Increased resistance to Verticillium dahliae in Arabidopsis plants defective in auxin signalling

Auxin signalling and transport participate in plant–microbe interactions as positive or negative regulators of disease resistance. The present study investigated the responses of Arabidopsis thaliana plants impaired in the auxin receptors TIR1, AFB1 and AFB3 and the auxin transporter AXR4, upon infection by the soilborne root pathogen Verticillium dahliae. Fewer symptoms were recorded in afb1, afb3 and axr4 plants compared to the wild type (wt). qPCR analysis revealed that the decrease in symptom severity in afb1, afb3 and axr4 was correlated with reduction in the growth of the pathogen in the plants. Therefore, afb1, afb3 and axr4 are partially resistant to V. dahliae. Upon V. dahliae infection, the expression of TIR1, AFB1, AFB3 and AXR4 was up‐regulated in roots, while indole‐3‐acetic acid levels were similar to mocks. The partial resistance of afb1, afb3 and axr4 against V. dahliae can be attributed in part to the up‐regulation of defence‐related genes, as it was observed that afb1 and axr4 had higher PR1 levels than wt, while afb3 had higher PDF1.2 levels than wt. The findings of the present study suggest that the auxin signalling defective mutants afb1, afb3 and axr4 show increased resistance against V. dahliae.     

Biological control of Pythium,Rhizoctonia and Sclerotinia in lettuce: association of the plant protective activity of the bacterium Paenibacillus alvei K165 with the induction of systemic resistance

The soilborne fungi Sclerotinia sclerotiorum, Rhizoctonia solani and the oomycete Pythium ultimum are among the most destructive pathogens for lettuce production. The application of the biocontrol agent Paenibacillus alvei K165 to the transplant soil plug of lettuce resulted in reduced S. sclerotiorum, R. solani and P. ultimum foliar symptoms and incidence compared to untreated controls, despite the suppressive effect of the pathogens on the rhizosphere population of K165. In vitro, K165 inhibited the growth of S. sclerotiorum and R. solani but not P. ultimum. Furthermore, the expression of the pathogenesis‐related (PR) gene PR1, a marker gene of salicylic acid (SA)‐dependent plant defence, and of the Lipoxygenase (LOX) and Ethylene response factor 1 (ERF1) genes, markers of ethylene/jasmonate (ET/JA)‐dependent plant defence was recorded. K165‐treated plants challenged with P. ultimum showed up‐regulation of PR1, whereas challenge with R. solani resulted in up‐regulation of LOX and ERF1, and challenge with S. sclerotiorum resulted in up‐regulation of PR1, LOX and ERF1. This suggests that K165 triggers the SA‐ and the ET/JA‐mediated induced systemic resistance against P. ultimum and R. solani, respectively, while the simultaneous activation of the SA and ET/JA signalling pathways is proposed for S. sclerotiorum.     

Plant–herbivore asynchrony necessitates augmentative releases of the exotic beetle,Zygogramma bicolorata,to enhance the biological control of P arthenium hysterophorus

Systematic information on the quantitative impact of Z ygogramma bicolorata on the biology of P arthenium hysterophorus is crucial as the seeds of this weed continue to germinate from the accumulated soil seed bank throughout the year in the form of different germinating flushes, while the activity of the beetle ceases during winter as it enters diapause. Therefore, plant–herbivore interactions need to be explored to develop predictions of the overall impact of the introduced beetle on the weed. The findings revealed that defoliation by Z . bicolorata had a significant impact on the plant height, density and flower production in flushes F ₃, F ₄ and F ₅, but not in F ₁ and F ₂ that exhibited longer periodicity, profuse branching, a longer flowering period and maximum flower production and contributed mostly to the existing seed soil bank. Therefore, total depletion of the existing soil seed bank was not possible. Consequently, the effect of augmentative field releases of laboratory‐reared beetles was explored on F ₁ and F ₂ in February for three consecutive years (2011–2013). Before initiating the trial, random soil samples were taken from the plots that were assigned to the paired treatments (i.e. with the beetle and without the beetle [insecticide‐treated]) and it was found that the seed bank in those samples did not differ. The single release of Z . bicolorata adults at five per plant at the six‐leaf stage significantly reduced the soil seed bank, compared to without the biocontrol agent, irrespective of the flushes at the end of the season.     

Detection of Tomato yellow leaf curl virus in imported tomato fruit in northern Europe

Imported tomato fruits infected with Tomato yellow leaf curl virus (TYLCV) were identified on the market in northern Europe using paper‐based FTA Classic Cards (Whatman), polymerase chain reaction (PCR) and partial DNA sequence analysis. Trade tomatoes originating from southern Europe, Africa and the Middle East were sampled in Estonia and Sweden, and tested for infection with begomoviruses. Out of 100 batches analysed with five fruits sampled in each batch (58 batches from Estonia and 42 from Sweden), 20 batches were positive (16 from Estonia and four from Sweden). Rolling circle amplification (RCA) and full‐length genome sequence analysis of one isolate collected in Estonia and one isolate in Sweden, revealed highest nucleotide sequence identity at 99% to TYLCV‐IL for the Estonian isolate and at 97% to TYLCV‐Mld for the Swedish isolate. In this study, TYLCV was identified for the first time in imported tomato fruits on the market in northern Europe. FTA cards proved to be an effective means to collect, extract and store begomovirus DNA from tomato fruits and the subsequent molecular analysis.     

Species composition and genetic structure of Fusarium graminearum species complex populations affecting the main barley growing regions of South America

Members of the Fusarium graminearum species complex (FGSC), such as F. graminearum and F. asiaticum, are the main cause of fusarium head blight (FHB) of wheat and barley worldwide. In this study, 117 FGSC isolates obtained from commercial barley grain produced in Argentina (n = 43 isolates), Brazil (n = 35), and Uruguay (n = 39) were identified to species and trichothecene genotypes, and analysed using amplified fragment length polymorphism (AFLP) and sequence‐related amplified polymorphism (SRAP) markers. In addition, reductase (RED) and trichothecene 3‐O‐acetyltransferase (Tri101) were sequenced for a subset of 24 isolates. The majority of the isolates (n = 103) were identified as F. graminearum, which was the only species found in Argentina. In Uruguay, only one F. cortaderiae isolate was found among F. graminearum isolates. In Brazil, F. graminearum also dominated the collection (22/35), followed by F. meridionale (8/35), F. asiaticum (2/35), F. cortaderiae (2/35) and F. austroamericanum (1/35). Species were structured by trichothecene genotype: all F. graminearum were of the 15‐acetyldeoxynivalenol (ADON), F. meridionale, F. asiaticum and F. cortaderiae were of the nivalenol (NIV), and F. austroamericanum was of the 3‐ADON genotype. Both AFLP and SRAP data showed high levels of genetic variability, which was higher within than among countries. Isolates were not structured by country of origin. SRAP analysis grouped F. graminearum in a separate cluster from the other species within the complex. However, AFLP analysis failed to resolve the species into distinct clades with partial clustering of F. meridionale, F. austroamericanum, F. asiaticum and F. graminearum isolates.     

Biological and molecular variation of Cherry leaf roll virus isolates from Malus domestica,Ribes rubrum,Rubus idaeus,Rumex obtusifolius and Vaccinium darrowii

Cherry leaf roll virus (CLRV) isolates from Malus domestica, Ribes rubrum, Rubus idaeus, Rumex obtusifolius and Vaccinium darrowii were characterized based on nucleotide sequences of a 371 bp fragment of the 3′ untranslated region (UTR) of their genomic RNAs, symptoms in the herbaceous hosts, Chenopodium amaranticolor, Chenopodium quinoa, Nicotiana benthamiana, Nicotiana occidentalis and Nicotiana tabacum, and seed transmission in N. occidentalis. The different isolates induced a range of localized and systemic disease symptoms, of varying severity, in the herbaceous hosts. The isolates from M. domestica, R. rubrum, R. obtusifolius and V. darrowii all showed greater than 80% seed transmission in N. occidentalis, but no seed transmission was observed for the R. idaeus isolate. Based on symptoms and seed transmission, the isolates appear to be biologically distinct strains of CLRV. Phylogenetic analysis of the nucleotide sequences from the 3′ UTR, commonly used to detect CLRV, showed that four isolates from M. domestica, R. rubrum, R. idaeus and V. darrowii were almost identical but an isolate from R. obtusifolius exhibited a pairwise nucleotide difference of up to 5·4% when compared to these isolates. There was no obvious correlation between sequence differences and symptomatology.     

Pathogenicity of 21 newly described Phytophthora species against seven Western Australian native plant species

The pathogenicity of some Phytophthora species recently described from Western Australia, together with P. cinnamomi as a control, was tested against seven Western Australian native plant species in the glasshouse. Host species were Banksia grandis, B. littoralis, B. occidentalis, Casuarina obesa, Corymbia calophylla, Eucalyptus marginata and Lambertia inermis. Twenty‐two Phytophthora species were grown on a vermiculite, millet seed and V8 substrate and used as soil inoculum when the plant hosts were approximately 3 months old. Pathogenicity was assessed after 6 weeks and plants were scored for death, root damage, and percentage reduction of shoot growth compared with control plants. The pathogenicity of P. cinnamomi was confirmed. Phytophthora niederhauserii was shown to be similar to P. cinnamomi in pathogenicity and of concern ecologically. Other species that killed one or more hosts were P. boodjera, P. constricta, P. elongata, P. moyootj and P. rosacearum, while P. condilina, P. gibbosa, P. gregata, P. litoralis and P. ‘personii’ caused significant reduction to shoot and/or root growth, but did not kill plants. Host species susceptible to the highest number of Phytophthora species were B. grandis, B. littoralis, B. occidentalis and E. marginata. No Phytophthora species tested killed C. calophylla.     

Involvement of FgMad2 and FgBub1 in regulating fungal development and carbendazim resistance in Fusarium graminearum

Resistance to carbendazim of Fusarium graminearum is conferred by point mutation in the β₂‐tubulin gene that plays an important role in spindle assembly. The spindle assembly checkpoint is a cellular surveillance system that is critical for maintaining genomic stability. Predicted protein Mad2‐ and Bub1‐encoding genes in F. graminearum (FgMad2 and FgBub1) were isolated and characterized. There was no difference in FgMad2 and FgBub1 expression levels between carbendazim‐sensitive and ‐resistant strains; however, after carbendazim treatment FgMad2 expression increased while FgBub1 expression stayed the same. Both the FgMad2 and FgBub1 deletion mutants became more sensitive to carbendazim. The FgMad2 deletion mutants grew more slowly, produced fewer conidia and both hyphae and conidia were malformed. Conversely, deletion of FgBub1 had no effect on fungal development other than a reduction in conidia production. FgMad2 deletion mutants exhibited a severe decrease in perithecia production and pathogenicity along with a down‐regulation of trichothecene production, whereas FgBub1 deletion mutants exhibited only a slight reduction in perithecia production and was accompanied by a twofold increase in trichothecene production. Overall, the results indicate that both FgMad2 and FgBub1 are involved in carbendazim resistance and trichothecene biosynthesis, and FgMad2 plays an important role in fungal development in F. graminearum.     

First report of Siphonatrophia cupressi and its parasitoids in Algeria and a first record for Africa

This paper presents the first finding of Siphonatrophia cupressi (Swain, 1918) (Hemiptera, Aphididae) for Algeria and also for the whole African continent. This aphid pest is reported for the first time from Khenchela Province, North‐Eastern Algeria, on Cupressus sempervirens L. var. horizontalis (Mill.). The first observations were made in a public garden, and other colonies were detected in a nursery producing forest seedlings. Monitoring of these colonies revealed that this aphid is parasitized by Ephedrus persicae Frog and Lysiphlebus testaceipes (Cress.) (Hymenoptera, Braconidae, Aphidiinae).     

Identification and characterization of bacteria isolated from crown galls on stone fruits in Poland

Eighty stone fruit nurseries located in different regions of Poland were examined for the presence of crown gall affected plants. The disease was observed in 39 nurseries, and galls were sampled for bacterial isolation. Out of 1213 isolates, 409 were pre‐identified as Agrobacterium/Rhizobium spp. with 23S rDNA‐based multiplex PCR, and out of these, 315 were pathogenic when tested on sunflowers. Sequence analysis of three housekeeping genes (fusA, recA, rpoD) revealed that 366 strains belonged to Rhizobium rhizogenes, 23 to Agrobacterium tumefaciens species complex, and the rest of the strains were allocated to new phylogenetic lineages. Of these, the most numerous was the lineage allocated in the Pararhizobium genus. Positive results obtained from pathogenicity tests were generally in agreement with results obtained by PCR with primers complementary to T‐DNA except for two strains, which were positive for PCR but negative for the pathogenicity test. All detected Ti plasmids were nopaline‐type. Independent of their pathogenicity, 59% of tested strains were not sensitive to agrocin 84 in in vitro tests. Analysis of biochemical and physiological features distinguished 50 groups with different phenotypic profiles, but the tested traits were not consistent for strains classified to one taxon. This finding shows limited value of biochemical tests in identification procedures. The bacteria causing tumours were heterogeneous and strains classified to different taxa were found even in a single tumour.     

Inoculation of Malus genotypes with a set of Erwinia amylovora strains indicates a gene‐for‐gene relationship between the effector gene eop1 and both Malus floribunda 821 and Malus ‘Evereste’

The Gram‐negative bacterium Erwinia amylovora, causal agent of fire blight disease in pome fruit trees, encodes a type three secretion system (T3SS) that translocates effector proteins into plant cells that collectively function to suppress host defences and enable pathogenesis. Until now, there has only been limited knowledge about the interaction of effector proteins and host resistance presented in several wild Malus species. This study tested disease responses in several Malus wild species with a set of effector deletion mutant strains and several highly virulent E. amylovora strains, which are assumed to influence the host resistance response of fire blight‐resistant Malus species. The findings confirm earlier studies that deletion of the T3SS abolished virulence of the pathogen. Furthermore, a new gene‐for‐gene relationship was established between the effector protein Eop1 and the fire blight resistant ornamental apple cultivar Evereste and the wild species Malus floribunda 821. The results presented here provide new insights into the host–pathogen interactions between Malus sp. and E. amylovora.     

Measuring lesion attributes and analysing their spatial patterns at the leaf scale using digital image analysis

Digital image analysis was used to quantify size, shape and relative positions of individual plant disease lesions to determine their spatial distribution pattern at the leaf scale. Rice brown spot was used as a necrotrophic pathogen causing numerous discrete lesions. A 50‐leaf subsample was selected from an existing data set of 350 images of leaves taken from the field, and analysed for disease severity using image analysis. Further measurements included size, shape and the relative positions of lesions for all leaves with severity > 8% (n = 25) and an additional 25‐leaf sample with severity <8%. A total of 3964 necrotic and/or halo areas were selected using a manually defined threshold in the computer program Assess . There were significant and positive associations (Pearson's r > 0.81; P < 0.001) between the size‐related measurements (lesion area, longest and shortest axis). Coalesced areas, formed by interconnection of lesions and associated haloes, and a high number of small lesions were found with an increase in severity, suggesting a secondary cycle and autoinfection process. Results from quadrat‐based (Poisson distribution and Spatial Analysis by Distance IndicEs) and distance‐based (point‐process Poisson) spatial methods were in good agreement and, together with a Taylor power law model, suggested a shift from random to predominantly aggregated patterns of lesions at severities approaching 10%. This framework, which is applicable to other foliar diseases, proved useful in providing quantitative knowledge of epidemic processes at the leaf scale. Finally, these results may be useful in improving simulation models and disease assessment methods.     

Development of three fusarium crown rot causal agents and systemic translocation of deoxynivalenol following stem base infection of soft wheat

Fusarium pseudograminearum, F. culmorum and F. graminearum are the most important fusarium crown rot (FCR) causal agents. They have the common ability to biosynthesize deoxynivalenol (DON). To elucidate the behaviour of each of the three species, a comparative study was carried out to investigate symptom progression, fungal systemic growth and translocation of DON following stem base inoculation of soft wheat. FCR symptoms were mainly localized in the inoculated area, which extended up to the second node for all inoculated species. Only the most aggressive strains caused symptoms up to the third node. Real‐time quantitative PCR showed that fungal colonization reached the third node for all the tested species, but a low percentage of plants showed colonization above the third node following inoculation with the most aggressive strains. Fungal growth was detected in symptomless tissues but none of the three species was able to colonize as far as the head tissues. However, even if the pathogens were not detected in the heads, DON was detected in head tissues of the plants inoculated with the most aggressive strains. These results demonstrate that F. pseudograminearum, F. culmorum and F. graminearum, under the same experimental conditions, follow a similar pattern of symptom progression, fungal colonization and DON translocation after stem base infection. Differences in the extent of symptoms, fungal colonization and mycotoxin distribution were mainly attributable to strain aggressiveness. These findings provide comparative information on the events following infection of the stem base of wheat by three of the most important FCR casual agents.     

Diversity of Avr‐vnt1 and AvrSmira1 effector genes in Polish and Norwegian populations of Phytophthora infestans

The oomycete Phytophthora infestans, the cause of late blight, is one of the most important potato pathogens. During infection, it secretes effector proteins that manipulate host cell function, thus contributing to pathogenicity. This study examines sequence differentiation of two P. infestans effectors from 91 isolates collected in Poland and Norway and five reference isolates. A gene encoding the Avr‐vnt1 effector, recognized by the potato Rpi‐phu1 resistance gene product, is conserved. In contrast, the second effector, AvrSmira1 recognized by Rpi‐Smira1, is highly diverse. Both effectors contain positively selected amino acids. A majority of the polymorphisms and all selected sites are located in the effector C‐terminal region, which is responsible for their function inside host cells. Hence it is concluded that they are associated with a response to diversified target protein or recognition avoidance. Diversification of the AvrSmira1 effector sequences, which existed prior to the large‐scale cultivation of plants containing the Rpi‐Smira1 gene, may reduce the predicted durability of resistance provided by this gene. Although no isolates virulent to plants with the Rpi‐phu1 gene were found, the corresponding Avr‐vnt1 effector has undergone selection, providing evidence for an ongoing ‘arms race’ between the host and pathogen. Both genes remain valuable components for resistance gene pyramiding.     

Potential impacts of climate change on the threat of potato cyst nematode species in Great Britain

Potato cyst nematode (PCN) species have different temperature optima for various life cycle stages, therefore a risk assessment of the threat of PCN species under future climates is essential to guide adaptation strategies. Data defining the spatial coverage of potato crops in Great Britain were combined with probabilistic climate change data and a newly developed PCN life cycle model to project the future risk to potato crops from PCN. The model was based on the results of controlled environment experiments to investigate the effect of temperature on survival to female maturity using three PCN populations: Globodera pallida (Lindley) and G. rostochiensis from the James Hutton Institute PCN collection, and a field population of G. pallida (S‐Fife). It was found that projected increases in soil temperature could result in increased survival to female maturity for all three PCN populations, with greater increases expected for Scotland, followed by Wales then England. The largest projected increases in Scotland were for G. pallida, whereas G. rostochiensis showed the largest increases in Wales and England. The potential impact of several agronomic adaptation strategies on projected PCN risk were also investigated. The results from the model suggest that soil infestation levels would have to be reduced by up to 40% in order to negate projected increases in PCN risk, and that advancing the start date of the growing season or modifying planting patterns could be successful strategies to reduce future PCN risk.     

HRM analysis provides insights on the reproduction mode and the population structure of Gnomoniopsis castaneae in Europe

Gnomoniopsis castaneae is an emergent nut rot agent of chestnut in southern Europe. To elucidate its population genetics, three simple sequence repeat (SSR) and two hypervariable markers were developed and assessed through high‐resolution melting (HRM) analysis on 132 isolates collected from 10 sites in Italy, France and Switzerland. High allele diversity (ranging from 0.23 to 0.40 depending on site) and number of haplotypes (49) were observed. More than 70% of the molecular variance could be accounted among isolates within sites. Multilocus analysis showed absence of linkage disequilibrium, suggesting a predominant role played by sexual reproduction and random mating. Data analyses indicated the presence of at least two putative distinct subpopulations and this was confirmed by several approaches, including analysis of shared haplotypes, multivariate and Bayesian analyses. Based on data of allelic diversity, the possibility that the pathogen could have been introduced is discussed. This work assessed the genetic variability and the sexual strategies of G. castaneae in Europe, adding useful information on the epidemiology of this fungal plant pathogen.     

Differences in constitutive and inducible defences in pine species determining susceptibility to pinewood nematode

The pinewood nematode, Bursaphelenchus xylophilus, originating from North America (NA), is a major invasive pine pest in Eurasia. It was first detected in Portugal in 1999 associated with maritime pine, Pinus pinaster, and has been differently affecting the main local pine species, P. pinaster and P. pinea. Field studies and direct inoculation experiments in Pinus spp. seedlings, under controlled conditions, were performed to assess whether the differences in constitutive and inducible defences are determining the different susceptibility of pine host species to B. xylophilus. Host co‐evolution with the pathogen was also assessed, including the NA P. radiata, widely used in forestry in the northeast of the Iberian peninsula. Pine mortality in the field was positively related with the abundance of B. xylophilus, and concentration of phenolics and condensed tannins in pines. In the greenhouse assay, seedling tissues were analysed for constitutive investment in defences, as well as the potential inducibility of those defences as driven by B. xylophilus inoculation. Slower growing P. pinea presented higher levels of constitutive defences than faster growing P. pinaster, with only P. pinaster being affected by B. xylophilus. Furthermore, co‐evolution with the pathogen is important, with the fast‐growing NA P. radiata presenting an inducible and effective response to B. xylophilus. Results point to the importance of integrating data on pine life history traits, including growth rate, and production of constitutive and inducible defences, into predictive models for this invasive forest pest.     

Co‐infection by Botryosphaeriaceae and Ilyonectria spp. fungi during propagation causes decline of young grafted grapevines

Decline of newly planted, grafted grapevines is a serious viticultural problem worldwide. In the Riverina (New South Wales, Australia), characteristic symptoms include low fruit yields, very short shoots and severely stunted roots with black, sunken, necrotic lesions. To determine the cause, roots and wood tissue from affected plants in 20 vineyards (Vitis vinifera cv. Chardonnay grafted to V. champini cv. Ramsey rootstock) were assayed for microbial pathogens. Ilyonectria spp. (I. macrodidyma or I. liriodendra, producers of phytotoxin brefeldin A, BFA, and cause of black foot disease of grapevines) and Botryosphaeriaceae spp. (predominantly Diplodia seriata) were isolated from rootstocks of 100 and 95% of the plants, respectively. Togninia minima and Phaeomoniella chlamydospora (cause of grapevine Petri disease) were isolated from 13 and 7% of affected plants, respectively. All Ramsey rootstock stems of grafted plants sampled from a supplier nursery were infected with Ilyonectria spp. and D. seriata. Diplodia seriata, but not Ilyonectria spp., was also isolated from 25% of canes sampled from the rootstock source block. Root inoculation of potted, disease‐free Chardonnay plants with Ilyonectria isolates from diseased vineyards caused typical disease symptoms, while co‐inoculation with Botryosphaeriaceae spp. increased disease severity. This is the first study to show that a major cause of young grapevine decline can be sequential infection by Botryosphaeriaceae from rootstock cuttings and Ilyonectria spp. from nursery soil. Although the Petri disease fungi were less common in young declining grafted grapevines in the Riverina, they are likely to contribute to the decline of surviving plants as they mature.     

Investigations into Encephalartos insect pests and diseases in South Africa and identification of Phytophthora cinnamomi as a pathogen of the Modjadji cycad

South Africa holds the greatest diversity of Encephalartos species globally. In recent years several reports have been received of Encephalartos species in the country dying of unknown causes. The aim of this study was to investigate the presence of, and identify the causal agents of, diseases of Encephalartos species in the Gauteng and Limpopo Provinces of South Africa. Plant material with symptoms and insects were collected from diseased plants in private gardens, commercial nurseries and conservation areas in these regions. Insects collected were identified based on morphology, and microbial isolates based on morphology and DNA sequence data. Insect species identified infesting cultivated cycads included the beetle Amorphocerus talpa, and the scale insects Aonidiella aurantii, Aspidiotus capensis, Chrysomphalus aonidum, Lindingaspis rossi, Pseudaulacaspis cockerelli, Pseudaulacaspis pentagona and Pseudococcus longispinus. Fungal species isolated from diseased plants included species of Diaporthe, Epicoccum, Fusarium, Lasiodiplodia, Neofusicoccum, Peyronellaea, Phoma, Pseudocercospora and Toxicocladosporium. The plant pathogen Phytophthora cinnamomi was identified from E. transvenosus plants in the Modjadji Nature Reserve. Artificial inoculation studies fulfilled Koch's postulates, strongly suggesting that P. cinnamomi is responsible for the deaths of these plants under field conditions.     

Effects of plant age on disease development and virulence of Clavibacter michiganensis subsp. michiganensis on tomato

The effect of plant age at the time of inoculation on the severity of bacterial wilt and canker disease caused by Clavibacter michiganensis subsp. michiganensis (Cmm) was examined in six greenhouse experiments. The period during which inoculations led to wilt and death of tomato plants was defined. This period, designated ‘window of vulnerability’, ranged from transplanting to the 17‐ to 18‐leaf stage. Plants inoculated after this period expressed disease symptoms but did not wilt or die. No significant changes in disease incidence were observed when leaves of different ages were inoculated. Yield accumulation was significantly reduced in plants inoculated within the window of vulnerability compared with those inoculated after this period. Expression of virulence genes, viz. celA, encoding a secreted cellulase, and the serine protease‐encoding pat‐1, chpC and ppaA, was induced during the early stages after inoculation in plants inoculated within the window of vulnerability. Differences in Cmm population between plants inoculated within and outside of this period were insignificant after the first week post‐inoculation, indicating that differences in disease severity, yield loss and expression of virulence determinants are not correlated with Cmm population level. Results suggest that implementation of precautionary measures during the window of vulnerability to avoid secondary spread of Cmm will have a season‐long effect on plant mortality and may minimize, or even prevent, yield losses.