A comparison of inoculation techniques for inducing aflatoxin contamination and<Emphasis Type="Italic">Aspergillus flavus</Emphasis> kernel infection on corn hybrids in the field

Aspergillus flavus inoculation techniques were compared on aflatoxin-resistant and -susceptible corn hybrids for inducing aflatoxin contamination andA. flavus kernel infection. A dry carrier technique was comparable to the standard inoculation techniques (the side-needle and a spray technique) in differentiating between the resistant and the susceptible hybrids in the first year of the study. However, only hybrids inoculated with the side-needle technique had statistically different levels of aflatoxin andA. flavus kernel infection in the second year of the study. In a second study, a modified pinbar technique with inoculations near the tip or base of the ear was compared with the side-needle technique. When developing ears were inoculated near the base with the modified pinbar, adequate levels of aflatoxin were induced both years to distinguish between the resistant and susceptible corn hybrids. The modified pinbar technique has the potential of being a useful tool in evaluating corn germplasm for aflatoxin resistance. http://www.phytoparasitica.org posting May 4, 2007.     

Note: Effect of ear bagging systems on<Emphasis Type="Italic">Aspergillus flavus</Emphasis> kernel infection and aflatoxin contamination of corn hybrids grown in the field

Field studies were conducted for 4 years to determine the effect of various ear bagging systems onAspergillus flavus kernel infection and aflatoxin production in developing ears of corn hybrids. Each year, corn hybrids were grown on a Myatt loam (low water-holding capacity) and a Leeper silty clay loam (high water-holding capacity). Corn ears were inoculated withA. flavus using the side-needle technique 7 days after midsilk (50% of the plants in the plot had silks emerged). For the first 2 years, inoculated ears were covered with either white or black paper pollination bags at approximately 14 days after inoculation. During the last 2 years, inoculated ears were covered with either a brown paper pollination bag or a clear plastic zip-lock bag. Daily maximum temperatures were increased 2 to 4°C in all of the bagging systems over ambient temperatures. The bagging systems had a limited effect on aflatoxin production andA. flavus kernel infection in the Leeper silty clay loam. In the Myatt loam, ears covered with plastic bags had higher levels of aflatoxin contamination andA. flavus kernel infection compared with ears covered with paper bags. http://www.phytoparasitica.org posting May 4, 2007.     

Period of susceptibility of almonds to aflatoxin contamination during development in the orchard

Almonds can be contaminated with aflatoxins, produced mainly by Aspergillus flavus and A. parasiticus. Infection can be facilitated by insect injuries during hull split, which begins four to six weeks before harvest. Within this period, it is unknown which kernel stages are most susceptible to aflatoxin contamination. Developing almonds of the Nonpareil cultivar were inoculated weekly with a spore suspension of A. flavus or A. parasiticus for five weeks after hull split in 2013. The almonds were infested with eggs of the lepidopteron navel orangeworm (NOW) (Amyelois transitella) before each spore inoculation. Aflatoxin levels were quantified at harvest using HPLC. Aflatoxin contamination was consistently higher in NOW-damaged kernels, although aflatoxins were also detected in undamaged kernels at each inoculation date. Insect injury is not required for kernel infection but it is a key risk factor for high aflatoxin contamination. Laboratory inoculations were also performed on Nonpareil almond kernels collected during the summers of 2013 and 2015. Aflatoxin levels were significantly lower on dried almonds but the ability to produce aflatoxins was restored when almonds were incubated with high humidity or when the Aspergillus species were inoculated on almond meal agar amended with ground kernels. Therefore, aflatoxins can accumulate in kernels with low a_w, should sufficient moisture favors aflatoxin production. In our field experiment, the orchard micro-climate had sufficient humidity to enable aflatoxin production in both damaged and undamaged dried kernels.     

Evaluating commercial maize hybrids for resistance to gibberella ear rot

An integral component of breeding maize for resistance to Fusarium graminearum ear rot is the identification of resistant genotypes. Since natural infection is not consistent from year to year, maize researchers must use manual techniques to inoculate the plant material with fungal spores. Information is presented here on site resistance of commercial maize hybrids to F. graminearum over three years and at two locations. Additionally, results of an investigation on the two predominant techniques of inoculating maize, the silk channel and kernel inoculation methods, are reported. Of 61 commercial hybrids tested, only two were ranked as moderately resistant to the fungus by both inoculation methods. These two hybrids also had a stable response to the F. graminearum infection across seven environments when the silk channel inoculation method was used. The majority of the hybrids were ranked as either susceptible or highly susceptible and less than 10% of the hybrids had a stable response to fungal infection. In the investigation of methodology, it was concluded that silk browning would be the least laborious way to identify the ideal time to complete silk channel inoculations. It was found that kernel inoculations using the pin inoculation method should take place between 11 and 15 days after 50% silking to achieve proper hybrid discrimination. Mist irrigation increased mold severity ratings and resulted in greater discrimination between hybrids with varying levels of resistance to F. graminearum infection.     

Systemic infection of sorghum and corn by conidia ofSclerospora Sorghi

Conidia ofSclerospora sorghi, obtained from either systemically-infected or local-lesion-infected leaves of sorghum (cv. Vidan), were capable of inducing typical downy mildew systemic infection, including oospore formation, in sorghum and corn hybrids. Very young inoculated seedlings displayed chlorotic systemic symptoms already on the first leaf, and often died at fourth-leaf stage. Systemic infection was induced by conidia on sorghum 1–14 days old at inoculation. Incidence of infection was much higher and symptoms less delayed when the shoot rather than coleorhizas of young sorghum and corn seedlings were inoculated; in two-week-old sorghum with three leaves, inoculation of the coleoptile or of the base of the second and third blades resulted in systemic infection; with coleoptile inoculation partial leaf chlorosis was delayed until the fourth-or fifth-leaf stage, showing that penetration without symptoms had occurred as far as the meristematic tissues of young leaves still within the leaf tube. Conidial inoculation of young sorghum tillers sprouting after cutting down healthy mother shoots resulted in systemic infection. Conidial inoculum is deemed to be the probable major means for systemic infection of corn and sorghum sown in fields in which oospores are not present; inoculation of new tillers of forage sorghum by conidia from infected plants in a neighboring field can explain the rise in numbers of plants systemically stricken. Two sweet corn hybrids — one considered resistant in the field, the other very susceptible — proved equally susceptible when inoculated with conidia at 5 days of age.     

A recessive resistance gene of alfalfa to alfalfa mosaic virus and impact of high ambient temperature on virus resistance

Twenty plants of alfalfa cv. Beaver were screened for resistance to alfalfa mosaic virus (AIMV) severe strain A-515. ELISA screening on both inoculated and apical leaves at fixed temperature (20°C day, 16°C night) suggested the following three types of clonal response to AIMV infection: extremely resistant, AIMV not normally detected from either inoculated or apical leaves; resistant, AIMV detected from inoculated leaves only and did not spread systemically; susceptible, AIMV detected from both inoculated and apical leaves. When plants in the second category were maintained at high temperature (30°C), AIMV was detected from inoculated and apical leaves 6 and 12 days after inoculation, but was not detectable in the apical leaves thereafter. Plants in the first category remained extremely resistant at all temperatures tested. The results of comparative tests using progeny plants of extremely resistant, resistant and susceptible plants, and of their hybrids, suggested that the resistance to AIMV A-515 was controlled by a temperature-dependent recessive gene.     

酿酒高粱品种、组合及亲本的丝黑穗病抗性鉴定

为了明确不同类型的酿酒高粱材料对西南地区丝黑穗病的抗性和促进抗丝黑穗病品种的选育,采用土壤接菌法对107份材料进行了丝黑穗病抗性鉴定;根据抗性鉴定结果,比较了酿酒高粱杂交组合与其亲本对丝黑穗病的抗性,并对供试菌种进行了致病力分析.结果显示:在感病对照恢1植株平均病株率为67.65％时,有13个不育系、6个保持系、10个恢复系、28个杂交组合的病株率为0,达免疫水平,其中,酿酒高粱不育系和杂交组合所占比例较大,分别为86.67％和75.68％;恢复系中免疫类型所占比例较小,为24.39％;常规品种和组合中无免疫类型;杂交高粱对丝黑穗病的抗性属显性遗传;泸州丝黑穗病菌具有较强的致病力.     

Infection of Rice Plants with the Sheath Blight Fungus Causes an Activation of Pentose Phosphate and Glycolytic Pathways

The response of key regulatory enzymes of the pentose phosphate and glycolytic pathways in disease development was assessed in genetically-related rice plants resistant and susceptible to the sheath blight fungus, Rhizoctonia solani. The plants were grown and maintained under greenhouse conditions and inoculated at 50% flowering. Uninoculated healthy plants served as controls. The activities of pentose phosphate pathway enzymes (glucose-6 phosphate dehydrogenase and 6-phosphogluconate dehydrogenase) increased more than two-fold in both the resistant and susceptible plants. Activities of ATP- and pyrophosphate-dependent phosphofructokinase and phosphoenolpyruvate phosphatase increased in infected plants while activity of phosphoenolpyruvate carboxylase in infected plants was lower than in the healthy plants. Furthermore, for enzymes with increased activity, the levels were higher in the resistant line than in the susceptible line. The enhancement of the enzyme activities correlated well with the post infection period. These data suggest that altered carbohydrate metabolism in sheath blight infections may play an important role in modulating the rice plant's response to infection. The isolation of an infection-induced gene encoding a basic enzyme of pentose phosphate and glycolytic pathways could be used to develop plants with more resistance towards sheath blight disease.     

Changes in polygalacturonase,pectin lyase and cellulase in tomato plants,associated withFusarium inoculation

Activities of polygalacturonase, pectin lyase, and cellulase increased in susceptible, catechol-treated, and resistant tomato plants, after inoculation withFusarium oxysporum f.lycopersici (Sacc.) Snyder and Hansen race 2. The catechol-treated and the resistant plants remained symptomless, while susceptible plants developed symptoms of disease. It is therefore suggested that increased activity of cell-wall-degrading enzymes in inoculated plants does not necessarily cause the development of disease symptoms.     

Inheritance of apple proliferation resistance by parental lines of apomictic <Emphasis Type="Italic">Malus sieboldii</Emphasis> as donor of resistance in rootstock breeding

To study inheritance of Malus sieboldii-derived apple proliferation resistance, 14 cross combinations were performed with the tetraploid apomictic M. sieboldii and first and second generation parental lines as donor of resistance and Malus x domestica scion cultivars and apple rootstocks as donor of pomological traits. In the progeny examined mainly three classes were present consisting of mother-like plants with the allele composition of the maternal apomict (ML), hybrids based on fertilization of an unreduced egg cell (hybrid I), and fully recombinant plants (hybrid II). Two-year screening of inoculated plants in the nursery revealed that progeny classes ML and H I responded similarly to infection and that about half of the progeny showed satisfactory resistance. No appropriate resistance was identified in progeny class H II. This might be due to the fact that in fully recombinant offspring M. sieboldii haplotypes have been reduced from 4n to 1-2n or were entirely lost. Following nursery-growing, promising trees were evaluated for six more years in the orchard. Nearly all of them showed satisfactory resistance but were mostly less productive and more vigorous than trees on clonal standard rootstock M9. However, mainly among the offspring of progeny 4608 × M9, resistant genotypes were identified showing pomological properties similar to M9.     

Metabolic profiling in the roots of coffee plants exposed to the coffee root-knot nematode, Meloidogyne exigua

To understand the effect of nematode Meloidogyne exigua infestation on coffee plants, resistant and susceptible coffee seedlings were inoculated with second-stage juveniles of M. exigua, and root metabolites were studied for four time intervals at 0, 24, 48 and 96 h. During this important period for parasite establishment, the concentrations of phenols, carbohydrates, amino acids and alkaloids in the roots were measured, and hydrogen nuclear magnetic resonance spectra of the root extracts were used to identify and quantify other metabolites. One of the most striking changes was the concentration of fumaric acid on resistant plants, which varied from 59 μg g(of root)^?1 to 138 μg g(of root)^?1 during the first 24 h of the nematode inoculation. The same level of variation was observed much later (96 h) in susceptible plants. Similarly, formic and quinic acid concentrations increased more rapidly in the resistant plants compared to the susceptibles. Sucrose concentrations increased to 370 % in the first 48 h in the resistant plants but showed no significant variation in the susceptible plant. Besides, the concentration of alkaloids was much higher at 24 and 48 h in the susceptibles compared to the resistant plants. These results suggest that the higher production of sucrose as well as formic, fumaric and quinic acids, and the lower production of alkaloids by the resistant cultivar in the first 48 h after the nematode inoculation are associated with the resistance of coffee plants to M. exigua.     

小偃6号成株期高温抗条锈性遗传分析

为揭示小偃6号抗病机制和培育持久抗病品种,采用常规杂交分析方法,在小麦抽穗期利用小麦条锈菌小种CYR30、CYR32和Su11-4对小偃6号、铭贤169及其杂交F1、F2、F2∶3接种,平均气温达到21℃时对小偃6号进行了抗条锈性调查和遗传分析。结果显示,接种CYR30、CYR32时,F1代表现高感,F2代群体中抗感分离比例符合1 R∶15 S的理论比例。接种Su11-4时,F1代表现高抗,F2代群体中抗感分离比例符合3R∶1S的理论比例。研究表明小偃6号对CYR30、CYR32的抗病性均由2对隐性基因累加作用控制,对Su11-4的抗病性由1对显性基因控制。     

Analysis of Accumulation Patterns of <Emphasis Type="Italic">Barley yellow dwarf virus-PAV</Emphasis> (BYDV-PAV) in Two Resistant Wheat Lines

Barley yellow dwarf (BYD) is one of the main viral diseases of small-grain cereals. This disease, reported on numerous plant species of the Poaceae family, is caused by a complex of eight viral species including the species Barley Yellow Dwarf Virus-PAV (BYDV-PAV), frequently found in western Europe. Resistance sources against BYDV-PAV are scarce and only identified in perennial Triticineae. Some BYDV-resistant wheat lines have been obtained by introgressing these resistances into bread wheat germplasms. Genetic and biological characterization of the resulting lines has been undertaken. However, little information on the resistant behaviour of these lines during the early stages of the infection process is available. To evaluate the resistance of two genetically distinct resistant lines (Zhong ZH and TC14), 1740 young plantlets, belonging to susceptible reference hosts (barley cv. Express and wheat cv. Sunstar), Zhong ZH or TC14 wheat lines, were inoculated in controlled conditions with French BYDV-PAV isolates. The infection process was monitored during the first 21 days after inoculation (DAI) using a semi-quantitative ELISA. A standardized protocol including five successive samplings of leaves from all inoculated plants and the collection of plant roots at the end of the monitored period was carried out. This protocol enabled an assessment of the infection percentage and the evolution of the viral load in plants from the 7th DAI to the 21st DAI. Statistical analyses of the BYDV infection kinetics using raw ELISA data, a model of the time-dependent variation of the percentage of infected plants and the area under concentration progress curves (AUCPC) demonstrated that Zhong ZH and TC14 lines (1) reduce the development rate of the BYD disease during the first days of infection, (2) decrease the infection efficiency of BYDV-PAV isolates, in the leaves, from 98.7% for susceptible plant genotypes to 81.9% and 71.7% for Zhong ZH and TC14, respectively, (3) reduce the virus load in the leaves of infected plants and (4) are not spared from BYDV infection, as 95.1% of Zhong ZH and 90.2% of TC14 inoculated plants accumulated viral particles in roots and/or in leaves at 21 DAI. These results confirm the BYDV-partial resistant behaviour of both Zhong ZH and TC14 lines. The development rate of the disease was the single parameter that allowed the distinction between the two resistant sources present in the tested lines.     

Pathogenic diversity within field populations of Orobanche cumana and different reactions on sunflower genotypes

Different races of the parasitic Orobanche cumana (sunflower broomrape) have been reported in Spain, race F being the most virulent. Full resistance in sunflower to races A–E is achieved with each of the single major genes Or₁ to Or₅ respectively. However, parasitised hybrids allegedly resistant to race F were observed in early 2002. The purpose of this study was to verify broomrape incidences (BI) on resistant sunflower genotypes, to assess the mixture of races within field populations and to test for partial resistance to race F in the sunflower hybrids showing a low degree of attack (DA) by the weed. Tests were conducted under field conditions in two locations of southern Spain. While no significant differences were found for yield and BI between locations, the DA on the cultivars depended on the location. With high infection levels and significantly lower yield in susceptible controls, marked differences in BI and DA were found within resistant cultivars, but all of them showed similar crop yield. When artificially inoculated with several populations of race F, line P96 and mainly line L86, were consistently slightly infected, suggesting they were inbred lines responsible for horizontal resistance in infested fields. L86 was extremely susceptible to race E populations, which is unusual as sunflower resistance to one race provided resistance to all the previously described races of O. cumana. No different virulences were detected within two groups of subpopulations (races E and F) inoculated onto resistant sunflower genotypes. However, race F subpopulations showed significant differences in aggressiveness, which seems to be related to horizontal (multigenic) resistance of the crop to the parasitic weed.     

Importance of Different Pathways for Maize Kernel Infection by Fusarium moniliforme

ABSTRACT The relative importance of several infection pathways (silks, stalks, and seed) leading to kernel infection of maize hybrids by Fusarium moniliforme was investigated in field experiments in 1993 and 1994. Systemic movement of specific fungal strains within plants was detected by using vegetative compatibility as a marker. Transmission of F. moniliforme from inoculated seed to stalks and developing kernels was detected in two of three field experiments; the seed-inoculated strain was detected in kernels on approximately 10% of ears. The percentage of kernels infected with the seed-inoculated strain ranged from 0 to 70%, with a mean of 0 to 2.5% (0 to 8.3% of F. moniliforme-infected kernels). Other pathways to kernel infection were more effective than seed transmission and systemic infection. F. moniliforme strains inoculated into the crowns and stalks of plants were found throughout the stalks and in up to 95% of the kernels in individual plants. Infection through the silks was clearly the most effective pathway to kernel infection. This was the only inoculation method that significantly increased overall incidence of F. moniliforme infection in kernels; the silk-inoculated strain infected up to 100% of the kernels in individual ears, with a treatment mean as high as 83.7% of kernels. When plants were silk-inoculated, the percentage of kernels infected by other F. moniliforme strains from the seed or stalk was reduced, apparently due to competition among strains. This study provides evidence that systemic development of F. moniliforme from maize seed and stalk infections can contribute to kernel infection, but silk infection is a more important pathway for this fungus to reach the kernels.     

Susceptibility of pure and hybrid willows to isolates ofMelampsora epitea rust

Pure species and F₁ hybrid families ofSalix viminalis andS. dasyclados were tested for resistance to four single uredinium isolates ofMelampsora rust in laboratory experiments using excised leaves. Rust isolates were derived from:S. viminalis, S. dasyclados, aS. viminalis x triandra hybrid, andS. daphnoides. Incidence of infection, number of uredinia per leaf, and numbers of spores per uredinium were measured. As expected, the isolate fromS. daphnoides did not infect any of the willow species or hybrids tested. For the other three rust isolates that were tested, the parent from which the isolate was derived was susceptible, the other parent was resistant, and hybrids were intermediate in resistance for incidence and uredinia per leaf. These patterns indicate additive inheritance of these resistance traits in hybrids. Numbers of spores per uredinium were similar on the hybrids and the susceptible parent for one rust isolate, suggesting dominant inheritance of this trait in the hybrids.     

Differences in the responses of melon accessions to fusarium root and stem rot and their colonization by Fusarium oxysporum f. sp. radicis‐cucumerinum

Fusarium root and stem rot caused by the fungus Fusarium oxysporum f. sp. radicis‐cucumerinum is a major disease in greenhouse cucumbers. Over the past decade, the disease has been documented in melon greenhouses in Greece, and recently it has been sporadically recorded in greenhouse melons in Israel. Variations in disease response were found among 41 melon accessions artificially inoculated with the pathogen: 10 accessions were highly susceptible (90–100% mortality), 23 exhibited an intermediate response (20–86%) and eight were resistant (0–4%). Two melon accessions – HEM (highly resistant) and TAD (partially resistant) – were crossed with the susceptible accession DUL. The responses of the three accessions and F₁ crosses between the resistant and susceptible parents were evaluated. HEM contributed higher resistance to the F₁ hybrid than TAD. Roots of susceptible and resistant accessions were 100 and 79% colonized, respectively, following artificial inoculation. However, only susceptible plants showed colonization of the upper plant tissues. Microscopic evaluation of cross sections taken from the crown region of the susceptible DUL revealed profuse fungal growth in the intercellular spaces of the parenchyma and in xylem vessels. In the resistant cultivar HEM, very little fungal growth was detected in the intercellular spaces of the parenchyma, and none in the xylem or any other vascular tissue. Finding resistant accessions may create an opportunity to study the genetics of resistance inheritance and to develop molecular markers that will facilitate breeding resistant melon cultivars.     

Reaction of three potato cultivars to potato virus S and alteration of miRNAs and mRNA targets in the resistant genotypes

Reactions of three Polish potato cultivars to potato virus S (PVS) were investigated at 22°C. Cultivars Tajfun and Tonacja exhibited partial resistance with systemic infection detected in some inoculated plants; cultivar Bryza was susceptible to PVS with systemic infection detected in all inoculated plants. The virus was not detectable by ELISA at 23 days postinoculation (dpi) but was detected after 40 dpi. Infection rate and viral accumulation were significantly lower in Tonacja and Tajfun than in Bryza, but no statistically significant difference between Tajfun and Tonacja was detected. Both susceptible and resistant genotypes displayed various, either common or cultivar-specific, symptoms. Delayed systemic infection at 56 dpi was observed in some cases in Tonacja and Tajfun. Resistance-related alteration of a set of miRNAs and mRNA targets in the tested cultivars in response to PVS at 22°C exhibited inter- and intracultivar variability. The majority of tested genes were altered only in the partially resistant Tajfun and Tonacja but not in the susceptible Bryza. Enhanced expression of AGO1-2, DCL1, stu-miR482 and its target Gpa2 was observed in Tonacja and plants of Tajfun in which PVS was detected, with the highest induction of Gpa2 in Tajfun (30.2-fold). However, their expression remained unchanged or decreased in plants of Tajfun in which PVS was undetected. Increased expression of stu-miR168a and stu-miR172e was observed in Tonacja and the PVS-undetected plants of Tajfun, respectively, but not in the PVS-detected plants of Tajfun. This is the first report on cultivar-specific alteration of miRNA in a potato–PVS resistance interaction.     

Reduced fusarium ear rot and symptomless infection in kernels of maize genetically engineered for European corn borer resistance

ABSTRACT Field experiments were conducted in 1994, 1995, and 1996 to evaluate the incidence and severity of Fusarium ear rot and the incidence of symp-tomless Fusarium infection in kernels of maize hybrids genetically engineered with Bacillus thuringiensis genes encoding for the delta-endotoxin CryIA(b). Treatments included manual infestation with European corn borer (ECB) larvae and insecticide applications to limit ECB activity to specific maize growth stages or mimic standard ECB control practices. Fusarium symptoms and infection were affected by the specific cryIA(b) transformation used in each hybrid that determines tissue-specific expression of CryIA(b). In hybrids expressing CryIA(b) in kernels, incidence and severity of Fusarium ear rot and incidence of symptomless kernel infection were reduced compared with near-isogenic hybrids lacking cryIA(b) genes. In plants that were manually infested with ECB, ear rot incidence was reduced by 87, 58, and 68%; severity was reduced by 96, 54, and 64%; and incidence of kernel infection by Fusarium species was reduced by 17, 38, and 38% in 1994, 1995, and 1996, respectively. Results were similar in treatments that were not manually infested, but differences between transgenic and nontransgenic hybrids were smaller. Most kernel infection was due to F. moniliforme, F. proliferatum, and F. subglutinans (section Liseola) collectively, and it was within this group that transgenic hybrids exhibited reduced infection. Expression of CryIA(b) in plant tissues other than kernels did not consistently affect Fusarium symptoms or infection. Disease incidence was positively correlated with ECB damage to kernels. Insecticide applications also reduced Fusarium symptoms and infection when applied to nontransgenic plants.