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Bacteria use selective membrane transporting strategies to support cell survival in different environments. Of the membrane transport systems, ATP-binding cassette (ABC) transporters, which utilize the energy of ATP hydrolysis to deliver substrate across the cytoplasmic membrane, are the largest and most diverse superfamily. These transporters import nutrients, export molecules, and are required for diverse cell functions, including cell division and morphology, gene regulation, surface motility, chemotaxis, and interspecies competition. Phytobacterial pathogens encode numerous ABC transporter homologs compared with related nonphytopathogens, with up to 160 transporters per genome, suggesting that plant pathogens must be able to import or respond to a greater number of molecules compared with saprophytes or animal pathogens. Despite their importance, ABC transporters have been little examined in plant pathogens. To understand bacterial phytopathogenesis and evolution, we need to understand the roles that ABC transporters play in plant-microbe interactions. In this review, we outline a multitude of roles that bacterial ABC transporters play, using both plant and animal pathogens as examples, to emphasize the importance of exploring these transporters in phytobacteriology. [ABSTRACT FROM AUTHOR]

Potato virus Y (PVY) causes one of the most serious and widespread diseases in North America. In recent years, the virus has become increasingly difficult to control. Durable dominant genes for resistance to PVY exist in potato germplasm and provide an effective control strategy. This paper describes a Solanum chacoense clone (M19) that is homozygous for a PVY resistance gene. The gene is linked to a previously published marker for Rychc found in Solanum chacoense. M19 was crossed with a diploid S. tuberosum clone to produce an adapted clone carrying the resistance gene. This hybrid clone is named M20. M20 tuberizes in the field, producing round tubers with white skin and flesh and moderate size. M20 is resistant to PVYO, PVYN:O, and PVYNTN. Both M19 and M20 are female and male fertile, so they are being released as sources of PVY resistance for breeding programs. [ABSTRACT FROM AUTHOR]

The third most important food crop worldwide, potato (Solanum tuberosum L.) is a tetraploid outcrossing species propagated from tubers. Breeders have long been challenged by polyploidy, heterozygosity, and asexual reproduction. It has been assumed that tetraploidy is essential for high yield, that the creation of inbred potato is not feasible, and that propagation by seed tubers is ideal. In this paper, we question those assumptions and propose to convert potato into a diploid inbred line-based crop propagated by true seed. Although a conversion of this magnitude is unprecedented, the possible genetic gains from a breeding system based on inbred lines and the seed production benefits from a sexual propagation system are too large to ignore. We call on leaders of public and private organizations to come together to explore the feasibility of this radical and exciting new strategy in potato breeding. [ABSTRACT FROM AUTHOR]

A new leaf spot disease was observed on leaves of Rheum palmatum (Chinese rhubarb) in Northwest China (Gansu Province) starting in 2005. A Septoria-like fungus was isolated and completion of Koch's postulates confirmed that the fungus was the casual agent of the leaf spot disease. Morphology and molecular methods were combined to identify the pathogen. The fungus produced conidiomata pycnidia and the conidia were 2~5 septate, 61.2~134.1 μm in length and 3.53~5.3 μm in width, which is much larger than the known Spetoria species that infects Polygonaceae species. Phylogenic analysis of the internal transcribed spacer region confirmed that this Septoria-like fungus is within the Septoria genus but distinct from known Septoria species. Together, these morphological and phylogenetic data support that the R. palmatum infecting Septoria strain is a newlydescribed plant pathogenic species. [ABSTRACT FROM AUTHOR]

Pectobacterium species cause soft rot, blackleg, and stem rot in potato and a wide range of other vegetable crops and ornamental plants. Diseases caused by Pectobacterium are controlled mainly through use of healthy planting material, sanitation and copper sprays. Environmental factors, such as temperature, moisture and soil oxygen concentration, have a large effect on development of diseases caused by Pectobacterium and disease incidence can be unpredictable. The pathogen is spread by various mechanisms including water, seed, equipment and insects. Little is understood about plant resistance to soft rot bacterial pathogens and no commercial potato cultivars are resistant to soft rot, although some have tolerance and some wild potato species are resistant. Pectobacterium is a diverse genus, with multiple species capable of infecting potato. Multiple Pectobacterium species may be found in the same field and even on the same plant. Pectobacterium strains vary in aggressiveness and the virulence genes they encode but there are many commonalities across the genus. Over the past decade, genomic studies have provided new insights into Pectobacterium biology. For example, some Pectobacterium strains may elicit plant cell death to promote disease in leaves. Strains of the pathogen also produce an orange pigment and volatile compounds that increase virulence and that may act as insect kairomones. Recent work with a supervised machine learning program has identified several novel target genes likely to contribute to plant-microbe interactions, suggesting that there is still much to learn about how soft rot bacteria cause disease. [ABSTRACT FROM AUTHOR]

Background A wealth of genome sequences has provided thousands of genes of unknown function, but identification of functions for the large numbers of hypothetical genes in phytopathogens remains a challenge that impacts all research on plant-microbe interactions. Decades of research on the molecular basis of pathogenesis focused on a limited number of factors associated with long-known host-microbe interaction systems, providing limited direction into this challenge. Computational approaches to identify virulence genes often rely on two strategies: searching for sequence similarity to known host-microbe interaction factors from other organisms, and identifying islands of genes that discriminate between pathogens of one type and closely related non-pathogens or pathogens of a different type. The former is limited to known genes, excluding vast collections of genes of unknown function found in every genome. The latter lacks specificity, since many genes in genomic islands have little to do with host-interaction. Result In this study, we developed a supervised machine learning approach that was designed to recognize patterns from large and disparate data types, in order to identify candidate hostmicrobe interaction factors. The soft rot Enterobacteriaceae strains Dickeya dadantii 3937 and Pectobacterium carotovorum WPP14 were used for development of this tool, because these pathogens are important on multiple high value crops in agriculture worldwide and more genomic and functional data is available for the Enterobacteriaceae than any other microbial family. Our approach achieved greater than 90% precision and a recall rate over 80% in 10-fold cross validation tests. Application of the learning scheme to the complete genome of these two organisms generated a list of roughly 200 candidates, many of which were previously not implicated in plant-microbe interaction and many of which are of completely unknown function. Conclusion These lists provide new targets for experimental validation and further characterization, and our approach presents a promising pattern-learning scheme that can be generalized to create a resource to study host-microbe interactions in other bacterial phytopathogens. [ABSTRACT FROM AUTHOR]

Soft-rot Enterobacteriaceae (SRE), which belong to the genera Pectobacterium and Dickeya, consist mainly of broad host-range pathogens that cause wilt, rot, and blackleg diseases on a wide range of plants. They are found in plants, insects, soil, and water in agricultural regions worldwide. SRE encode all six known protein secretion systems present in gram-negative bacteria, and these systems are involved in attacking host plants and competing bacteria. They also produce and detect multiple types of small molecules to coordinate pathogenesis, modify the plant environment, attack competing microbes, and perhaps to attract insect vectors. This review integrates new information about the role protein secretion and detection and production of ions and small molecules play in soft-rot pathogenicity. [ABSTRACT FROM AUTHOR]

Soft rot pathogens cause significant losses worldwide in fruit and vegetable production during the growing season and after harvest. These pathogens engage in molecular interactions with their plant hosts via acyl-homoserine lactone (AHL) quorum sensing, pectin metabolites, phenolic acids, and auxin. The AHL-based interactions can be disrupted enzymatically or with chemical inhibitors, which in some cases, leads to disease control. Since the wisdom of disrupting AHL-based quorum sensing for control of an agricultural problem is questionable when this may be a useful target for control of human pathogens, development of other control mechanisms more specific to plant pathogens would be beneficial. The recent identification of plant-specific compounds used in plant–bacterial interactions, such as auxin, o-coumaric acid, t-cinnamaic acid, as well as analogs that inhibit pathways induced by these compounds, may lead to novel control mechanisms aimed specifically at managing plant-infecting bacteria. [Copyright &y& Elsevier]

Abstract: RNA integrity is critical for successful RNA quantitation for mammalian tissues, but the level of integrity required differs among tissues. The level of integrity required for quantitation has not been determined for bacterial RNA. Three RNA isolation methods were evaluated for their ability to produce high quality RNA from Dickeya dadantii, a bacterium refractory to RNA isolation. Bacterial lysis with Trizol using standard protocols consistently gave low RNA yields with this organism. Higher yields due to improved bacterial cells lysis was achieved with an added hot SDS incubation step, but RNA quality was low as determined by the RNA Integrity Number (RIN). Contaminating DNA remained a problem with the hot SDS-Trizol method; RNA samples required repeated, rigorous DNase treatments to reduce DNA contamination to levels sufficient for successful real-time qRT-PCR. A hot SDS-hot phenol RNA method gave the highest RNA quality and required only two DNase treatments to remove DNA. The assessment of RNA integrity using the Agilent 2100 BioAnalyzer was critical for obtaining meaningful gene expression data. RIN values below 7.0 resulted in high variation and loss of statistical significance when gene expression was analyzed by real-time qRT-PCR. We found that RNA preparations of different quality yielded drastic differences in relative gene expression ratios and led to major errors in the quantification of transcript levels. This work provides guidelines for RNA isolation and quality assessment that will be valuable for gene expression studies in a wide range of bacteria. [Copyright &y& Elsevier]

Presents information on a study which focused on the characteristics of the plant pathogenic bacterium Pseudomonas syringae Hrp pathogenicity island. Plant bioassays; Materials and methods of the study; Results and discussion.

Focuses on a study on the role that hypersensitive reaction and pathogenicity (hrp) genes play in interactions of Pseudomonas syringae pv. syringae with its host plant as they occur naturally under field conditions. Methodology of the study; Results and discussion; Conclusion.

Presents information on an experiment which concluded that genes in the hrpC (hypersensitive response and pathogenicity) operons are necessary for translocation across the outer membrane, and that the Pss61 Hrp system permits the study of two genetically distinguishable stages in type III secretions. Discussion of the results that permitted this conclusion; Methodology used to conduct the study.

In the battle between bacteria and plants, bacteria often use a population density–dependent regulatory system known as quorum sensing (QS) to coordinate virulence gene expression. In response, plants use innate and induced defense mechanisms that include low-molecular-weight compounds, some of which serve as antivirulence agents by interfering with the QS machinery. The best-characterized QS system is driven by the autoinducer N-acyl-homoserine lactone (AHL), which is produced by AHL synthases (LuxI homologs) and perceived by response regulators (LuxR homologs). Several plant compounds have been shown to directly inhibit LuxI or LuxR. Gaining atomic-level insight into their mode of action and how they interfere with QS enzymes supports the identification and design of novel QS inhibitors.Such information can be gained by combining experimental work with molecular modeling and docking simulations. The summary of these findings shows that plant-derived compounds act as interkingdom cues and that these allomones specifically target bacterial communication systems. [ABSTRACT FROM AUTHOR]

Seed potato certification programs aim to limit disease incidence in planting material to levels below a threshold that causes significant losses to seed potato buyers. Records maintained for seed potato certification can be analyzed for trends in seed potato varietal mixture and disease incidences over time. We used logistic regression models to determine effects of year, potato variety, and their interaction on the incidences of potato diseases and disorders based on seed potato certification data collected in Colorado from 2012 to 2016. The effect of seed generation of important varieties on the incidence of common potato diseases was also quantified. Among the documented diseases, mosaic, caused primarily by Potato virus Y, is a persistent problem leading to high percentage of seed lot rejections in both summer and winter inspections, while blackleg, caused by Dickeya spp. and Pectobacterium spp., was a common disease causing summer rejections. The model demonstrated year, potato variety, and their interaction were key factors contributing to incidence of varietal mixture, or one or more potato diseases and disorders. For summer inspections, our models identified 53, 42, and six varieties sensitive to mosaic, blackleg, and leafroll, respectively. There were 17, 15, and six varieties that were tolerant to mosaic, blackleg, and leafroll regardless of pathogen pressures in the environment. For winter inspections, 51 varieties were sensitive to mosaic and three to leafroll, whereas 45 and one were relatively tolerant to mosaic and leafroll. The pattern of seed generation effects of selected potato varieties on mosaic and blackleg incidence was inconsistent across inspection years. In addition, we observed a significant negative correlation between the relative abundance of the green peach aphid, Myzus persicae, and mosaic in winter inspections. Mosaic incidences in summer and leafroll incidences were not influenced by common aphid species captured or total aphids in the valley. These results identify mosaic and blackleg as major causes of seed potato rejections and downgrades, sensitive and tolerant varieties, and provide suggestions for improving integrated crop management practices in Colorado. [ABSTRACT FROM AUTHOR]

Potato common scab is a widespread disease in which scab-like lesions develop on tubers. The disease is caused by pathogenic Streptomyces species, which synthesize the phytotoxin thaxtomin. The txtAB operon, responsible for thaxtomin production, can be used as a marker to identify pathogenic strains of the bacterium. Screening methods to assess scab susceptibility in breeding programs are time-consuming and can produce variable results. Management practices to control the disease vary and include crop rotation, tolerant varieties, monitoring soil pH, avoiding low soil moisture at tuber initiation, and application of soil- and/or seed-applied pesticides. There is a wide range in levels of tolerance among potato varieties. Many public research programs are committed to breeding for scab-tolerant varieties and evaluating management methods. Topics reviewed target readers focused on breeding and disease management objectives to reduce the incidence and severity of potato common scab. [ABSTRACT FROM AUTHOR]

In response to increasing interest in diploid potato (Solanum tuberosum) breeding and the production of diploid inbred hybrid potato varieties, the Breeding and Genetics section of the Potato Association of America (PAA) organized a symposium on diploid breeding that took place during the 2021 PAA annual meeting. Proceedings from that symposium are documented in this manuscript. Speakers from academia, government and industry presented their unique perspectives. Presentations covered a wide range of topics. Potential advantages of diploid breeding were introduced, and reasons to be skeptical about diploid breeding were highlighted. The impact that diploid breeding might have on the potato seed industry was discussed. Advantages for genetics research were emphasized. Aspects of tomato breeding and production were reviewed and considered as potential models for diploid potato breeding and production activities. Lastly, an industry-centered view of diploid potato breeding was provided. Taken together, these presentations are a snapshot of how diploid potato breeding was viewed in the moment, a vision for how diploid breeding might be implemented, and a thoughtful reflection on how diploid breeding and inbred hybrid varieties might change the potato variety development process and impact the potato industry. [ABSTRACT FROM AUTHOR]

Labor and equipment costs are the main expenses in potato micropropagation. To determine if we could reduce costs associated with media sterilization, a disinfectant, sodium hypochlorite (NaOCl), in combination with microwave heating, were assayed as media sterilants. Incorporating a common 5 % NaOCl household beach at a concentration of 9 ppm (active chlorine) in media sterilized with an autoclave or microwave oven controlled microorganism growth and maintained plantlet growth performance. Non-sterile 473 ml (16 oz.) clear deli containers were selected as an inexpensive replacement for traditional culture vessels and were effectively sterilized with a 50 ppm (active chlorine) NaOCl solution. Reuse of the non-sterile clear deli containers and alternate media water sources were also tested but this decreased plantlet growth performance. Comparison of a controlled growth chamber and ambient laboratory conditions was also investigated. Microorganism growth was significantly less in a controlled growth chamber (5 %) as compared to uncontrolled conditions (26-36 %). [ABSTRACT FROM AUTHOR]

Helminthosporium solani is a slow-growing fungal pathogen belonging to the family Massarinaceae. It causes blemishes on potato tubers, affecting processing and fresh market trade. Despite its worldwide distribution, little is known about the biology of H. solani. Here we report the generation of a draft genome sequence of H. solani with an estimated genome size of ~35 megabases, for use in phylogenetic and pathogenicity studies of this fungal pathogen. This sequence is also the first reference genome within the family Massarinaceae. We identified a large suite of genes in the H. solani genome that encode putative cell wall degrading enzymes. Based on comparison with other genomes, we speculate that H. solani is a hemibiotroph or necrotroph. The presence of a large number of genes in the glycoside hydrolase (GH) 10 and 43 families, which aid in the hydrolysis of glucoronoarabinoxylan, also suggests that H. solani may be able to survive on grass hosts and indicates the need to re-examine the life cycle and host range of this pathogen. [ABSTRACT FROM AUTHOR]

The article focuses on the factors influencing the sustainability of the Wisconsin Seed Potato Certification Program (WSPCP) and examines the changes in disease control with changing pathogen epidemiology. It presents the factors responsible for long term stability of these programs which include invention of tractors, synthetic fertilizers, and pesticides. It informs that WSPCP was founded with two aims which include reducing pathogen incidence and providing variety of seeds.

Vegetative propagation of potato leads to virus accumulation, resulting in significant yield losses and reduced quality. Virus identification is critical for developing disease management strategies and measuring seed lot health. The most widely used method of virus diagnosis in seed potatoes is a post-harvest test, for which the enzyme-linked immunosorbent assay (ELISA) is often used. ELISA was previously modified by substituting microtiter plates with membranes to develop a more flexible and inexpensive assay. We optimized a dot-blot immunoassay with viral proteins bound to a polyvinylidene fluoride (PVDF) membrane and detection of the proteins with alkaline phosphatase-labeled antibodies and a chemiluminescence reagent. The assay was tested for detection of viruses of seven genera. We have also altered the assay by spotting an antibody array onto a PVDF membrane and tested it for its potential uses as a diagnostic tool for Potato virus Y, Hosta virus X, and Potato leafroll virus. [ABSTRACT FROM AUTHOR]

Silver scurf, caused by Helminthosporium solani, and black dot, caused by Colletotrichum coccodes, cause tuber blemishes on potato ( Solanum tuberosum L.) which affect processing and fresh market trade. Tubers from ten cultivars were collected at harvest from three organic farms in Wisconsin and categorized as symptomatic or asymptomatic based on visual symptoms of silver scurf and black dot and/or signs of H. solani and C. coccodes. Tuber incubation and PCR assays were performed on asymptomatic tubers to detect H. solani and C. coccodes. Tuber incubation and PCR assays were in slight to fair agreement (kappa coefficient <0.4) for detecting both pathogens. Most asymptomatic tubers tested were positive by one or both assays for H. solani (75 %) or C. coccodes (94 %). Minituber inoculation assays were also performed to screen potato lines for resistance to silver scurf. Of the 14 lines tested, a diploid interspecific hybrid, C287, had consistently low sporulation, suggesting it has partial resistance to silver scurf. Since the majority of tubers harvested are already infected with one or both pathogens further research should focus on organically acceptable management practices that may inhibit disease development in field and in storage. [ABSTRACT FROM AUTHOR]

Pectobacteriu,n species cause disease worldwide in many crop and ornamental plants, including potato. A new Pectobacteriu,n subspecies, P carotovorum subsp. brasiliensis was recently described in Brazil and later found in the United States, Israel, and South Africa. Its virulence traits and host range remain unknown. A comparison of three taxa commonly found on potato showed that both P carotovorum subsp. carotovorum and subsp. brasiliensis are more aggressive in causing tuber and stem soft rot than P atrosepticum. Also, despite bacterial growth inhibition in vitro of P carotovorum subsp. carotovorum and P atrosepticum strains by P carotovorum subsp. brasiliensis, this new subspecies and P carorovorum subsp. carotovorurn are able to co-colonize in the same infected tissue. Both subspecies were motile in lesions. Pathogenesis assays showed that host ranges of all three overlap, but are not identical. The host ranges of individual strains of P carobvorum subsp. carotovorum and subsp. brasiliensis are limited, whereas P atrosepticum can macerate many plant species in addition to potato. There was high variability in virulence assays with potato tuber; thus physiological factors were investigated. Tuber size, maturity, and field location had significant effects on susceptibility to soft rot, with larger, more mature tubers being more susceptible. [ABSTRACT FROM AUTHOR]

Despite rapid advances on certain aspects of plant pathogenic bacteria, many economically important pathosystems are largely unexplored and biologically relevant life stages of even familiar systems remain poorly understood. We know remarkably little about end-stage disease, latent infections, survival away from the host, interactions among multiple microbes in a plant, and the effects of quantitative virulence factors. While no thoughtful researcher would dispute the effectiveness of reductionist experiments, we propose that this approach be combined with a broader perspective that includes the ecology, histopathology, and community population biology of phytopathogenic bacteria. We offer examples of exciting recent discoveries resulting from this natural history-based approach. In particular, in situ studies using biologically realistic inoculation followed by analyses with microscopy, gene expression profiling, community analyses, or application of key computational tools can offer new insights into old questions. Research that combines cutting-edge tools with a biological perspective is especially lacking on high-impact diseases of subsistence crops. Understanding the biology underlying important practical issues such as copper resistance, eradication from seed and cuttings, and rapid, sensitive detection could be of significant utility. Overall, we endorse a broader biological approach to research on plant pathogenic bacteria. [ABSTRACT FROM AUTHOR]

A rapid, high-temperature method of sample preparation from gram-negative bacteria for contour-clamped homogenous electric field (CHEF) electrophoresis is described, which utilizes a diethylpyrocarbonate nuclease inactivation step for preventing latent degradation of DNA. Also described is a simple 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid buffer system that permits high-voltage CHEF electrophoresis in standard apparatus while preventing the strand scission events associated with tris(hydroxymethyl)aminomethane buffers. Finally, the use of NuSieve agarose in combination with standard analytical agarose was compared to specialty CHEF agarose, demonstrating the potential use of this reagent in CHEF gel separations. PRACTICAL APPLICATIONS Contour-clamped homogenous electric field (CHEF) gel electrophoresis is a widely used technique in epidemiological and comparative genetic studies of microorganisms. Standard techniques work variably well, making options that can be used to “tweak” methodology useful. In this article, we describe a rapid, higher-temperature variation of sample preparation that yields inherently stable DNA with a more efficient usage of expensive reagents. Also detailed is a simple electrophoresis buffer that eliminates the tris(hydroxymethyl)aminomethane-mediated strand scission events that have complicated the comparison of bacterial strains in some studies. This buffer permits the use of the higher-voltage electrophoresis conditions used in many studies without the addition of potentially harmful chemicals. Finally, we demonstrate that the combination of NuSieve (Lonza) branded agarose with standard agarose can be used effectively in CHEF gels, thus expanding analytical options while also providing an alternative to maintaining other expensive specialty agaroses. The methods herein may be applied in whole or in part, allowing their possible inclusion in existing or future protocols as desired. [ABSTRACT FROM AUTHOR]

We used genetic and biochemical methods to examine the genomic diversity of the enterobacterial plant pathogen Erwinia carotovora subsp, carotovora. The results obtained with each method showed that E. carotovora subsp, carotovora strains isolated from one ecological niche, potato plants, are surprisingly diverse compared to related pathogens. A comparison of 23 partial mdh sequences revealed a maximum pairwise difference of 10.49% and an average pairwise difference of 2.13%, values which are much greater than the maximum variation (1.81%) and average variation (0.75%) previously reported for Escherichia coli. Pulsed-field gel electrophoresis analysis of I-CeuI-digested genomic DNA revealed seven rrn operons in all E. carotovora subsp. carotovora strains examined except strain WPP17, which had only six copies. We identified 26 I-CeuI restriction fragment length polymorphism patterns and observed significant poiymorphism in fragment sizes ranging from 100 to 450 kb for all strains. We detected large plasmids in two strains, including the model strain E. carotovora subsp, carotovora 71. The two least virulent strains had an unusual chromosomal structure, suggesting that a particular pulsotype is correlated with virulence. To compare chromosomal organization of multiple enterobacterial genomes, several genes were mapped onto I-CeuI fragments. We identified portions of the genome that appear to be conserved across enterobacteria and portions that have undergone genome rearrangements. We found that the least virulent strain, WPPI7, failed to oxidize cellobiose and was missing several hrp and hrc genes. The unexpected variability among isolates obtained from clonal hosts in one region and in one season suggests that factors other than the host plant, potato, drive the evolution of this common environmental bacterium and key plant pathogen. [ABSTRACT FROM AUTHOR]

Pseudomonas syringae is a plant pathogen whose pathogenicity and host specificity are thought to be determined by Hop/Avr effector proteins injected into plant cells by a type III secretion system. P. syringae pv. syringae B728a, which causes brown spot of bean, is a particularly well-studied strain. The type llI secretion system in P. syringae is encoded by hrp (hypersensitive response and pathogenicity) and hrc (hrp conserved) genes, which are clustered in a pathogenicity island with a tripartite structure such that the hrp/hrc genes are flanked by a conserved effector locus and an exchangeable effector locus (EEL). The EELs of P. syringae pv. syringae B728a, P. syringae strain 61, and P. syringae pv. tomato DC3000 differ in size and effector gene composition; the EEL of P. syringae pv. syringae B728a is the largest and most complex. The three putative effector proteins encoded by the P. syringae pv. syringae B728a EEL-HopPsyC, HopPsyE, and HopPsyV--were demonstrated to be secreted in an Hrp-dependent manner in culture. Heterologous expression of hopPsyC, hopPsyE, and hopPsyV in P. syringae pv. tabaci induced the hypersensitive response in tobacco leaves, demonstrating avirulence activity in a nonhost plant. Deletion of the P. syringae pv. syringae B728a EEL strongly reduced virulence in host bean leaves. EELs from nine additional strains representing nine P. syringae pathovars were isolated and sequenced. Homologs of avrPphE (e.g., hopPsyE) and hopPsyA were particularly common. Comparative analyses of these effector genes and hrpK (which flanks the EEL) suggest that the EEL effector genes were acquired by horizontal transfer after the acquisition of the hrp/hrc gene cluster but before the divergence of modern pathovars and that some EELs underwent transpositions yielding effector exchanges or point mutations producing effector pseudogenes after their acquisition. [ABSTRACT FROM AUTHOR]

Soft rot bacteria are the main cause of early seed decay and stand losses in field grown potatoes. These bacteria are present in most surface water and colonize lenticels of early generation seed potatoes and cause disease when conditions are favorable for bacterial growth. Soft rot bacteria, formerly in the genus Erwinia, have been reclassified into the genera Dickeya and Pectobacterium. Because of their ubiquitous association with potato tubers, it is assumed that the soft rot bacteria is spread during handling and planting of seed potatoes and can potentially cause seed decay and blackleg. A soft rot bacterium new to the United States, Dickeya dianthicola was first recognized in 2015 (Secor, unpublished) as the cause of serious stand losses of potatoes in production sites in the eastern US. Replicated and commercial field trials using seed laboratory inoculated and naturally infected with D. dianthicola were conducted in multiple locations to determine spread from infected seed potatoes to healthy seed potatoes during handling and cutting. There is no evidence of increased stand losses or blackleg after simultaneous handling and cutting of Dickeya infected seed potatoes with seed potatoes free of Dickeya. Laboratory trials with labeled soft rot bacteria demonstrated spread of bacteria by cutting knives. There may be other factors that prevent establishment and infection by soft rot bacteria that may spread during seed potato cutting that need to be investigated. We conclude that spread of Dickeya bacteria during handling and cutting of seed potatoes is not a major cause of subsequent seed decay and blackleg resulting in increased stand losses in the field. [ABSTRACT FROM AUTHOR]

Highlights • Primers targeting the highly conserved pipo region specifically recognized different Potato virus Y isolates by RPA. • The RPA assay efficiently detects viral RNA in low viral titer samples, including the peridermal tissues of potato tubers. • The RPA procedure was streamlined by using crude extract from infected-potato tissues. • The real-time detection of Potato virus Y using fluorescent probe picked up the viral RNA within 6 min. Abstract Potato virus Y (PVY) is a global challenge for potato production and the leading cause of seed crop downgrading and rejection for certification. Accurate and timely diagnosis is key to effective control of PVY. Here we optimized the isothermal recombinase polymerase amplification (RPA) for accurate detection of different PVY O and N types that were tested, present in different tissues of potato plants including tubers with a primer set that specifically targets the highly conserved pipo region within the viral genome. Combined with a simplified preparation of the template by tissue homogenization, we established a rapid RPA procedure, which can allow real time detection in less than 10 min with a fluorescent probe. Specificity of the reaction was determined by the lack of cross-reactivity with other common potato viruses. Although RPA reagents remain more expensive than PCR reagents, RPA technology is equivalent in that results can be visualized by gel electrophoresis or with a fluorescent probe with greater sensitivity; and it is superior to the common PCR-based assay in its versatility, speed, and lack of need for a highly purified RNA template. [ABSTRACT FROM AUTHOR]

Potato virus Y (PVY) is the most important virus in North American seed potato ( Solanum tuberosum L.) production. Planting virus-free minitubers in place of field-grown seed, which usually has a low PVY incidence, reduces initial PVY inoculum in the field. However, plants grown from minitubers are smaller and emerge later than those grown from conventional seed, which could make them more likely to become infected with PVY. We tested the effects of seed type of three potato cultivars (Dark Red Norland, Goldrush, and Red La Soda) on PVY incidence, tuber yield, and flowering time. The incidence of PVY in plants grown from minitubers did not differ from that of plants grown from conventional seed. Minituber-grown plants produced lower tuber yields than plants grown from conventional seed. Plants from minitubers also emerged and flowered later, but this did not increase their incidence of PVY. Cultivar-specific differences were observed in tuber yield and flowering times, suggesting that this variation may influence PVY incidence more than seed type. [ABSTRACT FROM AUTHOR]

The advent of genomics has advanced science into a new era, providing a plethora of “toys” for researchers in many related and disparate fields. Genomics has also spawned many new fields, including proteomics and metabolomics, furthering our ability to gain a more comprehensive view of individual organisms and of interacting organisms. Genomic information of both bacterial pathogens and their hosts has provided the critical starting point in understanding the molecular bases of how pathogens disrupt host cells to cause disease. In addition, knowledge of the complete genome sequence of the pathogen provides a potentially broad slate of targets for the development of novel virulence inhibitors that are desperately needed for disease management. Regarding plant bacterial pathogens and disease management, the potential for utilizing genomics resources in the development of durable resistance is enhanced because of developing technologies that enable targeted modification of the host. Here, we summarize the role of genomics studies in furthering efforts to manage bacterial plant diseases and highlight novel genomics-enabled strategies heading down this path. [ABSTRACT FROM AUTHOR]

Potato virus Y (PVY) is responsible for the majority of seed potato lot rejections in North America. Some commercial cultivars and breeding lines have major genes for PVY resistance, but their use in North American breeding programs has been limited. Marker assisted selection of PVY resistance can increase the efficiency of identifying resistant plants and integrating resistance into new cultivars. We used markers RYSC3 and YES3-3B, linked to genes Ry and Ry, respectively, to screen 46 breeding clones and cultivars, and identified 19 resistant clones. Resistant parents were crossed with susceptible parents with good market and agronomic traits and molecular marker analysis of each family showed a 1:1 segregation ratio. The persistence of this segregation ratio over 2 years of selection and the lack of major linkage drag as assessed by a breeder rating indicated that there was no evidence of an association of PVY resistance with undesirable traits. [ABSTRACT FROM AUTHOR]

Pectobacterium species are enterobacterial plant-pathogens that cause soft rot disease in diverse plant species. Unlike hemi-biotrophic plant pathogenic bacteria, the type III secretion system (T3SS) of Pectobacterium carotovorum subsp. carotovorum (P. carotovorum) appears to secrete only one effector protein, DspE. Previously, we found that the T3SS regulator HrpL and the effector DspE are required for P. carotovorum pathogenesis on leaves. Here, we identified genes up-regulated by HrpL, visualized expression of dspE in leaves, and established that DspE causes host cell death. DspE required its full length and WxxxE-like motifs, which are characteristic of the AvrE-family effectors, for host cell death. We also examined expression in plant leaves and showed that hrpL is required for the expression of dspE and hrpN, and that the loss of a functional T3SS had unexpected effects on expression of other genes during leaf infection. These data support a model where P. carotovorum uses the T3SS early in leaf infection to initiate pathogenesis through elicitation of DspE-mediated host cell death. [ABSTRACT FROM AUTHOR]

Pectobacterium species are necrotrophic bacterial pathogens that cause soft rot diseases in potatoes and several other crops worldwide. Gene expression data identified Pectobacterium carotovorum subsp. carotovorum budB, which encodes the aacetolactate synthase enzyme in the 2,3-butanediol pathway, as more highly expressed in potato tubers than potato stems. This pathway is of interest because volatiles produced by the 2,3-butanediol pathway have been shown to act as plant growth promoting molecules, insect attractants, and, in other bacterial species, affect virulence and fitness. Disruption of the 2,3-butanediol pathway reduced virulence of P. c. subsp. carotovorum WPP14 on potato tubers and impaired alkalinization of growth medium and potato tubers under anaerobic conditions. Alkalinization of the milieu via this pathway may aid in plant cell maceration since Pectobacterium pectate lyases are most active at alkaline pH. [ABSTRACT FROM AUTHOR]

The type III secretion system (T3SS) is a major virulence factor in many gram-negative bacterial pathogens. This secretion system translocates effectors directly into the cytosol of eukaryotic host cells, where the effector proteins facilitate bacterial pathogenesis by interfering with host cell signal transduction and other cellular processes. Plants defend themselves against bacterial pathogens by recognizing either the type 3 effectors or their actions and initiating a cascade of defense responses that often results in programmed cell death of the plant cell being attacked. Here we show that a plant phenolic compound, p-coumaric acid (PCA), represses the expression of T3SS genes of the plant pathogen Dickeya dadantii, suggesting that plants can also defend against bacterial pathogens by manipulating the expression of the T3SS. PCA repressed the expression of T3SS regulatory genes through the HrpX/Y two-component system, a core regulator of the T3SS, rather than through the global regulator GacS/A, which indirectly regulates the T3SS. A further analysis of several PCA analogs suggests that the para positioning of the hydroxyl group in the phenyl ring and the double bond of PCA may be important for its biological activity. [ABSTRACT FROM AUTHOR]

Pectobacrerium and Dickeva spp. are related broad-host-range enterobacteria! pathogens of angiosperms. A review of the literature shows that these genera each cause disease in species from at least 35% of angiosperm plant orders. The known host ranges of these pathogens partially overlap and, together, these two genera are pathogens of species from 50% of angiosperm plant orders. Notably, there are no reported hosts for either genus in the eudicots dade and no reported Dickeya hosts in the magnoliids or eurosids II clades, although Pectobacterium spp. are pathogens of at least one plant species in the magnoliids and at least one in each of the three eurosids 11 plant orders. In addition, Dickeva but not Pectobacterium spp. have been reported on a host in the rosids dade and, unlike Pectobacterium spp., have been reported on many Poales species. Natural disease among nonangiosperms has not been reported for either genus. Phylogenetic anaIyses of sequences concatenated from regions of seven housekeeping genes (acnA, gapA, icdA, mdh, mtlD, pgi, and proA) from representatives of these genera demonstrated that Dickeya spp. and the related tree pathogens, the genus Brenneria, are more diverse than Pectobacterium spp. and that the Pectobacterium strains can be divided into at least five distinct clades, three of which contain strains from multiple host plants. [ABSTRACT FROM AUTHOR]

Production of the plant hormone indole-3-acetic acid (IAA) is widespread among plant-associated micro- organisms. The non-gall-forming phytopathogen Erwinia chrysanthemi 3937 (strain Ech3937) possesses iaaM (ASAP16562) and iaaH (ASAP16563) gene homologues. In this work, the null knockout iaaM mutant strain Ech138 was constructed. The IAA production by Ech138 was reduced in M9 minimal medium supplemented with L-tryptophan. Compared with wild-type Ech3937, Ech138 exhibited reduced ability to produce local maceration, but its multiplication in Saintpaulia ionantha was unaffected. The pectate lyase production of Ech138 was diminished. Compared with wild-type Ech3937, the expression levels of an oligogalacturonate lyase gene, ogl, and three endopectate lyase genes, pelD, pelI, and pelL, were reduced in Ech138 as determined by a green fluorescent protein-based fluorescence-activated cell sorting promoter activity assay. In addition, the transcription of type III secretion system (T3SS) genes, dspE (a putative T3SS effector) and hrpN (T3SS harpin), was found to be diminished in the iaaM mutant Ech138. Compared with Ech3937, reduced expression of hrpL (a T3SS alternative sigma factor) and gacA but increased expression of rsmA in Ech138 was also observed, suggesting that the regulation of T3SS and pectate lyase genes by IAA biosynthesis might be partially due to the posttranscriptional regulation of the Gac-Rsm regulatory pathway. [ABSTRACT FROM AUTHOR]

Numerous Salmonella enterica food-borne illness outbreaks have been associated with contaminated vegetables, in particular sprouted seeds, and the incidence of reported contamination has steadily risen. In order to understand the physiology of S. enterica serovar Newport on plants, a screen was developed to identity transposon mutants that were defective in attachment to alfalfa sprouts. Twenty independent mutants from a pool of 6,000 were selected for reduced adherence to alfalfa sprouts. Sixty-five percentage of these mutants had insertions in uncharacterized genes. Among the characterized genes were strains with insertions in the intergenic region between agfB, the surface-exposed aggregative fimbria (curli) nucleator, and agfD, a transcriptional regulator of the LuxR superfamily, and rpoS, the stationary-phase sigma factor. Both AgfD and RpoS have been reported to regulate curli and cellulose production and RpoS regulates other adhesins such as pili. The intergenic and rpoS mutants were reduced in initial attachment to alfalfa sprouts by 1 log unit compared to the wild type. Mutations of agfA, curli subunit, and agfB in S. enterica serovar Enteritidis differentially affected attachment to plant tissue. The agfA mutation was not reduced in ability to attach to or colonize alfalfa sprouts, whereas the agfB mutation was reduced. Thus, agfB alone can play a role in attachment of S. enterica to plant tissue. These results reveal that S. enterica genes important for virulence in animal systems are also required for colonization of plants, a secondary host that can serve as a vector of S. enterica from animal to animal. [ABSTRACT FROM AUTHOR]

Enterobacterial animal pathogens exhibit aggregative multicellular behavior, which is manifested as pellicles on the culture surface and biofilms at the surface-liquid-air interface. Pellicle formation behavior requires production of extracellular polysaccharide, cellulose, and protein filaments, known as curli. Protein filaments analogous to curli are formed by many protein secretion systems, including the type III secretion system (TTSS). Here, we demonstrate that Erwinia chrysanthemi, which does not carry curli genes, requires the TTSS for pellicle formation. These data support a model where cellulose and generic protein filaments, which consist of either curli or TTSS-secreted proteins, are required for enterobacterial aggregative multicellular behavior. Using this assay, we found that hrpY, which encodes a two-component system response regulator homolog, is required for activity of hrpS, which encodes a σ54-dependent enhancer-binding protein homolog. In turn, hrpS is required for activity of the sigma factor homolog hrpL, which activates genes encoding TTSS structural and secreted proteins. Pellicle formation was temperature dependent and pellicles did not form at 36°C, even though TTSS genes were expressed at this temperature. We found that cellulose is a component of the E. chrysanthemi pellicle but that pellicle formation still occurs in a strain with an insertion in a cellulose synthase subunit homolog. Since the TTSS, but not the cellulose synthase subunit, is required for E. chrysanthemi pellicle formation, this inexpensive assay can be used as a high throughput screen for TTSS mutants or inhibitors. [ABSTRACT FROM AUTHOR]

Assesses the gene Erwinia amylovor and the avirulence locus avrE of Pseudomonas syringae. Details on the usefulness of bacterial genes encoding proteins involved in infection of plants; Method used in the assessment process; Indications of the findings

Dickeya dadantii is a plant-pathogenic enterobacterium responsible for the soft rot disease of many plants of economic importance. We present here the sequence of strain 3937, a strain widely used as a model system for research on the molecular biology and pathogenicity of this group of bacteria. [ABSTRACT FROM AUTHOR]

Presents an abstract of the study "Harpin Mediates Cell Aggregation in Erwinia chrysanthemi 3937." Secretion of the hypersensitive response elicitor harpin of soft rot pathogen Erwinia chrysanthemi strains 3937 and EC16; Importance of strain 3937 HrpN for cell aggregation.