Here we present genome sequences for twelve isolates of the invasive pathogen Phytophthora ramorum EU1. The assembled genome sequences and raw sequence data are available via BioProject accession number PRJNA177509. These data will be useful in developing molecular tools for specific detection and identification of this pathogen.

Non-targeted approaches are useful tools to identify new or emerging issues in bee health. Here, we utilise next generation sequencing to highlight bacteria associated with healthy and unhealthy honey bee colonies, and then use targeted methods to screen a wider pool of colonies with known health status. Our results provide the first evidence that bacteria from the genus Arsenophonus are associated with poor health in honey bee colonies. We also discovered Lactobacillus and Leuconostoc spp. were associated with healthier honey bee colonies. Our results highlight the importance of understanding how the wider microbial population relates to honey bee colony health.

Different fertilisation regimes, i.e. the use of inorganic or organic fertilisers used in agriculture, are thought to cause differential effects on soil bacteria. In this study, glasshouse experiments were used to test the effects caused by addition of inorganic fertiliser or digestate from sewage sludge on soil bacterial community structure and diversity assessed by pyrosequencing of the V1-V3 region of the 16S rRNA gene. Spring wheat (Triticum aestivum L., cultivar Paragon) was used as model crop and its growth (measured by total dry weight) was monitored as well as changes in soil nitrogen and phosphorous at three time points over 128 days. Overall, 40 different bacterial phyla were detected with Proteobacteria, Acidobacteria and Actinobacteria dominating the communities. Additionally, members of the Bacteroidetes, Gemmatimonadetes, Chloroflexi and WS3 were found in all samples. Members of the Planctomycetes, Firmicutes, Nitrospirae, candidate division SPAM and Armatimonadetes were found in all samples but at lower abundances. Within the phylum Proteobacteria the classes Alpha-and Betaproteobacteria were most prevalent. Over the course of the experiment, the major differences between treatments were observed for the Actino-, Proteo-and the Acidobacteria. Sequences related to the Planctomycetes, implicated in nitrogen cycling, decreased in all treatments during the course of the experiment. Statistical analyses revealed that mainly nutrient addition and plant growth influenced the bacterial community structure. The effects of the treatment itself could be attributed to different gain in wheat growth especially when the communities were compared at the end of the trial. Our results indicate that the usage of different fertilisers will not only affect the bacterial community by direct addition of nutrients, but also indirectly. Crown Copyright (C) 2014 Published by Elsevier B.V. All rights reserved.

Xanthomonas vasicola pathovar vasculorum (Xvv) is the bacterial agent causing gumming disease in sugarcane. Here, we compare complete genome sequences for five isolates of Xvv originating from sugarcane and one from maize. This identified two distinct types of lipopolysaccharide synthesis gene clusters among Xvv isolates: one is similar to that of Xanthomonas axonopodis pathovar citri (Xac) and is probably the ancestral type, while the other is similar to those of the sugarcane-inhabiting species, Xanthomonas sacchari. Four of six Xvv isolates harboured sequences similar to the Xac plasmid, pXAC47, and showed a distinct Type-IV pilus (T4P) sequence type, whereas the T4P locus of the other two isolates resembled that of the closely related banana pathogen, Xanthomonas campestris pathovar musacearum (Xcm). The Xvv isolate from maize has lost a gene encoding a homologue of the virulence effector, xopAF, which was present in all five of the sugarcane isolates, while xopL contained a premature stop codon in four out of six isolates. These findings shed new light on evolutionary events since the divergence of Xvv and Xcm, as well as further elucidating the relationships between the two closely related pathogens.

Melissococcus plutonius is the bacterial pathogen that causes European Foulbrood of honeybees, a globally important honeybee brood disease. We have used next-generation sequencing to identify highly polymorphic regions in an otherwise genetically homogenous organism, and used these loci to create a modified MLST scheme. This synthesis of a proven typing scheme format with next-generation sequencing combines reliability and low costs with insights only available from high-throughput sequencing technologies. Using this scheme we show that the global distribution of M.plutonius variants is not uniform. We use the scheme in epidemiological studies to trace movements of infective material around England, insights that would have been impossible to confirm without the typing scheme. We also demonstrate the persistence of local variants over time.

The bacterium Xanthomonas campestris pathovar musacearum (Xcm) is the causal agent of banana Xanthomonas wilt (BXW). This disease has devastated economies based on banana and plantain crops (Musa species) in East Africa. Here we use genome-wide sequencing to discover a set of single-nucleotide polymorphisms (SNPs) among East African isolates of Xcm. These SNPs have potential as molecular markers for phylogeographic studies of the epidemiology and spread of the pathogen. Our analysis reveals two major sub-lineages of the pathogen, suggesting that the current outbreaks of BXW on Musa species in the region may have more than one introductory event, perhaps from Ethiopia. Also, based on comparisons of genome-wide sequence data from multiple isolates of Xcm and multiple strains of X. vasicola pathovar vasculorum, we identify genes specific to Xcm that could be used to specifically detect Xcm by PCR-based methods.

Following publication of our article [1], we found errors in analyses performed by the corresponding author (DJS) related to the phylogenetic relationship between Xylella species and the other xanthomonads. These errors do not make any difference to the main findings and conclusions reported in our paper. For example, the phylogenetic positions of NCPPB1131, NCPPB1132 and NCPPB4393 within the Group 1 Xanthomonas species are unaffected. However, we wish to apologize to the authors of a previous work [2] for creating any negative impression on the quality of their phylogenetic analyses and to take this opportunity to rectify the errors. [...]

We present draft genome sequences for three strains of Xanthomonas species, each of which was associated with banana plants (Musa species) but is not closely related to the previously sequenced banana-pathogen Xanthomonas campestris pathovar musacearum. Strain NCPPB4393 had been deposited as Xanthomonas campestris pathovar musacearum but in fact falls within the species Xanthomonas sacchari. Strain NCPPB1132 is more distantly related to Xanthomonas sacchari whilst strain NCPPB 1131 grouped in a distinct species-level clade related to X. sacchari, along with strains from ginger, rice, cotton and sugarcane. These three newly sequenced strains share many genomic features with the previously sequenced Xanthomonas albilineans, for example possessing an unsual metE allele and lacking the Hrp type III secretion system. However, they are distinct from Xanthomonas albilineans in many respects, for example showing little evidence of genome reduction. They also lack the SPI-1 type III secretion system found in Xanthomonas albilineans. Unlike X. albilineans, all three strains possess a gum gene cluster. The data reported here provide the first genome-wide survey of non-Hrp Xanthomonas species other than Xanthomonas albilineans, which is an atypical member of this group. We hope that the availability of complete sequence data for this group of organisms is the first step towards understanding their interactions with plants and identifying potential virulence factors.

Soil Microbiology and Sustainable Crop Production . Dixon, G.R. and Tilston, E.L. ( eds ). Dordrecht, The Netherlands : Springer , 2010 . ISBN 978-90-481-9478-0 . The sustainability of global food security is on a knife-edge. That’s the conclusion of just about every recent analysis of the demands placed on world farming by the seven billion people who rely on it. The plant pathology community knows better than most the challenges of meeting this demand. How do we counter disease and improve yield while satisfying the need to conserve water, safeguard soils and meet demands to reduce chemical input? We’re all familiar with these questions, and quote them liberally in our publications and grant proposals. But the study of soil microbiology and its influence on the health and productivity of crops can sometimes seem like the poor cousin of research into agrochemicals, new crop varieties and epidemic diseases. Soil Microbiology and Sustainable Crop Production is therefore a welcome addition to the literature. Its publication is timely, coming as increasing attention is focused on the need to conserve soil health, but also as new and emerging analysis methods are set to revolutionize our understanding of soil microbes, their populations and function. The increasingly accessible ‘omic’ technologies for genetic and metabolomic analysis of community structure and function will undoubtedly redefine our view of soil health from a microbiological viewpoint. Dixon and Tilston’s book does not cover these technical advances in detail, but this is such a rapidly developing area that to freeze-frame it in an academic text would outdate the book in an instant. Instead the editors have chosen to review the underlying knowledge on soil microbiology in some detail: a fertile soil, if you like, in which both student and the more experienced researcher can grow their own appreciation of the more rapidly-developing literature. And the editors have chosen well. After setting the scene by describing the importance of healthy soils for the world’s food production, the book defines its own wide scope. Chapters describe how microbes, plants and their soil environment interact, the effect of land use on microbes, how soil microorganisms can suppress disease and the potential impact of a changing environment on these interactions. In these chapters the book repeatedly provides arguments for steering intensive farming towards conservation agriculture and, of course, supporting the research that enables this change. Of particular interest to readers of Plant Pathology will be the sections devoted to suppression of disease. In a work of this scope it is inevitable that some key examples are given only brief mention. The suppression of pathogenic Streptomyces, for example, is discussed only in the context of soil pH, although the agricultural practices that achieve the best control of this pathogen are more varied. Streptomyces should perhaps have been included in the otherwise extensive treatment of the effects of irrigation, as the literature raises interesting questions about the influence of this practice on beneficial microbial communities and suppression of this pathogen. But this is not a significant criticism; viewed as an extensive primer rather than an intensive review of the literature, the book does its job well in demonstrating the value of soil microbiology research to inform better management of disease. An interesting inclusion, which serves to demonstrate the ‘bigger picture’ is the chapter by Sandhu et al. on the economic and social impact of soil microbes, which could just as easily be called a review on the past effects of economics on soil microbes, as it discusses the effect of agricultural intensification and shift towards monocultures on the soil microbial resource. This alone demonstrates why a hard look at agricultural practices to conserve soil function is long overdue. The pure scientist may think it a shame that environmental value should be expressed in dollars, but this chapter provides compelling reasoning that rising energy and nutrient prices make the ecosystem services provided by soil microbes increasingly invaluable. In short this book is an important waymark on our journey to discover and value one of the most vital resources for agriculture worldwide.

Banana Xanthomonas wilt is a newly emerging disease that is currently threatening the livelihoods of millions of farmers in East Africa. The causative agent is Xanthomonas campestris pathovar musacearum (Xcm), but previous work suggests that this pathogen is much more closely related to species Xanthomonas vasicola than to X. campestris. We have generated draft genome sequences for a banana-pathogenic strain of Xcm isolated in Uganda and for a very closely related strain of X. vasicola pathovar vasculorum, originally isolated from sugarcane, that is nonpathogenic on banana. The draft sequences revealed overlapping but distinct repertoires of candidate virulence effectors in the two strains. Both strains encode homologues of the Pseudomonas syringae effectors HopW, HopAF1 and RipT from Ralstonia solanacearum. The banana-pathogenic and non-banana-pathogenic strains also differed with respect to lipopolysaccharide synthesis and type-IV pili, and in at least several thousand single-nucleotide polymorphisms in the core conserved genome. We found evidence of horizontal transfer between X. vasicola and very distantly related bacteria, including members of other divisions of the Proteobacteria. The availability of these draft genomes will be an invaluable tool for further studies aimed at understanding and combating this important disease.

Disease in compatible hosts and induction of the hypersensitive response in resistant plants by most plant-pathogenic bacteria require a functional type III secretion system (T3SS). Expression of T3SS genes responds to host and environmental factors and is induced within the plant. In Pseudomonas syringae, expression of the T3SS requires HrpL, which is transcriptionally upregulated by HrpR and HrpS. In some pathovars, expression of the hrpRS genes is upregulated by the GacA/S two-component system. Additionally, HrpA, the major component of the T3SS pilus, has also been linked to the regulation of the hrpRS gene expression. Previous studies concerning regulation of hypersensitive response and pathogenesis/hypersensitive response conserved (hrp/hrc) gene expression have used mostly in vitro inducing conditions, different pathovars, and methodology. Here, we analyze the roles of HrpL, GacA, and HrpA in the bean pathogen, using single, double, and triple mutants as well as strains ectopically expressing the regulators. We use real-time polymerase chain reaction analysis in vitro and in planta to quantify gene expression and competitive indices and other assays to assess bacterial fitness. Our results indicate that i) HrpL acts as a general virulence regulator that upregulates non-T3SS virulence determinants and downregulates flagellar function; ii) GacA modulates the expression of hrpL, and its contribution to virulence is entirely HrpL dependent; iii) there is a basal HrpL-independent expression of the T3SS genes in rich medium that is important for full activation of the system, maybe by keeping the system primed for rapid activation upon contact with the plant; and iv) HrpA upregulates expression of the T3SS genes and is essential to activate expression of the hrpZ operon upon contact with the plant.

European foulbrood (EFB) persists in England and Wales despite current treatment methods, all of which include feeding honey bee colonies with the antibiotic oxytetracycline (OTC). A large-scale field experiment was conducted to monitor a husbandry-based method, using comb replacement (known as Shook swarm), as a drug free EFB control option. The understanding of EFB epidemiology is limited, with little information on the presence of Melissococcus plutonius in disease free colonies. Additional samples were collected from diseased and disease free apiaries to identify symptomless infection. EFB reoccurrence was not significantly different between OTC and husbandry methods and real-time PCR data demonstrated that fewer Shook swarm treated colonies contained M. plutonius carryover to the Spring following treatment. Asymptomatic colonies from diseased apiaries showed an increased risk of testing positive for M. plutonius compared to asymptomatic colonies from disease free apiaries. The probability of a sample being symptomatic increased when a greater quantity of M. plutonius was detected in adult bees and larvae. The possibility of treating EFB as an apiary disease rather than a colony disease and the implications of a control strategy without antibiotics are discussed.

Genome-wide sequencing data revealsvirulence factors implicated in bananaXanthomonas wilt David J. Studholme,, Eric Kemen, Daniel MacLean, Sebastian Schornack, Valente Aritua, Richard Thwaites, Murray Grant, Julian Smith & Jonathan D.G. Jones University of Exeter, Exeter, UK; The Sainsbury Laboratory, Norwich, UK; National Agricultural Biotechnology Center, Kawanda Agricultural Research Institute, Kampala, Uganda; and The Food and Environment Research Agency, York, UK

A recently emerging bleeding canker disease, caused by Pseudomonas syringae pathovar aesculi (Pae), is threatening European horse chestnut in northwest Europe. Very little is known about the origin and biology of this new disease. We used the nucleotide sequences of seven commonly used marker genes to investigate the phylogeny of three strains isolated recently from bleeding stem cankers on European horse chestnut in Britain (E-Pae). On the basis of these sequences alone, the E-Pae strains were identical to the Pae type-strain (I-Pae), isolated from leaf spots on Indian horse chestnut in India in 1969. The phylogenetic analyses also showed that Pae belongs to a distinct clade of P. syringae pathovars adapted to woody hosts. We generated genome-wide Illumina sequence data from the three E-Pae strains and one strain of I-Pae. Comparative genomic analyses revealed pathovar-specific genomic regions in Pae potentially implicated in virulence on a tree host, including genes for the catabolism of plant-derived aromatic compounds and enterobactin synthesis. Several gene clusters displayed intra-pathovar variation, including those encoding type IV secretion, a novel fatty acid biosynthesis pathway and a sucrose uptake pathway. Rates of single nucleotide polymorphisms in the four Pae genomes indicate that the three E-Pae strains diverged from each other much more recently than they diverged from I-Pae. The very low genetic diversity among the three geographically distinct E-Pae strains suggests that they originate from a single, recent introduction into Britain, thus highlighting the serious environmental risks posed by the spread of an exotic plant pathogenic bacterium to a new geographic location. The genomic regions in Pae that are absent from other P. syringae pathovars that infect herbaceous hosts may represent candidate genetic adaptations to infection of the woody parts of the tree.