Evaluation of cucumber (<Emphasis Type="Italic">Cucumis sativus</Emphasis>) cultivars grown in Eastern Europe and progress in breeding for resistance to angular leaf spot (<Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">lachrymans</Emphasis>)

Increased occurrence of cucumber angular leaf spot, Pseudomonas syringae pv. lachrymans, has caused significant losses in cucumber, Cucumis sativus, yield in Poland in recent years. These losses necessitated evaluation of the level of resistance in cucumber cultivars of mainly Polish breeding, cultivated in Eastern Europe, and initiation of a breeding programme for resistance to this disease. Screening for resistance was performed on 84 cucumber accessions under growth chamber conditions using a highly aggressive strain of P. syringae pv. lachrymans. Most of the screened accessions were either susceptible or displayed intermediate resistance. The screening resulted in the identification of five F₁ hybrid cultivars moderately resistant to angular leaf spot. The identified F₁ hybrids were self-pollinated up to the F₄ generation. Individuals resistant to angular leaf spot were identified. These individuals can be used as a source of resistance to angular leaf spot in future breeding efforts.     

Isolation of Molecular Markers Linked to the <Emphasis Type="Italic">Cry</Emphasis> Locus Conferring Resistance to <Emphasis Type="Italic">Cucumber mosaic cucumovirus </Emphasis>Infection in Cowpea

We previously demonstrated that cowpea [Vigna unguiculata (L.) Walp.] cultivar Kurodane-Sanjaku contains the Cry gene, which confers resistance against Cucumber mosaic cucumovirus infection. In this paper, randomly amplified polymorphic DNA (RAPD) analysis was carried out to tag the Cry locus. Bulked segregant analysis for RAPD resulted in many polymorphisms in amplified DNA patterns. Candidates were further screened using parental and/or F₂ cowpea DNAs. As a result, we obtained three RAPD markers, D13/E14-350, WA3-850 and OPE3-500, flanking the Cry locus. In addition, we amplified cowpea sequences coding for the putative nucleotide-binding site (NBS). Degenerate primers based on NBS sequences of tobacco N and Arabidopsis RPS2 disease resistance genes were used for polymerase chain reaction, and resultant products were cloned and sequenced. Among eight independent clones, cowpea resistance gene analog (CRGA) 5 showed a distinct polymorphism when used as a probe for restriction fragment length polymorphism analysis against the susceptible cowpea cultivar PI 189375 and a near-isogenic line for the Cry. Linkage analyses of these molecular markers showed that genetic distances of CRGA5, D13/E14-350, WA3-850 and OPE3-500 to the Cry locus were 0.7, 5.2, 11.5 and 24.5 cM, respectively. Received 16 December 1999/ Accepted in revised form 22 March 2000     

Patterns of ascospore release in relation to phoma stem canker epidemiology in England (<Emphasis Type="Italic">Leptosphaeria maculans</Emphasis>) and Poland (<Emphasis Type="Italic">Leptosphaeria biglobosa</Emphasis>)

Experiments over five growing seasons at Rothamsted (1998/99–2002/03), four seasons at Boxworth (1998/99, 1999/2000, 2001/02, 2002/03) in England (Leptosphaeria maculans) and three seasons (1998/99–2000/01) at Poznan in Poland (Leptosphaeria biglobosa) suggest that differences in the development of phoma stem canker epidemics between England and Poland relate to differences in weather patterns between the two countries. The duration of ascospore release was longer in England, where winter weather is mild and wet, than in Poland, where winters are cold and often with snow cover, but there was little difference between two sites in England (Rothamsted and Boxworth). Wetness provided by rainfall was essential for release of ascospores of both L. maculans in England and L. biglobosa in Poland. Temperature did not affect release of ascospores over the range 5–20 °C. Diurnal periodicity in release of ascospores of L. maculans in England and L. biglobosa in Poland was similar. The timing (date) of first release of ascospores of L. maculans or L. biglobosa in autumn was related to rainfall in August and September; with increasing rainfall the date was earlier. The incubation periods from first release of ascospores to first appearance of phoma leaf spots for both L. maculans in England and L. biglobosa in Poland, and from first leaf spots to first stem base canker in England, were described using a thermal time (degree-day) approximation.     

The Problem of Protecting Crops Against Harmful Birds in Poland

From the point of view of Polish agriculture, the rook (Corpus frugilegus L.) causes the most damage. Jackdaws (C. monedula L.) usually forage together with rooks and are equally abundant as a breeding species in the southern and central regions of Poland. Fields under spring oats, barley, wheat, maize, peas and winter wheat are the main feeding grounds of rooks and jackdaws. Unlike the corvine birds, starlings (Sturnus vulgaris L.), in Poland, only cause damage to cherry orchards, although in certain areas they also attack strawberries and currants. The house sparrows (Passer domesticus L.) cause considerable damage to ripening crops of wheat, barley, and sunflowers. Domestic pigeons cause considerable damage to crops of peas, wheat and maize grown within a 50 km radius of a town.     

CCK1, a PMK1-type MAP kinase is required for hyphal growth,pigmentation, conidiation,enzyme activity,osmotic stress response,and pathogenicity in <Emphasis Type="Italic">Corynespora cassiicola</Emphasis>

The aim of this study was to confirm the presence of races in populations of the fungus Venturia inaequalis that are able to overcome specific apple scab resistance gene(s) within the major apple-growing areas of Poland. The monitoring was conducted in six orchards located in the north, centre and south of Poland. The study involved the use of 16 differential genotypes for pathogenicity testing conducted under both greenhouse and orchard conditions. In addition, the occurrence of apple scab on 10 apple cultivars containing the Rvi6 gene was assessed in four organic orchards in central Poland. Apple scab was found on their leaves for the first time in Poland in 2010. The use of differential genotypes containing specific resistance genes suggested that 10 apple scab resistance genes had been overcome by V. inaequalis in the orchards monitored in this study.     

Comparison of leaf proteomes of potato (<Emphasis Type="BoldItalic">Solanum tuberosum</Emphasis> L.) genotypes with ER- and HR-mediated resistance to PVY infection

The Ny-1 and Ry-fsto genes of potato (Solanum tuberosum L.) confer hypersensitive response (HR) and extreme resistance (ER), respectively, to Potato virus Y (PVY). ER-type resistance was also observed in potato plants with both alleles (Ny-1 and Ry-fsto). Using two-dimensional electrophoresis (2-DE), quantitative differences between PVY-infected and non-infected control plants were found for 35, 32 and 15 protein spots identified in leaves of potato cultivar Rywal (possessing Ny-1), and potato tetraploid clones PW 363 (with Ry-fsto) and PB 08–137 (with Ny-1 + Ry-fsto), respectively. We recognized 29, 12 and 21 PVY-induced protein spots involved in qualitative changes in Rywal, PW 363 and PB 08–137 plants, respectively, which were processed and analysed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. A database search indicated that these 62 proteins belong to various functional categories. Their expression was genotype-specific. In the case of cultivar Rywal with HR-mediated resistance, proteins involved in photosynthesis and primary metabolism were the most abundant. For PW 363 and PB 08–137, both with ER–mediated resistance, stress-responsive proteins were the most numerous. Only two proteins – glutamate–glyoxylate aminotransferase 2 (GGAT2) and monodehydroascorbate reductase 5 (MDHAR5) – were identified in all three genotypes. These two proteins are components of the reactive oxygen species (ROS) defence mechanism against pathogens in plants. The present study showed that the differences in PVY-induced proteins in the leaves of Ny-1, Ry-fsto and Ny-1 + R-fsto genotypes do not correspond to the type of gene conferring the resistance or to the observed phenotype.     

Increasing control reliability of <Emphasis Type="Italic">Orobanche cumana </Emphasis>through integration of a biocontrol agent with a resistance-inducing chemical

Fusarium oxysporum Schlecht. f.sp. orthoceras (Appel & Wollenw.) Bilai is a potential biocontrol agent against the root-parasitic weed Orobanche cumana. In pot experiments with different sunflower cultivars, the application of F. oxysporum was combined with a treatment of BTH (Benzo(1,2,3)thiadiazole-7-carbothioic acid S-methyl ester), a product known to induce resistance against O. cumana in sunflower. The combined treatments resulted in highly reliable control and sometimes in increased control level compared to the Fusarium treatment alone. In laboratory experiments, no enhancing effect of BTH on virulence and growth of the fungus was observed. Future experiments should further investigate the basic mechanisms of the effect achieved by combining the two control strategies as well as the transfer of this innovative control approach into field experiments.     

Identification and characterization of pathotypes in <Emphasis Type="Italic">Puccinia horiana</Emphasis>, a rust pathogen of <Emphasis Type="Italic">Chrysanthemum</Emphasis> x <Emphasis Type="Italic">morifolium</Emphasis>

Puccinia horiana is the causal agent of chrysanthemum white rust or Japanese rust. This microcyclic autoecious rust has a quarantine status and can cause major damage in the commercial production of Chrysanthemum x morifolium. Given the international and often trans-continental production of planting material and cut flowers of chrysanthemum and the decreasing availability of registered fungicides in specific regions, breeding for resistance against P. horiana will gain importance and will need to involve the appropriate resistance genes for the pathotypes that may be present. As pathotypes have not been well characterized in this system, the main objective was to build an international collection of isolates and screen these on a large collection of cultivars to identify different pathotypes. Using a robust and high throughput bioassay, we tested 36 selected cultivars with 22 individual single-pustule isolates of P. horiana. The isolates originated from three different continents over 4 different collection years and included some isolates from cultivars previously reported as resistant. In most cases the bioassays resulted in a clear scoring of interaction phenotypes as susceptible or resistant, while in several cases consistent intermediate phenotypes were found, often on specific cultivars. Twenty-four of the cultivars gave a differential interaction phenotype profile. All isolates produced a unique profile, infecting a minimum of 4 and a maximum of 19 differential cultivars. Based on the Person analysis of these profiles, this pathosystem contains at least seven resistance genes (and seven avirulence genes), demonstrating the highly complex race structure in this pathosystem.     

Control of Colorado beetle (Leptinotarsa decemlineata) in Poland1

Leptinotarsa decemlineata (Colorado beetle) was found in Poland for the first time in 1945 and a great invasion of this pest took place in 1950. It now plays a very important role in Polish plant protection, mainly because the acreage of potato in Poland amounts to about 1.8 million ha. The pest is controlled by repeated insecticide applications to a yearly total of 2 million ha, which is the highest amount of insecticides used in Poland. Research has been carried out in Poland on the following subjects: biology and ecology of Colorado beetle, influence of pest appearance and control on the biocenosis of fields, methods of control, biochemical factors influencing the life cycle, forecasts and warning of appearance, artificial diet, resistance of potato cultivars and reaction of leaf blades of potato to the eggs of Colorado beetle. Research is also carried out on chemical control, modernization of pesticide recommendations and resistance of Colorado beetle to pesticides. The influence of biotic and abiotic factors on the reduction of Colorado beetle numbers has been studied. With a view to possible biological control, predatory bugs (Perillus bioculatus and Podisus maculiventris) were introduced into Poland in 1959 and 1979. Research is also conducted on entomopathogenic fungi, parasitic nematodes, protozoans, spiroplasms and Bacillus thuringiensis.     

Phylogenetic relationship and genetic diversity of <Emphasis Type="Italic">Agrobacterium</Emphasis> spp. isolated in Poland based on <Emphasis Type="Italic">gyrB</Emphasis> gene sequence analysis and RAPD

The genetic diversity of 47 strains of Agrobacterium originating from different host plants and geographical locations in Poland, together with 12 strains from other countries was investigated. It was analyzed using RFLP of DNA fragment amplified with primers UP-1 and UP-2r flanking part of gyrB and parE genes, gyrB sequencing and randomly amplified polymorphic DNA (RAPD) technique. On the basis of obtained results, we found the majority of agrobacteria isolated in Poland belong to biovar 2. However, among others, three strains distinct from type strains of all the known Agrobacterium species, were discovered. All three methods showed no correlation between genetic diversity and geographical origin or the host plant of all studied strains but they revealed high diversity of the tested agrobacteria. The highest diversity was observed within strains of biovar 1, whereas those of biovar 2 were found to be the more homogenous group. The topology of the constructed gyrB tree corresponds to topologies of 16S and 23S rDNA trees obtained in this and other studies, but the gyrB tree had deeper branching. In the case of RAPD, it was possible to find a unique DNA fingerprint for almost each strain tested. The gyrB gene appeared to be a good phylogenetic marker with high discrimination power allowing better differentiation between species and strains, whereas the RAPD technique can serve as a tool for single strain typing.     

Validation of molecular markers for resistance among Pakistani chickpea germplasm to races of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">ciceris</Emphasis>

DNA markers in chickpea, targetting resistance genes for different races of Fusarium oxysporum f.sp. ciceris (Foc), have been identified in chickpea, but validation of these markers is essential for effective use in resistance breeding. In view of this, different simple sequence repeats (SSR) markers were analysed in Pakistani germplasm including induced mutants and some local lines. Most of the SSR markers showed good correlation with phenotypic evaluation of genotypes to different races of Foc and may be used effectively in resistance breeding, except those markers for race 3. Markers for race 3 showed deviations from phenotypic data and the reason might be that race 3 is actually Fusarium proliferatum as reported recently and resistance to this race might involve some other major resistance genes. Poor correlation of markers with foc-3 on LG2 in our study and a recent report of independent segregation of foc-2 and foc-3 in near isogenic lines suggested that linkage distances among different resistance genes need further investigation. Moreover three Pakistani mutant lines (97477, CM444/92 and CM368/93) depicted high levels of resistance to Foc races and can be deployed as a valuable source in resistance breeding programmes.     

Breeding <Emphasis Type="Italic">Lupinus albus</Emphasis> for resistance to the root pathogen <Emphasis Type="Italic">Pleiochaeta setosa</Emphasis>

Pleiochaeta root rot (PRR) caused by Pleiochaeta setosa is a serious, widespread fungal disease in lupin crops, especially in Lupinus albus (broad-leaf lupin, or white lupin). PRR resistance is common in the gene pool of L. albus with various landraces from the Mediterranean region being the most resistant, and suitable for use in breeding new cultivars. Heritability of resistance is sufficient to make good gains from selection but only when controlled-environment (CE) screening is used. Field disease nurseries on loamy soil gave much lower heritability of resistance. Field disease nurseries had spatially variable spore counts despite continuous lupin cropping, and this was partly responsible (along with climatic conditions) for their reduced precision compared to tests conducted in a CE. Giving infected L. albus roots a single, most-severe-lesion score on a 0–9 scale was adequate for CE screening but not as precise or discriminating as the more time-consuming method of six scores per root. Replication in CE experiments was reduced to two pots of 16 seedlings each without sacrificing genotype discrimination.     

Over-expression of the Arabidopsis <Emphasis Type="BoldItalic">NPR1</Emphasis> gene in citrus increases resistance to citrus canker

Citrus canker, caused by the bacterial pathogen Xanthomonas citri subsp. citri (Xcc), is a serious leaf and fruit spotting disease affecting many important citrus cultivars including grapefruit and certain sweet oranges. Currently, efficacious and economical disease control measures for highly susceptible citrus cultivars are lacking. Development of commercial cultivars with greater field resistance to citrus canker is the optimum strategy for effective disease management. In this study, we generated transgenic ‘Duncan’ grapefruit (DG) and ‘Hamlin’ sweet orange (Ham) expressing the Arabidopsis NPR1 gene (AtNPR1), which is a key positive regulator of the long-lasting broad-spectrum resistance known as systemic acquired resistance (SAR). Our results indicate that over-expression of AtNPR1 in citrus increases resistance to citrus canker and that the resistance is related with the expression levels of AtNPR1 in the transgenic plants. The line (DG 42-2) with the highest expression level of AtNPR1 was also the most resistant, which developed significant fewer lesions accompanied by a ten-fold reduction in Xcc population. The lesions developed on DG 42-2 were smaller and darker than those on the control and lacked callus formation. These lesion phenotypes resemble those on canker resistant kumquats and canker susceptible citrus trees treated with SAR-inducing compounds. Therefore, over-expression of AtNPR1 in citrus is a promising approach for development of more resistant cultivars to citrus canker.     

The <Emphasis Type="Italic">cyv-2</Emphasis> resistance to <Emphasis Type="Italic">Clover yellow vein virus</Emphasis> in pea is controlled by the eukaryotic initiation factor 4E

The same mutant allele of eukaryotic initiation factor 4E (eIF4E) that confers resistance to Pea seed-borne mosaic virus (sbm-1) and the white lupine strain of Bean yellow mosaic virus (wlv) also confers resistance to Clover yellow vein virus (ClYVV) in pea. The eIF4E genes from several pea lines were isolated and sequenced. Analysis of the eIF4E amino acid sequences from several resistant lines revealed that some lines, including PI 378159, have the same sequence as reported for sbm-1 and wlv. When eIF4E from a susceptible pea line was expressed from a ClYVV vector after mechanical inoculation of resistant PI 378159, the virus caused systemic infection, similar to its effects in susceptible line PI 250438. The resistance to ClYVV in line PI 378159 was characterized through a cross with PI 193835, which reportedly carries cyv-2. Mechanical inoculation of the F1 progeny with ClYVV resulted in no infection, indicating that the resistance gene in PI 378159 is identical to cyv-2 in PI 193835. Furthermore, particle bombardment of pea line PI 193835 with infectious cDNA of ClYVV (pClYVV/C3-S65T) resulted in the same resistance mode as that described for PI 378159. These results demonstrate that the resistance to ClYVV conferred by cyv-2 is mediated by eIF4E and that cyv-2 is identical to sbm-1 and wlv.     

The response of pyriproxyfen-resistant and susceptible Bemisia tabaci Genn (Homoptera: Aleyrodidae) to pyriproxyfen and fenoxycarb alone and in combination with piperonyl butoxide

Pyriproxyfen was effective against susceptible Bemisia tabaci eggs at a LC₅₀ of 0.003 mg litre⁻¹ and against nymphs at 0.02 mg litre⁻¹. In comparison, eggs of a laboratory selected, pyriproxyfen-resistant B tabaci strain, originating in an Israeli greenhouse, exhibited 6500-fold resistance and nymphs exhibited 1100-fold resistance. Eggs and nymphs of a strain from an Israeli sunflower field exhibited 450 and 210-fold resistance in comparison to the susceptible standard. Fenoxycarb was generally less effective than pyriproxyfen against B tabaci eggs and nymphs but was unaffected by pyriproxyfen resistance. Piperonyl butoxide (PB) was antagonistic to pyriproxyfen, and this increased with increasing pyriproxyfen resistance. PB had no effect on the toxicity of fenoxycarb. Collectively, these data imply that the modes of action of pyriproxyfen and fenoxycarb are distinct, despite the structural similarities of these molecules. Possible reasons for the antagonism of PB against pyriproxyfen are discussed. © 1999 Society of Chemical Industry     

Resistance to infection by stealth: <Emphasis Type="Italic">Brassica napus</Emphasis> (winter oilseed rape) and <Emphasis Type="Italic">Pyrenopeziza brassicae</Emphasis> (light leaf spot)

Light leaf spot (Pyrenopeziza brassicae) is an important disease on winter oilseed rape crops (Brassica napus) in northern Europe. In regions where economically damaging epidemics occur, resistance to P. brassicae in commercial cultivars is generally insufficient to control the disease without the use of fungicides. Two major genes for resistance have been identified in seedling experiments, which may operate by decreasing colonisation of B. napus leaf tissues and P. brassicae sporulation. Much of the resistance present in current commercial cultivars is thought to be minor gene-mediated and, in crops, disease escape and tolerance also operate. The subtle strategy of the pathogen means that early colonisation of host tissues is asymptomatic, so a range of techniques and molecular tools is required to investigate mechanisms of resistance. Whilst resistance of new cultivars needs to be assessed in field experiments where they are exposed to populations of P. brassicae under natural conditions, such experiments provide little insight into components of resistance. Genetic components are best assessed in controlled environment experiments with single spore (genetically fixed) P. brassicae isolates. Data for cultivars used in the UK Recommended List trials over several seasons demonstrate how the efficacy of cultivar resistance can be reduced when they are deployed on a widespread scale. There is a need to improve understanding of the components of resistance to P. brassicae to guide the development of breeding and deployment strategies for sustainable management of resistance to P. brassicae in Europe.     

Frequency of Avirulence Alleles in Field Populations of Leptosphaeria maculans in Europe

This paper describes the first large-scale Europe-wide survey of avirulence alleles and races of Leptosphaeria maculans. Isolates were collected from the spring rape cultivar Drakkar, with no known genes for resistance against L. maculans, at six experimental sites across the main oilseed rape growing regions of Europe, including the UK, Germany, Sweden and Poland. Additionally in Poland isolates were collected from cv. Darmor, which has resistance gene, Rlm9. In total, 603 isolates were collected during autumn in 2002 (287 isolates from Germany and the UK) and 2003 (316 isolates from Poland and Sweden). The identity of alleles at eight avirulence loci was determined for these isolates. No isolates had the virulence allele avrLm6 and three virulence alleles (avrLm2, avrLm3 and avrLm9) were present in all isolates. The isolates were polymorphic for AvrLm1, AvrLm4, AvrLm5 and AvrLm7 alleles, with virulence alleles at AvrLm1 and AvrLm4 loci and avirulence alleles at AvrLm7 and AvrLm5 loci predominant in populations. Virulent avrLm7 isolates were found at only one site in Sweden. Approximately 90% of all isolates belonged to one of two races (combinations of avirulence alleles), Av5-6-7 (77% of isolates) or Av6-7 (12%). Eight races were identified, with four races at frequencies less than 1%. The study suggested that Rlm6 and Rlm7 are still effective sources of resistance against L. maculans in oilseed rape in Europe. The results are comparable to those of a similar survey done in France in autumn 2000 and 2001.     

Induction of Systemic Resistance Against Bacterial Wilt in <Emphasis Type="Italic">Eucalyptus urophylla</Emphasis> by Fluorescent <Emphasis Type="Italic">Pseudomonas</Emphasis> spp

The ability of selected strains of fluorescent Pseudomonas spp. to cause induced systemic resistance (ISR) in Eucalyptus urophylla against bacterial wilt caused by Ralstonia solanacearum was investigated. Four of the five strains used can produce salicylic acid (SA) in vitro and, therefore, chemical SA, that is known to induce resistance in many plant species, was used as a reference treatment. Whereas a soil drench with SA did induce systemic resistance in E. urophylla, infiltration of SA into leaves did not. None of the fluorescent Pseudomonas spp. strains caused ISR against bacterial wilt when applied to the soil, but two strains, P. putida WCS358r and P. fluorescens WCS374r triggered ISR when infiltrated into two lower leaves 3–7 days before challenge inoculation. A mutant of strain WCS358r defective in the biosynthesis of the fluorescent siderophore pseudobactin, did not cause ISR, while the purified siderophore of WCS358r did, suggesting that pseudobactin358 is the ISR determinant of WCS358. A siderophore-minus mutant of WCS374r induced the same level of disease resistance as its parental strain, but the purified siderophore induced resistance as well, indicating that both the siderophore and another, unknown, inducing determinant(s) of WCS374r can trigger ISR in Eucalyptus. A possible role of WCS374r-produced SA remains uncertain. Transformation of a siderophore-minus mutant of WCS358 with the SA biosynthetic gene cluster from WCS374 did not enable this transformant to cause ISR in E. urophylla.     

Differences in bark beetle (<Emphasis Type="Italic">Ips typographus</Emphasis> and <Emphasis Type="Italic">Pityogenes chalcographus)</Emphasis> abundance in a strict spruce reserve and the surrounding spruce forests of Serbia

Frequent windthrow in the strict reserves of spruce in the Kopaonik National Park provides much material favorable for the development of Ips typographus (L.) and Pityogenes chalcographus (L.) (Coleoptera, Scolytidae). Therefore it was assumed that the abundance of these bark beetles in the reserves was high and that they dispersed to the surrounding spruce forests. To verify this opinion, their population levels were monitored in the strict reserve Jankove Bare and the adjoining spruce forests over the period 2006–2008. I. typographus parasitoids were identified and the parasitism percent was determined in spruce forests around the reserves. Throughout the monitoring period, the abundance of I. typographus and P. chalcographus was found to be lower in the reserve than in the surrounding spruce forests. Five species of I. typographus parasitoids and a low parasitization percent were identified in spruce forests surrounding the reserve.