Impact of primary infection site of <Emphasis Type="Italic">Fusarium</Emphasis> species on head blight development in wheat ears evaluated by IR-thermography

The effect of the primary infection site by Fusarium graminearum and F. culmorum within wheat ears on Fusarium head blight (FHB) was investigated under controlled conditions. FHB development was assessed visually and thermographically following inoculation by: (i) spraying ears, or injecting inoculum into spikelets on (ii) tip, (iii) centre and (iv) base of the ears, separately. Fusarium infection significantly increased the temperature span within ears 6 days post inoculation (dpi), especially infections starting at the ear tip. The temperature difference between air and ear was negatively correlated to FHB severity and enabled disease detection even 29 dpi. F. culmorum caused significant higher disease severity neither reflected in the frequency of infected kernels nor in thousand kernel weight (TKW). Spray inoculations had the strongest effect on TKW, whereas tip inoculations had no effect. Centre and base inoculations had intermediate effects on TKW, although FHB levels did not differ with the same trend among inoculation scenarios. The overall low correlations among FHB severity, infected kernels and TKW are explained by the pathogen spread within ears – downwards more than upwards – and the effect on yield formation which is lower for infections of the upper parts of ears. An exponential model showed high goodness of fit for gradients of infected kernels within ears (R ²  ≥ 70) except tip infection with F. culmorum. This study confirmed that FHB is a function of the primary infection site within ears. Thermography was useful to differentiate among infection scenarios and may be applied in breeding for FHB resistance.     

Suppression subtractive hybridization and microarray analysis reveal differentially expressed genes in the <Emphasis Type="Italic">Lr39/41</Emphasis>-mediated wheat resistance to <Emphasis Type="Italic">Puccinia triticina</Emphasis>

Wheat leaf rust caused by Puccinia triticina (Pt) is one of the most severe fungal diseases threatening the global wheat production. The use of leaf rust resistance (Lr) genes in wheat breeding programs is the major solution to solve this issue. Wheat isogenic line carrying the Lr39/41 gene has shown a moderate to high resistance to most of the Pt pathotypes detected in China. In the present study, a typical hypersensitive response (HR) was observed using microscopy in leaves of the Lr39/41 isogenic line inoculated with the avirulent Pt pathotype THTT from 48 h-post inoculation. Two Lr39/41 resistance-associated suppression subtractive hybridization (SSH) libraries with a total of 6000 clones were established. Microarray hybridizations were performed on all obtained SSH clones using RNAs extracted from leaves of the Pt-inoculated and non-inoculated Lr39/41 isogenic lines, and leaves of the Pt-inoculated and non-inoculated Thatcher susceptible lines. Differentially expressed clones were analyzed by significance analysis of microarrays (SAM), followed by further sequencing. A total of 36 Lr39/41-resistance-related differentially expressed genes (DEGs) were identified, many of which had been previously reported to be involved in the plant defense response. The expression levels of eight selected DEGs during different stages of the Lr39/41-mediated resistance were further quantified by a qRT-PCR assay. Several pathogenesis-related (PR) and HR-related genes seem to be crucial for the Lr39/41-mediated resistance. In general, a brief profile of DEGs associated with the Lr39/41-mediated wheat resistance to Pt was drafted.     

Resistance evaluation of differentials and commercial wheat cultivars to stripe rust (<Emphasis Type="Italic">Puccinia striiformis</Emphasis>) infection in hot spot regions of Canada

Stripe rust of wheat, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most important diseases of wheat in Canada. This study presents the results from resistance evaluation of Yr genes and western Canadian wheat cultivars from different milling classes, to natural infection in southern Alberta and British Columbia which are considered hot spots of stripe rust occurrence in Canada, due to proximity to Pacific Northwest of the United States where stripe rust epidemics are frequent. Genes Yr1, Yr5, Yr15, and YrSP were effective in all environments; Yr17 and Yr28, which were earlier reported ineffective to existing stripe rust races at the seedling stage in Canada, were effective at adult plant stages in most of the environments because of warmer climates in southerly locations, a favourable condition for expression of the genes. Yr17 is common in winter wheat cultivars and only reported spring wheat cultivar carrying it is CDC Stanley, which can serve as donor parent in breeding programs. Gene Yr24/26 was not very effective in western prairies although reported as effective in eastern prairies. Residual resistance from combination of defeated genes (Yr3, Yr7, Yr9, Yr27) in some supplementary differentials was observed. Most cultivars carry slow-rusting, pleiotropic adult-plant resistance gene Yr18 and some Yr29, which were effective in some locations. These genes failed to provide complete protection under high disease pressure. Seedling and adult plant resistance genes Yr5, Yr15, Yr17 and Yr18, Yr36, respectively could be good targets for resistance breeding. Stacking adult plant resistance genes with seedling resistance genes can provide durable resistance to stripe rust.     

Virulence profiling of <Emphasis Type="Italic">Xanthomonas oryzae</Emphasis> pv. <Emphasis Type="Italic">oryzae</Emphasis> isolates,causing bacterial blight of rice in India

Bacterial blight (BB) of rice caused by Xanthomonas oryzae pv. oryzae (Xoo), remains a major production constraint in rice cultivation especially in irrigated and rainfed lowland ecosystems in India. The pathogen is highly dynamic in nature and knowledge on pathotype composition among the Xoo population is imperative for designing a scientific resistance breeding program. In this study, four hundred isolates of Xoo collected from diverse rice growing regions of India were analyzed for their virulence and genetic composition. Virulence profiling was carried out on a set of differentials consisting of 22 near isogenic lines (NILs) of IR24 possessing different BB resistance genes and their combinations along with the checks. It was observed that different NILs possessing single BB resistance gene were susceptible to about 59–94% of the Xoo isolates except IRBB 13 (containing BB resistance gene xa13), which showed susceptibility to about 35% of the isolates. Based on the reaction of the Xoo isolates on the differentials, they were categorized into 22 pathotypes. Among the 22 pathotypes, IXoPt-1 and IXoPt-2 were least virulent and IXoPt # 18–22 were highly virulent. Pathotype IXoPt-19 which was virulent on all single BB resistance genes except xa13 constituted the major pathotype (22.5% isolates) and was widely distributed throughout India (16 states). This was followed by pathotype IXoPt-22 (17.25%) which was virulent on all the NILs possessing single BB resistance genes. Molecular analysis was carried out using two outwardly directed primers complementary to sequence of IS1112, a repetitive element of Xoo. A high level of genetic polymorphism was detected among these isolates and the isolates were grouped into 12 major clusters. The data indicated complex nature of evolution of the Xoo pathotypes and there was no strong correlation between pathotypes and genetic clusters as each genetic cluster was composed of Xoo isolates belonging to different pathotypes. The study indicated that none of the single BB resistance genes can provide broad spectrum resistance in India. However, two-gene combinations like xa5 + xa13 and different 3 or 4 genes combination like Xa4 + xa5 + xa13, Xa4 + xa13 + Xa21, xa5 + xa13 + Xa21 and Xa4 + xa5 + xa13 + Xa21 are broadly effective throughout India.     

Evaluation of resistance of current wheat cultivars and breeding lines to stripe rust from three Gorges reservoir area

Nearly 100,000 ha in the Three Gorges Reservoir Area (TGRA) are in wheat production and the area is a junction where wheat stripe rust overwinters and causes epidemics the next spring; thus the area plays a pivotal role in wheat stripe rust epidemics in China. To better understand wheat resistance levels and the application of Yr genes in this area, 116 wheat cultivars (lines) were collected from the TGRA to investigate stripe rust resistance during the 2014–2016 cropping seasons. Seedling resistance evaluation results indicated that only nine accessions (7.8%) were immune or nearly immune to three predominant races of CYR32, CYR33 and PST-V26. In the field evaluation, 51 accessions (43.9%) showed adult-plant resistance, whereas 56 accessions (48.3%) were susceptible. The application of resistant sources focused on ineffective Yr9 (26.7%) and Yr17 (18.9%), and gradual ineffective Yr26 (34.5%), while effective Yr5, Yr10 and Yr15 were absent. Among them, 21 accessions (18.1%) were combined with two resistance genes. Both low resistance and more concentrated use of Yr genes indicated that this region faces a major risk for a wheat stripe rust epidemic. To improve the wheat resistance level in the TGRA, it is important to discover new all-stage resistance resources and diversify resistance resources for breeding.     

Evaluation of the temporal distribution of <Emphasis Type="Italic">Fusarium graminearum</Emphasis> airborne inoculum above the wheat canopy and its relationship with Fusarium head blight and DON concentration

With the aim of unravelling the role of airborne Fusarium graminearum inoculum in the epidemic of Fusarium head blight (FHB) caused by this species in wheat spikes, a network of Burkard air samplers was set up in five wheat fields distributed in Belgium from 2011 to 2013. Each year from April to July, the daily amounts of F. graminearum inoculum above the wheat canopy were quantified using a newly developed TaqMan qPCR assay. The pattern of spore trapping observed was drastically different per year and per location with a frequency of detection between 9 and 66% and a mean daily concentration between 0.8 and 10.2 conidia-equivalent/m³. In one location, air was sampled for a whole year. Inoculum was frequently detected from the wheat stem elongation stage until the end of the harvesting period, but high inoculum levels were also observed during the fall. Using a window-pane analysis, different periods of time around wheat flowering (varying in length and starting date) were investigated for their importance in the relation between airborne inoculum and FHB parameters (FHB severity, frequency of F. graminearum infection and DON). For almost all the combinations of variables, strong and significant correlations were found for multiple window lengths and starting times. Inoculum quantities trapped around flowering were highly correlated with F. graminearum infection (up to R?=?0.84) and DON (up to R?=?0.9). Frequencies of detection were also well correlated with both of these parameters. DON concentrations at harvest could even be significantly associated with the F. graminearum inoculum trapped during periods finishing before the beginning of the anthesis (R?=?0.77). Overall, these results highlight the key role of the airborne inoculum in F. graminearum epidemics and underline the importance of monitoring it for the development of disease forecasting tools.     

Identification,virulence factors characterization,pathogenicity and aggressiveness analysis of <Emphasis Type="Italic">Fusarium</Emphasis> spp., causing wheat head blight in Iran

Fusarium head blight (FHB), mainly caused by Fusarium graminearum species complex (FGSC) and also by other species of this genus, is one of the most destructive cereal diseases with high yield losses and mycotoxin contamination worldwide. The aim of this study was to identify Fusarium species, characterize their virulence factors such as trichothecene genotypes and cell wall degrading enzymes (CWDEs), and also investigate virulence of the isolates obtained from wheat plants with FHB symptoms in Golestan province of Iran. Among 41 isolates tested, 24 were F. graminearum sensu stricto (s.s.), six were F. proliferatum, four were F. culmorum, three isolates belonged to each of F. subglutinans and F. meridionale species and one isolate of F. asiaticum was identified. Among Fusarium isolates, the nivalenol (NIV) genotype could be found more frequently, followed by 3-acetyl deoxynivalenol (3-ADON) and 15-acetyl deoxynivalenol (15-ADON) genotypes. Production of trichothecenes in autoclaved rice cultures was analyzed by gas chromatography (GC) and confirmed by GC–MS. The mean levels of NIV, 3-ADON and 15-ADON produced by Fusarium spp. were 824, 665 and 622 μg kg^?1, respectively. All Fusarium isolates were capable of producing CWDEs, mainly cellulase and xylanase. Lipase and pectinase activities appeared later and at less quantities. In overall, the isolates FH1 of F. graminearum and FH8 of F. proliferatum showed the maximum activity of CWDEs, which was correlated with high level of their virulence and aggressiveness on wheat. On the other hand, correlation was observed between the level and type of trichothecene produced by each isolate and its virulence on wheat. Virulence of trichothecene producing isolates was higher than that of non-trichothecene producing isolates. Our results suggested that CWDEs and trichothecenes, as virulence factors, have considerable roles on virulence and aggressiveness of the pathogen. This is the first report on the effect of trichothecenes and CWDEs on virulence and aggressiveness of Fusarium spp. associated with FHB disease in wheat growing regions of Iran.     

<Emphasis Type="Italic">Fusarium</Emphasis> head blight incidence and detection of <Emphasis Type="Italic">Fusarium</Emphasis> toxins in wheat in relation to agronomic factors

We investigated incidences of Fusarium head blight (FHB) and concentrations of six mycotoxins (deoxynivalenol, nivalenol, 3-acetyldeoxynivalenol, T-2 toxin, HT-2 toxin and zearalenone) in wheat from 2010 to 2013. Field trials were conducted at the Experimental Station of Cultivar Testing in Chrz?stowo, Poland (53^o11’N, 17^o35’E). We examined the effects of four agronomic factors, including pre-crop type (corn, sugar beets and wheat), date of sowing (late autumn: November 8–December 9 or spring: March 29–April 19), fungicidal application (untreated or treated with two applications) and cultivar (Monsun, Cytra), on FHB index (FHBi) and mycotoxin levels in order to minimize the risk of wheat grain contamination by mycotoxins via integrated pest management methods. The dominant Fusarium species observed on wheat heads were F. culmorum, F. avenaceum (Gibberella avenacea) and F. graminearum (Gibberella zeae), at 21.1%, 17.2% and 7.1%, respectively. A monthly rainfall sum of 113.9 mm and a relatively low air temperature (monthly average 15.5 °C) resulted in the highest FHBi in untreated wheat (25.1%). Agronomic factors crucial for the FHB incidence were the pre-crop, fungicidal treatments and cultivar selection. In wheat planted after wheat or corn, the FHBi was higher compared with a pre-crop of sugar beet. A double application of fungicides at BBCH 30–32 with prothioconazole and spiroxamine and at a BBCH 65 with fluoxastrobin and prothioconazole effectively reduced the FHBi and mycotoxin concentrations, respectively, in grain. The cultivar ‘Cytra’ had a greater FHBi (10.4%) than ‘Monsun’ (4.6%), and grain infestations by Fusarium species were also greater in ‘Cytra’, at 16.5%, than in ‘Monsun’, at 11.2%. Untreated cv. Cytra grown after corn in spring produced grains with the highest amounts of the mycotoxins, deoxynivalenol, 3-acetyldeoxynivalenol, zearalenone and HT-2 (605, 103, 17.5 and 5.53 μg/kg, respectively). Total mycotoxin levels in wheat were correlated with five determinants: duration of the period between the end of flowering and the beginning of kernel abscission, FHBi, F. culmorum isolation, G. zeae isolation and Fusarium ratio (FR) as a % of total mould isolations. Although, the mean concentration of mycotoxins in grain did not exceed the maximum permissible values for unprocessed wheat our study suggests necessity to monitor and mitigate FHB risk for susceptible cultivars, when wheat spring sowing follows corn or wheat.     

Trichothecene genotype and genetic variability of <Emphasis Type="Italic">Fusarium graminearum</Emphasis> and <Emphasis Type="Italic">F. cerealis</Emphasis> isolated from durum wheat in Argentina

Fusarium head blight (FHB) is one of the most important fungal diseases affecting wheat worldwide and it is caused mainly by species within the Fusarium graminearum species complex (FGSC). This study evaluated the presence of FGSC in durum wheat from the main growing area in Argentina and analyzed the trichothecene genotype and chemotype of the strains isolated. Also, the genetic variability of the strains was assayed using ISSR markers. Molecular analysis revealed that among the strains isolated and identified morphologically as F. graminearum, there were 14 strains identified as F. cerealis. Also, it revealed that durum wheat grains were mostly contaminated by F. graminearum, being this the only species reported so far, within the FGSC, affecting durum wheat in Argentina. Analysis of molecular variance (AMOVA) indicated a high genetic variability within rather than between F. graminearum populations. All F. graminearum strains presented 15ADON genotype and were able to produce DON while all F. cerealis strains presented the NIV genotype and most of them were able to produce this toxin. The finding of F. cerealis in durum wheat grains indicates the need for investigating if this fungus is the responsible for the NIV contamination found in wheat in Argentina.     

AlgU contributes to the virulence of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">tomato</Emphasis> DC3000 by regulating production of the phytotoxin coronatine

Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), which causes bacterial speck disease of tomato, has been used as a model pathogen to investigate the molecular basis of plant–pathogen interactions. The function of many potential virulence factors encoded in the Pst DC3000 genome and their modes of action are not fully understood. P. syringae is known to produce the exopolysaccharide alginate. Although AlgU, a sigma factor, is known to regulate the expression of genes such as algD related to alginate biosynthesis, the molecular mechanisms of AlgU in the virulence of Pst DC3000 is still unclear. To investigate the function of AlgU and alginate in plant–bacterial pathogen interactions, we generated ΔalgU and ΔalgD mutants. After inoculation with ΔalgU but not ΔalgD, host plants of Pst DC3000 including tomato and Arabidopsis had milder disease symptoms and reduced bacterial populations. Expression profiles of Pst DC3000 genes revealed that AlgU can regulate not only the expression of genes encoding alginate biosynthesis, but also the expression of genes related to type III effectors and the phytotoxin coronatine (COR). We also demonstrated that the ΔalgU mutant showed full virulence in the Arabidopsis fls2 efr1 double mutant, which is compromised in the recognition of PAMPs. Further, the application of COR was able to restore the phenotype of the ΔalgU mutant in the stomatal response. These results suggest that AlgU has an important role in the virulence of Pst DC3000 by regulating COR production.     

<Emphasis Type="Italic">Fusarium oxysporum</Emphasis> and <Emphasis Type="Italic">Fusarium avenaceum</Emphasis> associated with yield-decline of pyrethrum in Australia

Fusarium wilt, one of the destructive diseases of cucumber can be effectively controlled by using biocontrol agents such as Trichoderma harzianum. However, the mechanisms controlling T. harzianum-induced enhanced resistance remain largely unknown in cucumber plants. Here we screened the potent T. harzianum isolate TH58 that could effectively control F. oxysporum (FO). Glasshouse efficacy trials also showed that TH58 decreased disease incidence by 69.7 %. FO induced ROS over accumulation, while TH58 inoculation suppressed ROS over accumulation and improved root cell viability under F. oxysporum infection. TH58 inoculation could reverse the FO-induced cell division block and regulate the proportional distribution of nuclear DNA content through inducing 2C fraction. Moreover, the expression levels of cell cycle-related genes such as CDKA, CDKB, CycA, CycB, CycD3;1 and CycD3;2 in TH58 - pre-inoculated seedlings were up-regulated compared with those infected with FO alone. Taken together, these results suggest that T. harzianum improved plant resistance against Fusarium wilt disease via alterations in nuclear DNA content and cell cycle-related genes expression that might maintain a lower ROS accumulation and higher root cell viability in cucumber seedlings.     

Korean <Emphasis Type="Italic">Brassica oleracea</Emphasis> germplasm offers a novel source of qualitative resistance to blackleg disease

Blackleg disease, caused by the hemibiotrophic fungal pathogen Leptosphaeria maculans, is one of the most devastating disease of Brassica species worldwide. To date, a total of 20 race-specific blackleg resistance (R) genes have been reported and all of those loci are located in either the A or B genomes of various Brassica species. The B. oleracea genome (CC) shares a high ancestral synteny with the A genome of B. rapa, suggesting the presence of qualitative (race specific) resistance to blackleg disease is also possible in B. oleracea germplasm. In the present study the C genome of Korean B. oleracea germplasm was screened for the presence of blackleg R genes. Thirty-two inbred cabbage lines with unknown resistance profiles, along with five control B. napus lines with well-characterised race-specific R genes, were assessed for cotyledon resistance against two L. maculans isolates with known and highly-contrasting avirulence gene (Avr) profiles. Two cabbage accessions were identified which produced a strong resistance when challenged with either isolate, demonstrating the presence of effective blackleg R genes in the cabbage C genome. Additionally, 16 microsatellite markers linked to seven different R genes of the B. napus A genome were converted into markers for their homologous regions on the B. oleracea C genome. These markers were used to screen all B. oleracea lines to assess if the novel C genome R genes were syntenous to known R gene-homologous regions of the A genome. The resistant cabbage lines offer C genome R genes for the protection of B. oleracea varieties against incursion of blackleg disease, as well as novel additional resistance sources for introgression into B. napus and B. carinata breeding material.     

Conidial sporulation of <Emphasis Type="Italic">Stemphylium solani</Emphasis> under laboratory conditions and infectivity of the inoculum produced <Emphasis Type="Italic">in vitro</Emphasis>

Potato virus Y (PVY) is the type-species of the genus Potyvirus, family Potyviridae, being reported as a major tomato (Solanum lycopersicum L.) pathogen in several regions of the world. Pepper yellow mosaic virus (PepYMV) was originally described as a resistance-breaking Potato virus Y (PVY) isolate on Capsicum annuum L. cultivars, and afterwards it was also reported infecting tomatoes in Brazil. In the present work, a search for sources of resistance to both PepYMV and PVY was conducted in a collection of 119 accessions belonging to seven Solanum (section Lycopersicon) species. This germplasm was initially evaluated to PepYMV reaction by mechanical inoculation followed by symptom observations and ELISA. Potential PepYMV resistance sources were identified for the first time in S. habrochaites, S. peruvianum, S. corneliomuelleri, S. chilense, S. pimpinellifolium, and one accession derived from an interspecific cross (S. lycopersicum x S. peruvianum). A sub-group of 24 accessions with negative serology for PepYMV was also challenged with a PVY isolate, followed by serological and molecular detection with universal primers. Solanum habrochaites ‘L.03683’ and ‘L.03684’ were the only accessions found with stable resistance to both viruses. These results confirm S. habrochaites as the most important source of multiple resistance factor(s) to distinct Potyvirus species.     

Molecular and biological characterization of an isolate of <Emphasis Type="Italic">Tomato mottle mosaic virus</Emphasis> (ToMMV) infecting tomato and other experimental hosts in eastern Spain

Fusarium Head Blight is a major disease of wheat and an important contributor to the reduced cultivation of wheat in South Africa, where the crop often is grown under irrigation. We collected Fusarium isolates from 860 Fusarium Head Blight-infected wheat heads in seven irrigated wheat-growing areas of South Africa. Six Fusarium species, i.e., F. chlamydosporum, F. crookwellense, F. culmorum, F. equiseti, F. graminearum and F. semitectum were recovered, three of which, i.e., F. chlamydosporum, F. equiseti and F. semitectum, were not previously associated with Fusarium Head Blight in South Africa. Fusarium graminearum occurred at high frequencies at all seven locations. Based on polymerase chain reaction (PCR) assays of diagnostic sequences, more isolates were predicted to produce deoxynivalenol than nivalenol. Fusarium graminearum (sensu lato) appears to be the primary causal agent of Fusarium Head Blight in irrigated wheat in South Africa, which may not be the case for wheat cultivated under rain-fed conditions. Rotations of irrigated wheat with other graminaceous crops and maize could increase fungal inoculum and disease pressure. The establishment of Fusarium Head Blight in the irrigated wheat region of the country means that resistant lines and alternative agronomic practices are needed to limit disease severity, yield losses and mycotoxin contamination.