addition, mycotoxins can make infected tissues unfit for silage. Our objectives were to evaluate the natural variation in stalk rot resistance among maize inbreds, to establish whether deoxynivalenol (DON)- and zearalenone (ZEA)-deficient strains are pathogenic on a panel of diverse inbreds, and to quantify the accumulation of DON in infected stalk tissue. Wild-type F. graminearum and mycotoxin mutants (DON and ZEA) were used to separately inoculate stalks of 9-week-old plants of 20 inbreds in the greenhouse. Plants were evaluated for lesion area at the inoculation point at 0, 2, 14, and 28 days postinoculation and tissues around lesions were sampled to determine the DON content. Regardless of their ability to produce DON or ZEA, all tested F. graminearum strains caused stalk rot; however, significant differences in disease levels were detected. Among the tested inbreds, Mp717 was resistant to all three F. graminearum strains while Mp317 and HP301 were only partially resistant. Accumulation of DON was significantly lower in infected stalks of the resistant and partially resistant inbreds than the susceptible inbreds. Analysis of the 20 inbreds using data from 17 simple-sequence repeats revealed population structure among the individuals; however, there was no association between genetic clustering and stalk rot resistance. These findings are an additional step toward breeding maize inbreds suitable for planting in fields infested with F. graminearum.

Comparison Between CKD-EPI Creatinine and MDRD Equations to Estimate Glomerular Filtration Rate in Kidney Transplant Patients.

BACKGROUND: The Modification of Diet in Renal Disease (MDRD) equation is widely used to estimate glomerular filtration rate (eGFR) in kidney transplant (KT) patients. The novel Chronic Kidney Disease-Epidemiology Collaboration equation (CKD-EPI) could improve accuracy of GFR estimation. Our aim was to compare both equations for staging of CKD in KT patients. METHODS: In a cohort of KT patients, correlation of eGFR according to MDRD and CKD-EPI with 24-hour creatinine clearance (24h-CrCl) was evaluated. Concordance between both equations for CKD staging was performed. MDRD was used for initial CKD staging. The mean difference of GFR between 24h-CrCl and each equation was calculated and Bland-Altman analysis applied. RESULTS: A cohort of 463 KT patients were studied: 67% female, overall average age 46 ± 14 years, 41% living donor, mean time of transplantation 71 months (3-95), and mean serum creatinine 1.68 ± 1.03 mg/dL. For the whole cohort (all CKD stages), eGFR by CKD-EPI was 5.33 mL/min/1.73 m(2) higher than by MDRD (P < .01). For CKD stages 1, 2, and 3A, the mean eGFR differences (CKD-EPI - MDRD) were 13.98 ± 3.27, 8.2 ± 1.98, and 5.34 ± 1.32 mL/min/1,73 m(2), respectively. The percentage of patients with eGFR <60 mL/min/1.73 m(2) decreased from 63.8% according to MDRD to 53.9% with the use of CKD-EPI. In women and patients ≤65 years old, eGFR by CKD-EPI was 5.98 and 5.81 mL/min/1.73 m(2) higher, respectively, than by MDRD (P < .01). CONCLUSIONS: The novel CKD-EPI reduces the number of patients with eGFR <60 mL/min/1.73 m(2) and consequently assigns lower CKD stages to our KT population.

Regional and Temporal Population Structure of Pseudoperonospora cubensis in Michigan and Ontario.

Cucurbit downy mildew (CDM), caused by the oomycete pathogen Pseudoperonospora cubensis, is a devastating disease that affects cucurbit species worldwide. This obligate, wind-dispersed pathogen does not overwinter in Michigan or other northern regions and new isolates can enter the state throughout the growing season. To evaluate the regional and temporal population structure of P. cubensis, sporangia from CDM lesions were collected from cucurbit foliage grown in Michigan and Ontario field locations in 2011. Population structure and genetic diversity were assessed in 257 isolates using nine simple sequence repeat markers. Genetic diversity was high for isolates from Michigan and Canada (0.6627 and 0.6131, respectively). Five genetic clusters were detected and changes in population structure varied by site and sampling date within a growing season. The Michigan and Canada populations were significantly differentiated, and a unique genetic cluster was detected in Michigan.

Cultural, Chemical, and Alternative Control Strategies for Rhizopus Soft Rot of Sweetpotato.

Rhizopus soft rot, caused primarily by Rhizopus stolonifer, is one of the most common postharvest diseases of sweetpotato and is often considered the most devastating. Traditionally, Rhizopus soft rot has been effectively controlled using postharvest dips in dicloran fungicides; however, due to changes in market preferences, use of these fungicides is now limited. This, along with the lack of labeled and effective fungicides for control of Rhizopus soft rot in sweetpotato, creates the need for integrated strategies to control the disease. The effects of storage temperature (13, 23, and 29°C), relative humidity (80, 90, and 100%), and initial inoculum levels (3-, 5-, and 7-mm-diameter mycelial plugs) on progression of Rhizopus soft rot in 'Covington' sweetpotato were examined. Percent decay due to Rhizopus soft rot infection was significantly reduced (P < 0.0001) at a low temperature (13°C) but was not significantly affected by changes in relative humidity or initial inoculum level (P >0.05). Sporulation of R. stolonifer was also significantly reduced at the lowest temperature of 13°C. High relative humidity (>95%) significantly increased sporulation of R. stolonifer and sporulation also increased as initial inoculum level increased. Efficacy of chlorine dioxide (ClO2) fumigation, UV-C irradiation, and postharvest dips in alternative control products were also investigated for control of Rhizopus soft rot. Static ClO2 treatments were effective in reducing sporulation on treated roots but had no significant impact on incidence of Rhizopus soft rot. UV irradiation at 3.24 KJ/m2 1 h after inoculation as well as dips in aqueous ClO2 and StorOx 2.0 significantly (P < 0.05) reduced disease incidence. Understanding the epidemiological factors favoring Rhizopus soft rot and identifying alternative control strategies allow for improved recommendations to limit postharvest losses in sweetpotato.

Resistance to Crown and Root Rot Caused by Phytophthora capsici in a Tomato Advanced Backcross of Solanum habrochaites and Solanum lycopersicum.

Phytophthora capsici causes devastating disease on many vegetable crops, including tomato and other solanaceous species. Solanum habrochaites accession LA407, a wild relative of cultivated tomato, has shown complete resistance to four P. capsici isolates from Michigan cucurbitaceous and solanaceous crops in a previous study. Greenhouse experiments were conducted to evaluate 62 lines of a tomato inbred backcross population between LA407 and the cultivated tomato 'Hunt 100' and 'Peto 95-43' for resistance to two highly virulent P. capsici isolates. Roots of 6-week-old seedlings were inoculated with each of two P. capsici isolates and maintained in the greenhouse. Plants were evaluated for wilting and plant death three times per week for 5 weeks. Significant differences were observed in disease response among the inbred tomato lines. Most lines evaluated were susceptible to P. capsici isolate 12889 but resistant to isolate OP97; 24 tomato lines were resistant to both isolates. Heritability of Phytophthora root rot resistance was high in this population. Polymorphic molecular markers located in genes related to resistance and defense responses were identified and added to a genetic map previously generated for the population. Resistant lines and polymorphic markers identified in this study are a valuable resource for development of tomato varieties resistant to P. capsici.

Pharmacokinetics of an injectable long-acting parenteral formulation of doxycycline hyclate in pigs.

Based on its ideal PK/PD ratios, doxycycline hyclate (DOX-h), a time-dependant antibacterial, is ideally expected to achieve sustained plasma drug concentrations at or slightly above the MIC level for as long as possible between dosing intervals. Pursuing this end, a poloxamer-based matrix was used to produce a 10% long-acting injectable preparation (DOX-h-LA) and its serum concentrations vs. time profile investigated after its injection to pigs in the pericaudal s.c. by parallel design. Results were compared with the forced oral bolus dose and i.v. pharmacokinetics of DOX-h. For this study, 12 recently weaned pigs per group were included in this trial, and a dose of 20 mg/kg was injected in all cases. DOX-h-LA showed the greatest values for bioavailability (115.38%); maximum serum concentration (Cmax) value was 1.5 ± 0.2 with a time to reach Cmax of 3.41 ± 0.04 h and an elimination rate constant of 70.93 ± 0.87( ) h. Considering minimum effective serum concentration of 0.5 µg/mL, a dose interval of at least 5 days can be achieved for DOX-h-LA, whereas p.o. and i.v. dosing of DOX-h may only last 11 and 15 h, respectively. Pigs were slaughtered on day 30 after this trial, and no visible remnants of the preparation were detected neither fibrosis was observed after a thorough macroscopic and histopathological analysis.