New evidence of intra-race diversity in Fusarium oxysporum f. sp. dianthi populations based on Vegetative Compatibility Groups

Fusarium oxysporum f. sp. dianthi (Fod) causes vascular wilt, the most important carnation disease worldwide. We have analyzed vegetative compatibility in a collection of Fod isolates, obtained from both soils and carnation plants in the most important growing areas in Spain, by pairing all isolates in all possible combinations. Results showed that isolates of race 1 and race 2 were distributed among three Vegetative Compatibility Groups (VCG) which correlated with the molecular Groups previously described. Isolates of race 1 and race 2 in molecular Group I grouped in VCG 0021, isolates of race 1 type were in VCG 0022, and isolates of race 1 and race 2 in molecular Group II constituted a new VCG (002-), not previously reported. Isolates in each VCG contained the same mating type gene (MAT1-1 or MAT1-2), with the exception of the new VCG 002- that contained both idiomorphs. This work identifies a new VCG in Fod populations and reports for the first time the presence of isolates of race 1 and race 2 in the same VCG.     

Prevalence of and risk factors for bovine respiratory syncytial virus (BRSV) infection in non-vaccinated dairy and dual-purpose cattle herds in Ecuador

A cross-sectional study was carried out to determine the seroprevalence and risk factors associated with bovine respiratory syncytial virus (BRSV) infection in non-vaccinated dairy and dual-purpose cattle herds from Ecuador. A total of 2,367 serum samples from 346 herds were collected from June 2008 to February 2009. A questionnaire, which included variables related to cattle, health, management measures, and the environment, was filled out in each herd. Presence of antibodies against BRSV was analyzed using a commercial indirect ELISA test. A logistic regression model was used to determine risk factors associated with BRSV at herd level. The individual seroprevalence against BRSV in non-vaccinated herds in Ecuador was 80.48% [1,905/2,367; 95% confidence interval (CI)?=?78.9-82.1]. The herd prevalence was 91.3% (316/346; 95% CI?=?88.3-94.3), and the intra-herd prevalence ranged between 25% and 100% (mean, 90.47%). The logistic regression model showed that the existence of bordering cattle farms, the dual-purpose farms, and the altitude of the farm (more than 2,338?m above sea level) were risk factors associated with BRSV infection. This is the first study about BRSV prevalence in Ecuador. It shows the wide spread of the BRSV infection in the country. The risk factors found will help to design effective control strategies.     

The effect of plant extracts as seed treatments to control bacterial leaf spot of tomato in Tanzania

Bacterial leaf spot (BLS) caused by seed-borne xanthomonads is a serious disease of tomato (Solanum lycopersicum L.), causing significant losses in both yield and quality. To identify more effective control measures, we evaluated crude extracts from 84 plant species in in vitro and in planta assays for antibacterial activity against BLS of tomato. In the in vitro assays, 20.2?% of the tested plant extracts totally inhibited growth of bacteria when seed washings from treated seeds were plated on nutrient agar medium. In the in planta assays, 17.8?% of the tested plant extracts reduced BLS incidence by 100?% in tomato seedlings. The most effective seed treatments were obtained with extracts from Aloe vera, Betula pendula, Coffea arabica, Glycyrrhiza uralensis, Juniperus communis, Ocimum basilicum, Quercus robur, Rheum palmatum, Rosmarinus officinalis, Ruta graveolens, Sinapis alba, Yucca schidigera and Salvia officinalis. Seed treatment of tomato with these extracts completely inhibited Xanthomonas perforans in both in vitro and in planta assays. Extracts from A. vera, C. arabica and Y. schidigera were tested three times using tomato seeds of cultivars Tanya, Cal-J and Moneymaker in Tanzania. Treatment of tomato seeds with these extracts had a positive effect on the number of normal seeds and had no effect on seedling vigor, height and weight. These results indicate that plant extracts from A. vera, C. arabica and Y. schidigera are potential candidates for seed treatment against seed-borne xanthomonads of tomato in Tanzania.     

Induction of an antioxidant enzyme system and other oxidative stress markers associated with compatible and incompatible interactions between chickpea (Cicer arietinum L.) and Fusarium oxysporum f. sp.ciceris

To ascertain if active oxygen species play a role in fusarium wilt of chickpea caused by Fusarium oxysporum f. sp. ciceris, the degree of lipid peroxidation (malondialdehyde formation) and the activity levels of diamine oxidase (DAO), an apoplastic H₂O₂-forming oxidase, and several antioxidant enzymes, namely ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR), guaiacol-dependent peroxidase (GPX) and superoxide dismutase (SOD), were determined spectrophotometrically in roots and stems of ‘WR315’ (resistant) and ‘JG62’ (susceptible) chickpea cultivars inoculated with the highly virulent race 5 of the pathogen. Moreover, APX, CAT, GPX and SOD were also analysed in roots and stems by gel electrophoresis and activity staining; and the protein levels of APX and SOD in roots were determined by Western blotting. In roots, infection by the pathogen increased lipid peroxidation and CAT and SOD activities, although such responses occurred earlier in the incompatible compared with the compatible interactions. APX, GPX and GR activities were also increased in infected roots, but only in the compatible interaction. In stems, infection by the pathogen increased lipid peroxidation and APX, CAT, SOD and GPX activities only in the compatible interaction, and DAO activity only in the incompatible one. In general, electrophoregrams agreed with the activity levels determined spectrophotometrically and did not reveal any differences in isoenzyme patterns between cultivars or between infected and non-infected plants. Further, Western blots revealed an increase in the root protein levels of APX in the compatible interaction and in those of SOD in both compatible and incompatible interactions. In conclusion, whereas enhanced DAO activity in stems, and earlier increases in lipid peroxidation and CAT and SOD activities in roots, can be associated with resistance to fusarium wilt in chickpea, the induction of the latter three parameters in roots and stems along with that of APX, GR (only in roots) and GPX (only in stems) activities are rather more associated with the establishment of the compatible interaction.     

Inhibition of pheromone action in Sesamia nonagrioides by Haloacetate analogues

The electrophysiological activity of some halogenated analogues of the major component of the sex pheromone of the corn stalk borer Sesamia nonagrioides Lef. (1) is presented. The analogues comprise a series of fluoro-, chloro- and bromoacetate analogues 4 – 10 as well as trifluoromethyl ketone 11. The fluoro derivatives 4 – 6 displayed remarkable electro-antennogram (EAG) intrinsic activities in comparison with the parent acetate 1, while the remaining analogues elicited significantly lower response. The compounds have also been tested as inhibitors of the sex pheromone perception in EAG and in the field. In the laboratory. fluoro analogues 4 – 6 were better inhibitors than chloro derivatives 7 – 9 , which in turn behaved similarly to the bromoacetate 10 . Trifluoromethyl ketone 11, however, was a poor inhibitor of the pheromone action. In the field, baits of mixtures of compounds 5 – 11 with the corn stalk borer pheromone in 10 :1 ratio inhibited the concomitant attraction of the clover cutworm moth Scotogramma irifolii Rott., while the difluoro analogue 5, trichloroacetate 9 and trifluoromethyl ketone 11 also diminished the number of catches of the armyworm Mythimna unipuncta Haw. The monofluoroacetate 4. trifluoro analogue 6 and bromo derivative 10 significantly disrupted the pheromone action of the corn borer, whereas trifluoromethyl ketone 11 synergistically increased the number of males attracted to the pheromone trap alone. Addition of 11 to baits containing the corn borer pheromone caught S. nonagrioides selectively with regard to the other habitat-sharing species M. unipuncta and S. trifolii.     

Artenspektrum und Befallshäufigkeit von Fusarium spp. in Bt- und konventionellem Mais im Maiszünsler-Befallsgebiet Oderbruch

The European corn borer (Ostrinia nubilalis) is one of the most important pests of maize (Zea mays). Injuries to the plants caused by the larvae of the European corn borer may represent entrance gates for fungal-spores. The cultivation of Bacillus thuringiensis maize (Bt-maize) is one possibility to reduce infestation by the European corn borer. The aim of the present project was to determine and to compare the number of species and the frequency of Fusarium spp. infestation in Bt-maize (cry1Ab) and conventional maize. In 2003, we analysed the Fusarium spp. infestation of samples of chaffed Bt-maize and its isogenic variety on two experimental fields in the Oderbruch region (Germany), an European corn borer infested area. The conventional variety on the first of the experimental fields (previous crop wheat and forking cultivation) showed a small infestation (16%) of Ostrinia nubilalis while in the conventional variety on the second field (previous crop maize and not forking cultivation) the infestation of the European corn borer was almost three times higher (47%). In the conventional variety on both of the experimental fields we found a high Fusarium spp. infestation (70%). Especially species of the section Liseola dominates, among them: F. subglutinans, F. proliferatum und F. verticillioides. The Fusarium infestation in the samples of Bt-maize from the field with previous crop wheat and forking cultivation was just as high as in the conventional variety (70%). The infestation of Fusarium spp. in the samples of Bt-maize from the field with previous crop maize and forking cultivation was more than 20?% lower.     

Assessment of real-time PCR as a method for determining the presence of <Emphasis Type="Italic">Verticillium dahliae</Emphasis> in different Solanaceae cultivars

Real-time PCR was used to detect and quantify Verticillium dahliae and to assess the susceptibility of four Capsicum annuum cultivars (Luesia, Padrón, SCM331 and PI201234) and the Capsicum chinense cv. C118 to this pathogen. The symptoms which developed after infection included stunting and yellowing, and were more acute in the cv. SCM331, which also suffered defoliation in later stages of the disease and in C118, which suffered severe stunting. Quantification of the pathogen DNA in roots 23 and 34 days post-inoculation (dpi) revealed that there were significantly higher amounts of Verticillium dahliae DNA in C118 than in the other cultivars, followed by SCM331, Padrón and PI201234. The lowest amounts of fungal DNA in roots were found in Luesia. In hypocotyls, the highest amounts of fungal DNA were found in SCM331, while Luesia, Padrón and PI201234 had much lower amounts, and C118 had intermediate levels. When a compatible versus an incompatible system was studied, using the near-isogenic tomato lines LA3030 (susceptible) and LA3038 (resistant to V. dahliae), we were able to detect fungal DNA in both lines. As expected, the fungus/plant DNA ratio was lower in LA3038 than in LA3030 and it decreased with time in LA3038. The amount of Verticillium dahliae DNA in the roots of LA3030 remained constant between days 23 and 34 post-inoculation, but increased 10-fold in collars. Finally, when real-time PCR was applied as a diagnostic method to samples from pepper plants, soil and water collected from farms in northwest Spain, we were able to detect V. dahliae DNA in these samples even when symptoms of the disease were not evident.     

Genetic Structure of the Population of Pepino mosaic virus Infecting Tomato Crops in Spain

ABSTRACT The population structure of Pepino mosaic virus (PepMV), which has caused severe epidemics in tomato in Spain since 2000, was analyzed. Isolates were characterized by the nucleotide sequence of the triple gene block and coat protein gene and, for a subset of isolates, a part of the RNA-dependent RNA polymerase gene. The full-length sequence of the genomic RNA of a Solanum muricatum isolate from Peru also was determined. In spite of high symptom diversity, the Spanish population of PepMV mostly comprised highly similar isolates belonging to the strain reported in Europe (European tomato strain), which has been the most prevalent genotype in Spain. The Spanish PepMV population was not structured spatially or temporally. Also, isolates highly similar to those from nontomato hosts from Peru (Peruvian strain) or to isolate US2 from the United States (US2 strain) were detected at lower frequency relative to the European strain. These two strains were detected in peninsular Spain only in 2004, but the Peruvian strain has been detected in the Canary Islands since 2000. These results suggest that PepMV was introduced into Spain more than once. Isolates from the Peruvian and US2 strains always were found in mixed infections with the European tomato strain, and interstrain recombinants were detected. The presence of different strains of the virus, and of recombinant isolates, should be considered for the development of control strategies based on genetic resistance.     

Phenotypic and Genetic Diversity in Strains of Common Blight Bacteria (Xanthomonas campestris pv. phaseoli and X. campestris pv. phaseoli var. fuscans) in a Secondary Center of Diversity of the Common Bean Host Suggests Multiple Introduction Events

ABSTRACT Common bacterial blight (CBB) disease of the common bean (Phaseolus vulgaris) is caused by Xanthomonas campestris pv. phaseoli and the brown-pigmented variant X. campestris pv. phaseoli var. fuscans. CBB first was described in Castilla y León County, Spain, in 1940, and is now a major constraint on common bean production. In this secondary center of diversity of the common bean, large-seeded Andean cultivars predominate, although medium-seeded Middle American cultivars also are grown. Xanthomonad-like bacteria associated with CBB in Castilla y León were characterized on the basis of carbohydrate metabolism, brown pigment production, genetic analyses (repetitive-element polymerase chain reaction [rep-PCR] and random amplified polymorphic DNA [RAPD]) and pathogenicity on cultivars representing the two common bean gene pools (Andean and Middle American). X. campestris pv. phaseoli was more prevalent (80%) than X. campestris pv. phaseoli var. fuscans (20%). Patterns of carbohydrate metabolism of Spanish CBB bacteria were similar to those of known strains; and only X. campestris pv. phaseoli var. fuscans strains utilized mannitol as a sole carbon source. rep-PCR and RAPD analyses revealed relatively little genetic diversity among Spanish X. campestris pv. phaseoli strains, and these strains were placed together with New World strains into a large cluster. Similar to other New World strains, representative Spanish X. campestris pv. phaseoli strains were highly pathogenic on bean cultivars of both gene pools, showing no gene pool specialization such as that found in certain East African strains. Genetic analyses and pathogenicity tests confirmed and extended previous results, indicating that these East African strains represent distinct xanthomonads that independently evolved to be pathogenic on common bean. X. campestris pv. phaseoli var. fuscans strains were more closely related and genetically distinct from X. campestris pv. phaseoli strains. However, two distinct clusters of X. campestris pv. phaseoli var. fuscans strains were identified, one having the most New World strains and the other having the most African strains. Spanish strains were placed in both clusters, but all strains tested were highly pathogenic on bean cultivars of both gene pools. Together, our results are consistent with multiple introductions of CBB bacteria into Spain. These findings are discussed in terms of breeding for CBB resistance and the overall understanding of the genetic diversity and evolution of CBB bacteria.     

Polyphenolics and Antioxidant Capacity of White and Blue Corns Processed into Tortillas and Chips

White and blue corns of Mexican and American origins were lime-cooked to obtain nixtamals with optimal moisture (48–50%) for tortillas and chips. Blue kernels had less bulk density, softer endosperm and, consequently, required less cooking time than the white kernels. The optimum cooking regime for the white kernels was 100°C for 20 min, while the optimum for both pigmented genotypes was 90°C for 0 min (until the lime-cooking solution reached 90°C). Doughs, tortillas, and chips were characterized by total soluble phenolics (TSP), anthocyanins (ACN), and antioxidant capacity (AOX). A dough acidification procedure using fumaric acid (pH 5.2) was assessed as a means to improve TSP, ACN, and AOX retention. The Mexican blue corn had higher AOX (16%) than the American blue genotype, although the latter had a threefold higher TSP content (12.1 g/kg, dwb). Mexican and American blue corns had higher AOX capacity (29.6 and 25.6 μM trolox equivalents [TE]/g dwb), respectively, than the white corn (17.4 μM TE/g). White corns did not have detectable amounts of ACN, while blue Mexican and American kernels contained 342 and 261 mg/kg. Lime cooking had the greatest negative impact on the stability of TSP, ACN, and AOX. However, the acidification reduced ACN, TSP, and AOX losses by 8–23, 3–14, and 4–15%, respectively. Similar ACN losses were observed for both types of blue kernels when processed into nixtamal/dough (47%); however, ACN losses in tortillas and chips manufactured from the American blue genotype were higher (63 and 81%, respectively) than those of Mexican blue corn products (54 and 75%). ACN losses were highly correlated to TSP (r = 0.91) and AOX capacity losses (r = 0.94).