No evidence that homologs of key circadian clock genes direct circadian programs of development or mRNA abundance in Verticillium dahliae

Many organisms harbor circadian clocks that promote their adaptation to the rhythmic environment. While a broad knowledge of the molecular mechanism of circadian clocks has been gained through the fungal model Neurospora crassa, little is known about circadian clocks in other fungi. N. crassa belongs to the same class as many important plant pathogens including the vascular wilt fungus Verticillium dahliae. We identified homologs of N. crassa clock proteins in V. dahliae, which showed high conservation in key protein domains. However, no evidence for an endogenous, free- running and entrainable rhythm was observed in the daily formation of conidia and microsclerotia. In N. crassa the frequency (frq) gene encodes a central clock protein expressed rhythmically and in response to light. In contrast, expression of Vdfrq is not light-regulated. Temporal gene expression profiling over 48 h in constant darkness and temperature revealed no circadian expression of key clock genes. Furthermore, RNA-seq over a 24 h time-course revealed no robust oscillations of clock-associated transcripts in constant darkness. Comparison of gene expression between wild-type V. dahliae and

Vegetative compatibility groups partition variation in the virulence of Verticillium dahliae on strawberry

Verticillium dahliae infection of strawberry (Fragaria x ananassa) is a major cause of disease-induced wilting in soil-grown strawberries across the world. To understand what components of the pathogen are affecting disease expression, the presence of the known effector VdAve1 was screened in a sample of Verticillium dahliae isolates. Isolates from strawberry were found to contain VdAve1 and were divided into two major clades, based upon their vegetative compatibility groups (VCG); no UK strawberry isolates contained VdAve1. VC clade was strongly related to their virulence levels. VdAve1-containing isolates pathogenic on strawberry were found in both clades, in contrast to some recently published findings. On strawberry, VdAve1-containing isolates had significantly higher virulence during early infection, which diminished in significance as the infection progressed. Transformation of a virulent non-VdAve1 containing isolate, with VdAve1 was found neither to increase nor decrease virulence when inoculated on a susceptible strawberry cultivar. There are therefore virulence factors that are epistatic to VdAve1 and potentially multiple independent routes to high virulence on strawberry in V. dahliae lineages. Genome sequencing a subset of isolates across the two VCGs revealed that isolates were differentiated at the whole genome level and contained multiple changes in putative effector content, indicating that different clonal VCGs may have evolved different strategies for infecting strawberry, leading to different virulence levels in pathogenicity tests. It is therefore important to consider both clonal lineage and effector complement as the adaptive potential of each lineage will differ, even if they contain the same race determining effector

A new three-locus model for rootstock-induced dwarfing in apple revealed by genetic mapping of root bark percentage

Rootstock-induced dwarfing of apple scions revolutionized global apple production during the twentieth century, leading to the development of modern intensive orchards. A high root bark percentage (the percentage of the whole root area constituted by root cortex) has previously been associated with rootstock induced dwarfing in apple. In this study, the root bark percentage was measured in a full-sib family of ungrafted apple rootstocks and found to be under the control of three loci. Two QTL for root bark percentage were found to co-localise to the same genomic regions on chromosome 5 and chromosome 11 previously identified as controlling dwarfing, Dw1 and Dw2, respectively. A third QTL was identified on chromosome 13 in a region that has not been previously associated with dwarfing. The development of closely linked 3 Sequence-tagged site STS markers improved the resolution of allelic classes thereby allowing the detection of dominance and epistatic interactions between loci, with high root bark percentage only occurring in specific allelic combinations. In addition, we report a significant negative correlation between root bark percentage and stem diameter (an indicator of tree vigour), measured on a clonally propagated grafted subset of the mapping population. The demonstrated link between root bark percentage and rootstock-induced dwarfing of the scion leads us to propose a three-locus model that is able to explain levels of dwarfing from the dwarf ‘M.27’ to the semi-invigorating rootstock ‘M.116’. Moreover, we suggest that the QTL on chromosome 13 (Rb3) might be analogous to a third dwarfing QTL, Dw3 that has not previously been identified

Likelihood ratio tests assessing effects of VC group and race on pathogenicity.

<p>Likelihood ratio tests assessing effects of VC group and race on pathogenicity.</p

Indicative symptoms six weeks post inoculation of the strawberry cultivar ‘Hapil’ with <i>Vertcillium dahliae</i>.

<p>Disease scores six weeks post-inoculation (B) of the strawberry cultivar ‘Hapil’ inoculated with 13 isolates of <i>Verticillium dahliae</i>. ‘Race 1’ isolates are unshaded while ‘Race 2‘ isolates are shaded grey, subclade II-1 and II-2 isolates are denoted by thin and thick borders respectively. Error bars are standard errors.</p

Whole-genome phylogenetic tree of <i>Verticillium dahliae</i> isolates.

<p>Generated using RealPhy based on the alignment of short or shotgun reads from each genome to the reference genome of JR2.</p

Genomic statistics of Verticillium strains.

<p>Genomic statistics of Verticillium strains.</p

Neighbour joining phylogeny of <i>Verticillium dahliae</i> using primers DB19/22 to classify VC clades.

<p>Reference sequences were obtained from Collado-Romero et. al [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191824#pone.0191824.ref035" target="_blank">35</a>]. Bootstrap support values are the result of 1000 resampling events.</p

Divergence of ‘race 2’ isolates from UK strawberry can be divided into 2 groups depending upon VC subclade.

<p>Alignment of short reads to the <i>VdAve1</i> region of the JR2 genome highlights different patterns of gene loss around the <i>VdAve1</i> region which are dependent upon VC subclade; Subclade II-2 isolates (12008,12251,12253) show similar patterns of read alignments to each other and subclade II-1 isolates (12158,12161) again show similarity within VC subclade but not between groups.</p

Variation in gene numbers of <i>Verticillium dahliae</i> strains in secreted and putative effector orthogroups.

<p>Variation in gene numbers of <i>Verticillium dahliae</i> strains in secreted and putative effector orthogroups.</p