Effects of fungicide on growth of Leptosphaeria maculans and L. biglobosa in relation to development of phoma stem canker on oilseed rape (Brassica napus)

Controlled‐environment and field experiments were done to investigate effects of the fungicide Punch C (flusilazole plus carbendazim) on growth of Leptosphaeria maculans and L. biglobosa in oilseed rape. In controlled‐environment experiments, for plants inoculated with L. maculans, fungicide treatment decreased lesion size and amount of L. maculans DNA in leaves; for plants inoculated with L. biglobosa, fungicide did not affect lesion size or amount of pathogen DNA. When release of ascospores was monitored using a Burkard spore sampler, the timing and pattern of ascospore release differed between the four seasons. In 2006/2007, the majority of ascospores released were L. maculans, whilst in 2007/2008 the majority were L. biglobosa; in both seasons L. maculans ascospores were released before L. biglobosa ascospores. In field experiments in 2002/2003 and 2003/2004, fungicide treatment decreased severity of stem canker on cv. Apex, but gave no significant yield response. In 2006/2007 and 2007/2008, fungicide treatment decreased phoma leaf spot incidence in autumn and stem canker severity at harvest, and increased yield. Fungicide treatment decreased stem canker severity more on cv. Courage, with a good yield response, than on cv. Canberra. In 2002/2003 and 2003/2004, fungicide treatment decreased the frequency of spread of L. maculans into stem pith tissues and in 2006/2007 fungicide decreased the amount of L. maculans DNA in stem tissues (measured by quantitative PCR). These results are used to suggest how effects of fungicides on interactions between L. maculans and L. biglobosa might affect severity of phoma stem canker and yield response.     

Geographic variation in severity of phoma stem canker and <Emphasis Type="Italic">Leptosphaeria maculans/ L. biglobosa</Emphasis> populations on UK winter oilseed rape (<Emphasis Type="Italic">Brassica napus</Emphasis>)

Phoma stem canker, caused by Leptosphaeria maculans and L. biglobosa, is the most important disease of oilseed rape in Europe. Differences between L. maculans and L. biglobosa in their life-cycles enable the two species to co-exist on oilseed rape crops over a cropping season. This review considers the factors affecting geographic variation in the severity of phoma stem canker epidemics and in the structure of the population of the pathogens in the UK, where the most severe epidemics occur in the south of England and cankers do not develop in Scotland. It is concluded that this variation is directly related to differences in climate, since weather-based models show that stem canker severity increases with increasing winter/spring temperature and temperatures are greater in the south of the UK. It may also be related to differences in pathogen populations, since surveys showed that the proportion of the more damaging L. maculans in stem cankers was greatest in southern England, with most L. biglobosa in northern England. Regional variation in agronomic practices such as cultivar choice and fungicide use may also indirectly influence phoma stem canker severity. Differences in cultivar choice result in differences in L. maculans race structure, which may influence the severity of epidemics. Differences in fungicide use may also influence pathogen populations, since L. maculans and L. biglobosa differ in their sensitivities to different azole fungicides. These factors are discussed in relation to strategies for sustainable production of oilseed rape by adaptation to threats posed by climate change.     

Morphological and molecular characterization of <Emphasis Type="Italic">Diaporthe (</Emphasis>anamorph <Emphasis Type="Italic">Phomopsis)</Emphasis> complex and pathogenicity of <Emphasis Type="Italic">Diaporthe aspalathi</Emphasis> isolates causing stem canker in soybean

The complex of Diaporthe (asexual morph) species occurring on soybean constitutes an important pathogenic group associated with diseases such as pod and stem blight, seed decay and stem canker. Stem canker, caused by Diaporthe aspalathi, has been reported as the most aggressive form of canker and its occurrence has limited soybean crop productivity in the southern United States. The main form of pathogen control is the use of stem canker resistant soybean varieties. In this study, strains of Diaporthe and Phomopsis were isolated from stem and seeds of soybean in different locations in South America during the years 1989–2014. Genomic DNA from 26 isolates were analyzed by PCR-restriction fragment length polymorphism (RFLP) and Amplified Fragment Length Polymorphism (AFLP) techniques, and sequencing of internal transcribed spacer (ITS) regions of ribosomal DNA. The molecular analysis of ITS sequences by alignment with those of ex-type strains deposited in GenBank and morphological characteristics allowed the identification of Phomopsis longicolla, D. phaseolorum var. sojae, D. caulivora and D. aspalathi. An analysis of the pathogenicity of 13 isolates of D. aspalathi inoculated in soybean genotypes carrying different resistance genes to stem canker (Rdm1, Rdm2, Rdm3, Rdm4, Rdm5 and Rdm?) enabled us to identify the occurrence of at least three races of D. aspalathi occurring in Brazil. Among the isolates identified as D. aspalathi, both molecular and phenotypic analyses showed clustering depending on the date of collection and pathogenicity, which revealed the existence of variability of the pathogen.     

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.     

Relating plant and pathogen development to optimise fungicide control of phoma stem canker (<Emphasis Type="Italic">Leptosphaeria maculans</Emphasis>) on winter oilseed rape (<Emphasis Type="Italic">Brassica</Emphasis><Emphasis Type="Italic">napus</Emphasis>)

In winter oilseed rape experiments at Rothamsted in 2000/01 to 2002/03 growing seasons, the severity of phoma stem canker epidemics in summer depended on the timing of phoma leaf spot epidemics in the previous autumn, and hence on the timing of Leptosphaeria maculans ascospore release. The first major release of L. maculans ascospores was earlier in 2000 (26 September) and 2001 (18 September) than in 2002 (21 October). Consequently, the autumn phoma leaf spot epidemic was also earlier in 2000 and 2001 than in 2002. The resulting stem canker epidemics were severe by harvest (July) in 2001 and 2002 but not in 2003. No correlation was found between the severity or duration of phoma leaf spotting (lesion days or lesion °C-days) and the subsequent severity of phoma stem canker epidemics. Rates of leaf production and loss were similar in the three growing seasons. Out of ca. 25 leaves produced on plants during each season, leaf numbers 10–14 generally remained on plants for the longest. Treatment with flusilazole + carbendazim in autumn decreased the severity of phoma leaf spotting for several weeks after treatment, decreased the severity of stem canker the following summer and increased yield significantly in 2001 and 2002 but not in 2003. The most effective timings for flusilazole + carbendazim application were when leaves 7–11 were present on most plants and at least 10% of plants were affected by phoma leaf spot. Two half-dose applications of fungicide reduced phoma stem canker and increased yield more than a single full dose application when phoma leaf spot epidemics were early (<800 °C-days after sowing).     

Variations in relative humidity modulate <Emphasis Type="Italic">Leptosphaeria</Emphasis> spp. pathogenicity and interfere with canola mechanisms of defence

Blackleg (phoma stem canker), caused by Leptosphaeria spp., is an important disease of canola (oilseed rape, Brassica napus). Control strategies rely on the use of resistant cultivars, chemical and disease-reducing cropping practices. In Canada, the pathogen population is represented by L. maculans and L. biglobosa, which are considered to be highly and weakly aggressive, respectively. It is largely admitted that L. biglobosa isolates are not able to cause a significant amount of stem canker and develop on the plant only when it becomes senescent, late in the season. The prevalence of L. maculans over L. biglobosa has been considered to be linked to the low aggressiveness of the latter. However, in this study, we show that L. biglobosa isolates could become highly aggressive in terms of lesion appearance on cotyledons, if the right conditions of temperature and relative humidity (RH) are provided. Percent germination of inoculated pycnidiospores was not affected by the RH regimes tested. This is the first study to show the importance of RH as a factor conditioning the pathogenicity of L. biglobosa isolates on canola cotyledons. Concurrent changes in the host defence mechanisms against L. biglobosa isolates in response to variations in the RH were also investigated. Under high RH, the increase in disease caused by the weakly aggressive isolates coincided with a reduced accumulation of lignin at the early stages of infection.     

Soil application of imidacloprid and related SAR-inducing compounds produces effective and persistent control of citrus canker

Soil application of the systemic insecticide imidacloprid (Admire®, Bayer Crop Science) produced season-long control of citrus canker caused by Xanthomonas citri sbsp. citri. Imidacloprid is a neo-nicotinoid that breaks down in planta into 6-chloronicotinic acid, a compound closely related to the systemic acquired resistance (SAR) inducer isonicotinic acid. Potted Swingle citrumelo seedlings (Citrus paradisi × Poncirus trifoliata) were treated with imidacloprid and the SAR inducers, isonicotinic acid, and acibenzolar-s-methyl as soil drenches or with acibenzolar-s-methyl as a foliar spray 1week prior to inoculation of immature leaves with X. citri sbsp. citri. Seedlings were re-inoculated four times over a 24-week period. SAR induction was confirmed by expression of the PR-2 gene (β-1,3 glucanase). Soil drenches of imidacloprid, isonicotinic acid, and acibenzolar-s-methyl induced a high and persistent up-regulation of PR-2 gene expression and reduced the number of canker lesions for up to 24 weeks compared to 4 weeks for foliar acibenzolar-s-methyl. Soil applied inducers of SAR reduced canker lesions up to 70% compared with the untreated inoculated plants. Lesions on leaves were small, necrotic, and flat compared to pustular lesions on inoculated untreated plants. Populations of X. citri sbsp. citri per leaf were reduced 1–3 log units in soil-treated plants compared to inoculated untreated plants.     

World-Wide Importance of Phoma Stem Canker (Leptosphaeria maculans and L. biglobosa) on Oilseed Rape (Brassica napus)

Phoma stem canker is an internationally important disease of oilseed rape (Brassica napus, canola, rapeseed), causing serious losses in Europe, Australia and North America. UK losses of €56M per season are estimated using national disease survey data and a yield loss formula. Phoma stem canker pathogen populations comprise two main species, Leptosphaeria maculans, associated with damaging stem base cankers, and Leptosphaeria biglobosa, often associated with less damaging upper stem lesions. Both major gene and quantitative trait loci mediated resistance to L. maculans have been identified in B. napus, but little is known about resistance to L. biglobosa. Leptosphaeria maculans, which has spread into areas in North America and eastern Europe where only L. biglobosa was previously identified, now poses a threat to large areas of oilseed rape production in Asia. Epidemics are initiated by air-borne ascospores; major gene resistance to initial infection by L. maculans operates in the leaf lamina of B. napus. It is not clear whether the quantitative trait loci involved in the resistance to the pathogen that can be assessed only at the end of the season operate in the leaf petioles or stems. In countries where serious phoma stem canker epidemics occur, a minimum standard for resistance to L. maculans is included in national systems for registration of cultivars. This review provides a background to a series of papers on improving strategies for managing B. napus resistance to L. maculans, which is a model system for studying genetic interactions between hemi-biotrophic pathogens and their hosts.     

Enhanced resistance to citrus canker in transgenic sweet orange expressing the sarcotoxin IA gene

Citrus canker, caused by the bacterial pathogen Xanthomonas citri subp. Citri (Xcc), is a serious disease reported in most citrus-producing areas around the world. Although different levels of field resistance to citrus canker have been reported in sweet oranges, they are usually not sufficient to provide adequate control of the disease. Ectopic over-expression of antibacterial genes is one of the potential strategies to increase plant resistance to bacterial diseases. Previous in vitro results showed that sarcotoxin IA, an antimicrobial peptide isolated from the flesh fly (Sarcophaga peregrina), can be efficient to control different plant pathogenic bacteria, including Xcc. Transgenic “Pera” sweet orange (Citrus sinensis [L.] Osbeck) plants constitutively expressing the sarcotoxin IA peptide fused to the PR1a signal peptide from Nicotiana tabacum for secretion in the intercellular space were obtained by Agrobacterium-mediated transformation using thin sections of mature explants. Citrus canker resistance evaluation in leaves of transgenic and non-transgenic plants was performed through inoculations with Xcc by infiltration and spraying. The Xcc population was up to 2 log unit lower in leaves of the transgenic plants compared to those of non-transgenic controls. Incidence of canker lesions was significantly higher in non-transformed controls (>10 lesions/cm²) than in the transgenic plants (<5 lesions/cm²) after injection infiltration or spraying with Xcc inoculum. Accumulation of sarcotoxin IA peptide in sweet orange tissue did not cause any deleterious effects on the growth and development of the transgenic plants, indicating this approach is suitable to provide resistance to citrus canker.     

Overexpression of <Emphasis Type="Italic">SlARG2</Emphasis> or <Emphasis Type="Italic">SlTD2</Emphasis> in Arabidopsis enhances resistance against <Emphasis Type="Italic">Plutella xylostella</Emphasis> L.

Tomato (Solanum lycopersicum L.) ARGINASE2 (ARG2) and THREONINE DEAMINASE2 (TD2) are involved in plant defense. These enzymes act in the midgut of herbivores fed on tomato plants to degrade the essential amino acids Arg and Thr, respectively. Although it has been demonstrated that overexpression of the SlARG2 gene in tomato enhanced its resistance against M. sexta larvae, knock-down the expression of SlTD2 reduced the resistance of tomato to lepidopteran herbivores; it remains unclear whether overexpression of SlTD2 could enhance the resistance of the host plants to herbivores, or whether combined overexpression of SlARG2 and SlTD2 could lead to synergistically enhanced resistance to insects. Here, we generated transgenic Arabidopsis plants overexpressing SlARG2 (SlARG2 OE) and SlTD2 (SlTD2 OE) individually as well as in combination (SlARG2-SlTD2 OE). Overexpression of these genes did not affect Arabidopsis development, seed yield, or Arg and Thr content. Insect-feeding bioassay was performed by feeding diamondback moth (Plutella xylostella L.) larvae on detached leaves of wild-type, SlARG2 OE, SlTD2 OE, and SlARG2-SlTD2 OE plants. Larvae fed on SlARG2 OE leaves showed approximately 31% to 35% reduction in weight and 6% to 10% reduction in survival rate compared to those fed on wild-type leaves. Although larvae fed on SlTD2 OE leaves showed no reduction in survival rate, they gained less weight. Whereas larvae fed on SlARG2-SlTD2 OE leaves showed neither reduction in weight nor reduction in survival rate. We further investigated the arginase enzymatic activity of the SlARG2 OE and SlARG2-SlTD2 OE transgenic plants. The SlARG2 OE line most resistant to diamondback moth larvae displayed the highest arginase activity. Our data indicate that overexpression of SlARG2 or SlTD2 in Arabidopsis can enhance its resistance against diamondback moth, whereas combined overexpression of SlARG2 and SlTD2 did not generate synergistically increased resistance to diamondback moth.     

Biological control of green mould on <Emphasis Type="Italic">Agaricus bisporus</Emphasis> by a native <Emphasis Type="Italic">Bacillus subtilis</Emphasis> strain from mushroom compost

Fifty bacterial isolates obtained from compost were tested in vitro against the causal agents of green mould in Agaricus bisporus. Isolate B-38 which induced 48.08% in vitro growth inhibition of T. harzianum T54 and 52.25% of T. aggressivum f. europaeum T77 was identified as Bacillus subtilis, based on 16S rDNA sequence and used in mushroom growing room experiments. B. subtilis B-38 did not decrease mycelial growth rate of Agaricus bisporus A15 in mushroom compost in glass tubes. After applying prochloraz-manganese, B. subtilis B-38 and B. subtilis QST 713, no significant differences in BE values among treatments were found concerning both total yield and the weight of healthy mushrooms. Statistical analyses showed that only inoculation significantly influenced the healthy mushroom yield. In plots inoculated with T. harzianum T54 disease incidence was significantly lower after treatments with prochloraz-manganese (11.81%), B. subtilis QST 713 (12.26%) and B. subtilis B-38 (14.19%) compared to the control (28.16%), as well as in plots inoculated with T. aggressivum f. europaeum T77 11.88%, 12.2% and 15.03%, respectively, in comparison with the control (23.47%). Statistically significant differences were not found among the efficacy values of tested bio-fungicides based on B. subtilis and the commercial fungicide prochloraz-manganese suggesting the use of B. subtilis B-38 and B. subtilis QST 713 as good alternatives to chemical fungicides.     

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.     

<Emphasis Type="Italic">Alternaria alternata</Emphasis>: A new seed-transmitted disease of coriander in South Africa

This is the first report of Alternaria leaf spot disease on coriander (Coriandrum sativum L.) in South Africa. Using the agar plate method, Alternaria alternata was isolated from coriander seed lots together with four other fungal genera, which included Aspergillus, Fusarium, Penicillium and Rhizopus. Standard seed germination tests of coriander seed lots infected with seed-borne mycoflora showed a positive correlation with the number of diseased seedlings (r?=?0.239, p?<?0.01). Pathogenicity tests demonstrated that this seed-borne A. alternata was pathogenic on coriander and symptoms on leaves first appeared as small, dark brown to black, circular lesions (<5 mm diam.) that enlarged and coalesced to form dark brown blotches as time progressed. Leaf spot disease was most severe (64%) on wounded leaves inoculated with A. alternata. Re-isolation of A. alternata from diseased coriander plants satisfied the Koch’s postulates, thus confirming it as the causal agent of Alternaria leaf spot disease. Parsimony analysis based on rpb2 (GenBank Accession No. KT895947), gapdh (KT895949) and tef-1α (KT895945) sequences confirmed identity of the Alternaria isolate, which grouped within the A. alternata clade. Alternaria alternata was shown to be transmitted from infected coriander seed to the developing plants.     

Mitigation of Salt Stress Negative Effects on Sweet Pepper Using Arbuscular Mycorrhizal Fungi (AMF), <Emphasis Type="Italic">Bacillus megaterium</Emphasis> and Brassinosteroids (BRs)

A study was conducted at the experimental farm of Faculty of Agriculture, Ain Shams University, Cairo, Egypt, during two successive summer seasons (2014 and 2015) to investigate the effects of arbuscular mycorrhizal fungi (Glomus irradicans 10% w/w), Bacillus megaterium (10?ml/pot) and brassinosteroids (24-EBL, C₂₈H₄₈O_6; 2?µM) on growth, nutrient absorption, chlorophyll, proline content, antioxidant enzymes activity and fruit yield of sweet pepper plants (Capsicum annuum L.) cv. Marconi. Plants were grown under three levels of salinity (0, 25 and 50?mM). The obtained results showed that plants grown under non-saline water (0?mM NaCl), with or without treatments, significantly gave the most vigorous vegetative growth and had the highest fruit yield compared with those grown under salt stress conditions. All anti-salinity treatments (Mycorrhiza, Bacillus and Brassinosteroids) improved growth when compared with untreated plants (control). Plants inoculated with mycorrhiza or treated with brassinosteroids showed better vegetative growth and shoot biomass (total fresh and dry weight per plant), chlorophyll a and b concentrations, antioxidant content expressed as total soluble phenols and proline concentrations at all studied salinity levels followed by plants inoculated with Bacillus megaterium compared with control plants which showed severe growth retardant especially under higher salt concentration (50?mM). Carotenoids concentration increased proportionally with the increase of salinity concentration. The maximum leaf relative water content (LRWC) and lowest values of membrane permeability (MP) were significantly observed with mychorhiza inoculated plants and brassinosteroid application respectively, followed by Bacillus inoculated plants. Antioxidant enzyme activity were highest in plants irrigated with moderate saline water (25?mM) than plants under high salinity irrigation water (50?mM) except polyphenol oxidase (PPO) as compared with unstressed plants (0?mM). Mycorrhizal inoculated plants accumulated higher K and lower Na and Cl followed by plants treated with brassinosteroids and then plants inoculated with Bacillus megaterium. Anti-salinity treatments positively enhanced fruit yield of sweet pepper plants under all salinity stress levels and the highest fruit yield were significantly observed with brassinosteroid application followed by mychorhiza inoculated plants and then Bacillus inoculated plants.     

Factors affecting <Emphasis Type="Italic">Neofuscicoccum ribis</Emphasis> infection and disease progression in blueberry

Botryosphaeria stem blight is an economically important disease of blueberry worldwide. In this study, factors affecting inoculum production, infection and disease progression of Neofusicoccum spp. in blueberries were investigated. Under laboratory conditions conidia of the main three Neofusicoccum species (N. australe, N. parvum and N. ribis) were released from pycnidia at 15–30 °C and under relative humidities (RHs) of 80–100%, with greatest numbers released by N. parvum. The greatest numbers of oozing pycnidia and conidial release occurred at higher temperatures (25–30 °C) and RHs (92–100%). Inoculation of green shoots with different N. parvum and N. ribis conidial concentrations (50 μL of 5 × 10⁴?5 × 10⁶ conidia/ mL) caused 100% incidence but lesion lengths increased with increasing concentrations. Wound age affected N. ribis lesion development, with lesions only observed for 0–7-day-old wounds in soft green shoots and 0–4-day-old wounds for both hard green shoots and trunks. Colonisation length decreased with increasing wound age. Lesions developed on wounded shoots when plants were exposed to 20 or 25 °C and 90 or 100% RH during the early infection processes; and in non-wounded shoots spot-like lesions were observed although N. ribis colonised the stem tissue. Seasons (summer, autumn and winter) had no effect on susceptibility of wounded plants to N. ribis. External lesions only developed in summer-inoculated plants and colonisation length was lower in winter-inoculated plants. Information on host and environmental factors that affect disease development determined by the study will be used to inform the development of control strategies.     

Population features of <Emphasis Type="Italic">Xanthomonas arboricola</Emphasis> pv. <Emphasis Type="Italic">pruni</Emphasis> from <Emphasis Type="Italic">Prunus spp.</Emphasis> orchards in northern Italy

Bacterial leaf/fruit spot and canker of stone fruits, caused by Xanthomonas arboricola pv. pruni, is a recurrent disease in Italy. A set of 23 strains has been isolated in peach and plum orchards in an intensively stone fruit cultivated area located in north-eastern Italy. They were all identified as X. arboricola pv. pruni by means of phytopathological and serological features: hypersensitive reaction on bean pods, pathogenicity test on immature peach or plum fruitlets, identification by immunofluorescence assay and conventional PCR. Phylogenetic analysis based on sequencing of the gyrB housekeeping gene of the isolates showed that they formed a unique clade, well characterised and separated from other xanthomonads. An insight into the genetic population features was attempted by rep-PCR analysis, using the ERIC, REP and BOX primers. The combined rep-PCR fingerprints showed a slight intra-pathovar variation within our isolates, which grouped in five close clusters. Copper resistance has been assessed in vitro for our whole X. arboricola pv. pruni collection, highlighting that two isolates show a level of resistance in vitro up to 200 ppm of copper. Nonetheless, the copLAB gene cluster, present in many other species of Xanthomonads, was not detected in any isolate, confirming the presence of a still unknown mechanism of copper detoxification in our Xanthomonads arboricola pv. pruni tolerant/resistant strains.     

Improved control of black rot of broccoli caused by <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> pv. <Emphasis Type="Italic">campestris</Emphasis> using a bacteriophage and a nonpathogenic <Emphasis Type="Italic">Xanthomonas</Emphasis> sp. strain

We examined the potential for biological control of black rot of broccoli, caused by Xanthomonas campestris pv. campestris (Xcc), using nonpathogenic Xanthomonas sp. strain 11-100-01 (npX) mixed with bacteriophage XcpSFC211 (pXS). Inoculation of intact broccoli plants in greenhouse trials with either npX or pXS did not control black rot. After injured plant inoculation, however, npX alone or npX with pXS significantly controlled black rot. When a mixed suspension of npX with pXS was placed on a membrane filter, then washed with distilled water and air-dried, a substantial amount of pXS adsorbed to the surface of npX. In a field trial, broccoli plants were sprayed with a suspension of npX with pXS, then inoculated with Xcc. A meta-analysis of the results from five field trials showed an integrated risk ratio (IRR, the ratio of disease incidence in inoculated broccoli plants to the incidence in control plants) of 0.69 after treatment with only npX and 0.59 with npX with pXS, indicating that black rot incidence was significantly reduced by each treatment. The difference between these two treatments was also significant. IRR was 1.24 when comparing suppression by npX with pXS and that by basic copper sulfate wettable powder; thus, their control was comparable. The combination of npX with pXS improved the preventive effect against black rot. This is the first report describing that a nonpathogenic Xanthomonas sp. strain mixed with a bacteriophage effectively controlled black rot of broccoli in field trials.     

Increased susceptibility of potato to <Emphasis Type="Italic">Rhizoctonia</Emphasis> diseases in <Emphasis Type="Italic">Potato leafroll virus</Emphasis>-infected plants

In Hokkaido potato fields, tubers produced from the plants with leaf curl symptoms caused by potato leaf roll virus (PLRV) were noted to be more densely covered with Rhizoctonia sclerotia. This observation led us to hypothesize that potato infected with PLRV would have an increased susceptibility to Rhizoctonia solani. To test this hypothesis, in a pot experiment, we inoculated PLRV-infected mother tubers with Rhizoctonia. As a result, PLRV-infected plants produced significantly fewer and smaller tubers than virus-free plants did, suggesting that PLRV-infected plants are more susceptible than virus-free plants to R. solani. Virus-free seed tubers should thus be used to reduce Rhizoctonia diseases.