Molecular characterization,comparison of screening methods,and evaluation of cross‐pathogenicity of black rot (Xanthomonas campestris pv. campestris) strains from cabbage,choy sum,leafy mustard and pak choi from Taiwan

Choy sum (Brassica rapa var. parachinensis), leafy mustard (Brassica juncea) and pak choi (B. rapa var. chinensis) are highly nutritious components of diets in Taiwan and other Asian countries, and bacterial black rot caused by Xanthomonas campestris pv. campestris (Xcc) is a major biotic constraint in these crops. As very little was known about the Xcc strains from these crops in these regions, including their cross‐pathogenicity and aggressiveness on different hosts, Xcc strains were obtained from cabbage (Brassica oleracea var. capitata), choy sum, leafy mustard and pak choi crops in Taiwan. Two previously published PCR‐based assays reliably distinguished the Xcc strains from other Xanthomonas species and subspecies. Phylogenetic analysis based on repetitive sequence‐based PCR assays placed the Xcc strains in a clade distinct from other Xanthomonas species, and also showed host specificity. Although all of the Xcc strains from the different host species were pathogenic on all five Brassica test species in both a detached leaf assay and an intact plant assay, in the intact plant assay they showed differences in virulence or aggression on the different test hosts. The Xcc strains from leafy mustard and pak choi were consistently highly aggressive on all the test host genotypes, but the strains from choy sum and cabbage were less aggressive on leafy mustard and choy sum. The intact plant assay proved more discriminating and reliable than the detached leaf assay for comparing the aggressiveness of Xcc strains on different host genotypes, and so, with the new Xcc strains isolated in this study, will be useful for screening leafy brassica germplasm accessions for resistance to black rot.     

Discrimination of <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> pv. <Emphasis Type="Italic">campestris</Emphasis> races among strains from northwestern Spain by <Emphasis Type="Italic">Brassica</Emphasis> spp. genotypes and rep-PCR

Black rot, caused by Xanthomonas campestris pv. campestris (Xcc) is a severe seedborne disease of Brassica crops around the world. Nine races are recognized, being races 1 and 4 the most aggressive and widespread. The identification of Xcc races affecting Brassica crops in a target area is necessary to establish adequate control measures and breeding strategies. The objectives of this study were to isolate and identify Xcc strains from northwestern Spain by using semi-selective medium and pathogenicity tests, determine the existing races of Xcc in this area by differential series of Brassica spp., and evaluate the use of repetitive DNA polymerase chain reaction-based fingerprinting (rep-PCR) to differentiate among the nine existing Xcc races. Seventy five isolates recovered from infected fields were identified as Xcc. Race-typing tests determined the presence of the following seven pathogen races: 1, 4, 5, 6, 7, 8 and 9. Race 4 was the most frequent in Brassica oleracea and race 6 in Brassica rapa crops, therefore breeding should be focussed in obtaining resistant varieties to both races. Cluster analysis derived from the combined fingerprints showed four groups, but no clear relationship to race, crop or geographical origin was found. Rep-PCR analysis was found not to be a reliable method to discriminate among Xcc races, therefore race typing of Xcc isolates should be done by using the differential series of Brassica spp. genotypes or another alternative approach.     

Multilocus sequence typing analysis of Italian Xanthomonas campestris pv. campestris strains suggests the evolution of local endemic populations of the pathogen and does not correlate with race distribution

Xanthomonas campestris pv. campestris (Xcc) is the causal agent of black rot in Brassicaceae. It is widespread in Italy and severe outbreaks occur under conditions that favour disease development. In this study a multilocus sequence typing approach (MLST) based on the partial sequence of seven loci was applied to a selection of strains representative of the main areas of cultivation and hosts. The aim was to investigate whether the long tradition of brassica crops in Italy has influenced the evolution of different Xcc populations. All loci were polymorphic; 14 allelic profiles were identified of which 13 were unique to Italian strains. Based on the seven loci, the most common genotype within the Italian Xcc strains (AP1) was also the most representative genotype found in worldwide Xcc strains. This genotype was included in a new clonal complex in addition to three other clonal complexes already identified in Xcc populations. The phylogenetic reconstruction using a concatenated dataset of four conserved protein-coding genes, dnaK, fuyA, gyrB and rpoD, showed that the Italian strains belonged to two genetic groups. Physiological races were also investigated for the first time in Italy. The race structure of Xcc was determined by inoculating eight differential Brassica lines belonging to five species and showed that, in Italy, race 4 is the most widespread, followed by races 1 and 6. No correlation was found between allelic profiles, host of isolation, geographical origin and races, although a prevalent race was identified within the same clonal complex.     

Genetic diversity and virulence of Xanthomonas campestris pv. campestris isolates from Brassica napus and six Brassica oleracea crops in Serbia

The present study provides insight into the diversity of 147 Xanthomonas campestris pv. campestris (Xcc) isolates obtained from six Brassica oleracea vegetable crops (broccoli, cabbage, cauliflower, collard greens, kale, kohlrabi) and the winter oilseed rape crop Brassica napus, collected from different regions in Serbia in 2014. The XCF/XCR pathovar-specific primer set was used for fast preliminary identification. In repetitive sequence-based PCR (BOX, ERIC and REP) of all isolates, a higher level of genetic diversity was found in winter oilseed rape isolates compared to isolates from the other hosts. ERIC and REP-PCR showed the highest heterogeneity, with 10 and nine banding patterns, respectively. The REP-PCR results showed the highest correlation (70%) with those obtained with multilocus sequence analysis (MLSA), performed with 10 housekeeping genes (fusA, gap-1, gltA, gyrB1, lacF, lepA, rpoD, dnaK, fyuA and gyrB2). Three distinct phylogenetic groups of winter oilseed rape isolates were detected using MLSA. Two genes, gltA and rpoD, showed the greatest ability to identify and discriminate winter oilseed rape Xcc isolates from isolates of the other six hosts. The lepA gene exhibited specific three-nucleotide changes in sequences of some of the isolates. Results of virulence testing of 18 representative isolates showed statistically significant host–pathogen specialization for Xcc isolates from winter oilseed rape, cauliflower, kale and kohlrabi. In conclusion, oilseed rape isolates are more genetically diverse and show greater specialization to their host in comparison to the rest of the tested isolates from other brassica hosts.     

Characterization of isolates that cause black rot of crucifers in East Africa

A study was conducted in the East African countries of Kenya, Tanzania and Uganda in the months of July and August 2009 with the objectives of assessing the status of black rot and race structure of Xanthomonas campestris pv. campestris in the three countries. Samples infected with black rot were collected from farmers’ fields mainly from Brassica oleracea crops (broccoli, cabbage, cauliflower and kales). A total of 399 farms were surveyed of which 260 were from Kenya, 91 from Tanzania and 48 from Uganda. Following successful isolations, a total of 249 isolates of the causal agent, Xanthomonas campestris pv. campestris were recovered. Pathogenicity of all isolates was confirmed on B. oleracea susceptible cultivars Copenhagen Market F1 and Wirosa F1. Sixty of the 250 isolates were race-typed using a differential set Brassica spp. Only two races, 1 (Kenya and Tanzania) and 4 (Kenya, Tanzania and Uganda) were observed however, another race (5) was observed from one isolate recovered from a B. rapa sample obtained from Tanzania in 2003. Genomic fingerprinting with repetitive-PCR revealed clusters that did not depict significant correlations between isolates and geographical location, isolates and host adaptation or isolates and race. However, it did demonstrate existence of genetic differences within the East African X. campestris pv. campestris population indicating that it is not a similar clonal population of the same genetic background.     

Survival of Xanthomonas campestris pv. campestris in the phyllosphere and rhizosphere of weeds

The phyllosphere and rhizosphere of weeds are important niches for phytobacterial survival. The absence of information in Brazil regarding Xanthomonas campestris pv. campestris (Xcc), the causal agent of black rot in crucifers, motivated this study. Twenty‐six weed species belonging to 14 botanical families were included in field experiments between August 2014 and October 2015. Lepidium virginicum and Raphanus raphanistrum (Brassicaceae) demonstrated great potential for survival of Xcc in the phyllosphere, with the bacterium isolated after 56 and 70 days, respectively. Low variation between maximum and minimum temperatures, high rainfall and high relative humidity at specific times of the year contributed to longer Xcc survival periods in the phyllosphere of some species. Xcc survived in the rhizosphere only in R. raphanistrum, where it was isolated for up to 28 days. No relation was found between climatic factors and survival in the rhizosphere. The data indicate that control of brassicaceous weeds will contribute to the control of black rot.     

Genetic fingerprinting of Iranian Xanthomonas campestris pv. campestris strains inducing black rot disease of crucifers

Northern Iran has one of the largest and most diverse populations of cultivated crucifers in Iran. Symptoms of black rot disease were observed in 40 % of fields. To assess the genetic diversity of Xanthomonas campestris pv. campestris (Xcc) strains, associated with black rot disease, 40 strains were isolated from infected samples of crucifers such as cabbage, radish, cauliflower, turnip and kohlrabi, and were collected from different geographic regions of northern Iran including West and East Azarbayjan and Ardabil provinces. Bacterial strains were characterized by their morphological, biochemical and physiological features and pathogenicity tests. Four races were found in northern Iran (1, 4, 5 and 6) and the majority of the tested strains belonged to either race 4 (45 %) or race 6 (20 %). To examine the distribution of dispersed repetitive DNA, Enterobacterial Repetitive Intergenic Consensus (ERIC), BOX, Repetitive Extragenic Palindromic (REP) and random amplified polymorphic DNA (RAPD) sequences in the genome of Xcc using conserved primers. The different markers produced characteristic banding patterns and the similarity matrices from binary banding data was derived with the similarity for qualitative data program (SIMQUAL). On the basis of the fingerprint patterns generated by the combination data set of both rep-PCR and RAPD, the Xcc strains were differentiated into seven clusters (A–G) at 76 % similarity level. The geographical origin of the Iranian strains does not seem to be correlated with the RAPD and rep-PCR clusters. The clusters seem to be more related to the race of the strains. This is the first study on genetic diversity of Xcc strains inducing black rot disease of crucifers in Iran.     

Application of polyclonal and monoclonal antibodies for the detection of Xanthomonas campestris pv. campestris in crucifer seeds using immunofluorescence microscopy

Polyclonal and monoclonal antibodies (PCAs and MCAs) were tested for the detection ofXanthomonas campestris pv.campestris (Xcc) in cabbage seeds using immunofluorescence microscopy (IF). It was concluded that PCA 94, MCAs 20H6, 2F4, 18G12 and a mixture of MCAs 20H6, 18G12, 2F4 and 16B5 could be used to detect Xcc in seed extracts when 5 min and 2.5 h shaking of seeds are used as extraction methods. The reliability of confirming suspect colonies with MCAs and PCA 94 in IF depended in part on the seed lot tested and the antibody used. Some virulent Xcc strains derived from seed lots, did not react with MCAs 10C5, 2F4, 18G12, 17C12 and 16B5. On the other hand, saprophytic isolates obtained from one seed lot cross-reacted with MCA 17C12 and to a lesser extent with MCAs 2F4, 18G12 and PCA 94. No relationship was found between IF-reactions of Xcc strains using MCAs and reactions of Xcc strains in pathogenicity testing. Xcc andX. c. pv.amoraciae (Xca) could in general not be distinguished on the basis of reactions with MCAs and PCAs. Also in pathogenicity tests Xcc and Xca were hard to distinguish.     

Specificity of polycllonal and monoclonal antibodies for the identification of Xanthomonas campestris pv. campestris

Polyclonal and monoclonal antibodies (PCAs and MCAs), produced to whole cells and flagellar extracts ofXanthomonas campestris pv.campestris (Xcc), respectively, were tested for specificity. In immunofluorescence microscopy (IF) the three PCAs tested, reacted at low dilutions with all Xcc strains, some other xanthomonads and non-xanthomonads. At higher dilutions most cross-reactivity with non-xanthomonad strains disappeared. However, the cross-reactivity with strains ofX. c. pv.vesicatoria (Xcv),X. c. pv.amoraciae (Xca) andX. c. pv.phaseoli var. fuscans (Xcpf) remained.Six MCA-producing cell clones viz. 20H6, 2F4, 18G12, 10C5, 17C12 and 16B5 were selected for specificity tests with an enzyme immunoassay (EIA), IF and a dot-blot immunoassay (DBI). None of the MCAs reacted with all Xcc strains in IF and EIA. In DBI, only MCAs 17C12 and 16B5 reacted with all Xcc strains. All six MCAs tested, cross-reacted in one of either tests with other pathovars ofX. campestris, such as Xcv or Xca. The MCAs were also tested in immunoblotting experiments using total bacterial extracts, cell envelope and flagellar extracts. MCAs 20H6, 2F4, 18G12 and 10C5 reacted with the lipopolysaccharide (LPS) of Xcc. MCAs 16B5 and 17C12 reacted with a 39 kilodalton and a 29 kilodalton protein, respectively.It is concluded that the PCAs and MCAs discussed in this study may be used for routine identification and differentiation of (a group of) Xcc strains. The significance of the cross-reactions with other pathovars ofX. campestris needs to be determined by testing seed lots.     

Biological Control of Black Rot (Xanthomonas Campestris Pv. campestris) of Brassicas with an Antagonistic Strain of Bacillus Subtilis in Zimbabwe

Biological control efficiency of an antagonistic, endophytic strain of Bacillus subtilis (strain BB) was evaluated against three strains of the black rot pathogen, Xanthomonas campestris pv. campestris (Xcc), in four Brassica crops (cabbage, cauliflower, rape and broccoli) grown during three consecutive growing seasons and on two soil types, in two different areas in Zimbabwe. Strain BB controlled the disease caused by strain Xcc B-147 in all Brassica crops during the dry and short rainy seasons. A similar effect was observed in cabbage using the strain Xcc 33908. Biological control was effective in broccoli, but not in cabbage and rape during the main rainy season in clay loam soil and limited biological control effect was still observed when these crops were grown in sandy loam soil. The endophytic colonisation of cabbage roots by strain BB was confirmed by immuno-blotting during the whole growing season. Biological control of black rot with strain BB is discussed in relation to its effect on Xcc strains, Brassica crops and to the effect of weather and soil conditions.     

Characterisation of Xanthomonas campestris pv. campestris isolates from South Africa using genomic DNA fingerprinting and pathogenicity tests

The genetic diversity of Xanthomonas campestris pv. campestris isolates from South Africa was evaluated using 28 isolates obtained from the Johannesburg Fresh Produce Market. Samples were collected from cabbage supplies from farms in Gauteng, Mpumalanga and North West Provinces. Strains were isolated from small sections of infected cabbage leaf samples and cultured on Yeast Dextrose Agar. Isolates identity was confirmed by ELISA and Pathogenicity test. Pathogenicity tests were performed by inoculating leaves of known susceptible cabbage seedlings. Infection symptoms induced could be categorized into three groups, ranging from typical to non-typical black rot symptoms. Four differential Brassica cultivars with known avirulence genes were used for race typing done by spray inoculation. Four races, namely 1, 3, 4 and 6, were identified. Of the 28 isolates, four were identified as race 1, two as race 3, 19 as race 4 and three as race 6. Repetitive DNA polymerase chain reaction-based fingerprinting using Eric- and Box-primers was used to assess the genetic diversity. Generated fingerprints of X. c pv. campestris were relatively similar. Cluster analysis could not strictly group isolates by their geographical origin, suggesting limited diversity of Xanthomonas campestris pv. campestris strains within cabbage producing regions in South Africa.     

Resistance to a highly aggressive isolate of Sclerotinia sclerotiorum in a Brassica napus diversity set

Sclerotinia stem rot (SSR) of oilseed rape (OSR, Brassica napus), caused by Sclerotinia sclerotiorum, is a serious problem in the UK and worldwide. As fungicide‐based control approaches are not always reliable, identifying host resistance is a desirable and sustainable approach to disease management. This research initially examined the aggressiveness of 18 Sclerotinia isolates (17 S. sclerotiorum, one S. subarctica) on cultivated representatives of B. rapa, B. oleracea and B. napus using a young plant test. Significant differences were observed between isolates and susceptibility of the brassica crop types, with B. rapa being the most susceptible. Sclerotinia sclerotiorum isolates from crop hosts were more aggressive than those from wild buttercup (Ranunculus acris). Sclerotinia sclerotiorum isolates P7 (pea) and DG4 (buttercup), identified as ‘aggressive’ and ‘weakly aggressive’, respectively, were used to screen 96 B. napus lines for SSR resistance in a young plant test. A subset of 20 lines was further evaluated using the same test and also in a stem inoculation test on flowering plants. A high level of SSR resistance was observed for five lines and, although there was some variability between tests, one winter OSR (line 3, Czech Republic) and one rape kale (line 83, UK) demonstrated consistent resistance. Additionally, one swede (line 69, Norway) showed an outstanding level of resistance in the stem test. Resistant lines also had fewer sclerotia forming in stems. New pre‐breeding material for the production of SSR resistant OSR cultivars relevant to conditions in the UK and Europe has therefore been identified.     

Identification and Characterisation of Xanthomonas campestris pv. campestris Strains from Tanzania by Pathogenicity Tests, Biolog, rep-PCR and Fatty Acid Methyl Ester Analysis

Black rot, caused by Xanthomonas campestris pv. campestris (Xcc), is a major disease constraint to cabbage production by smallholder farmers in Africa. Variability exists within the pathogen, and yet differentiation of Xcc strains from other closely-related xanthomonads attacking crucifers is often difficult. The Biolog system, fatty acid methyl ester analysis using microbial identification system (MIS), rep-PCR and pathogenicity tests were used to identify and characterise Xcc strains from Tanzania. Great diversity was observed among Xcc strains in their Biolog and rep-PCR profiles. Specific rep-PCR genomic fingerprints were linked to some geographical areas in the country. Most of the Xcc strains were clustered in two groups based on their fatty acid profiles and symptom expression in cabbage although some deviant strains were found. Each of the methods allowed a degree of identification from species, pathovar to the strain level. Biolog and MIS identified all Xcc strains at least to the genus level. Additionally, Biolog identified 47% of Xcc strains to the pathovar and 43% to strain level, whereas MIS identified 43% of the strains to pathovar level. In the absence of a database, the utility of rep-PCR for routine diagnosis of strains was limited, although the procedure was good for delineation of Xcc to the strain level. These findings indicate the existence of Xcc strains in Tanzania that are distinct from those included in Biolog and MIS databases. The limitations noticed warrant continued improvement of databases and inclusion of pathogenicity testing, using universally susceptible cultivars, as an integral part of strain identification.     

Growth of Xanthomonas campestris pv. campestris populations at constant and variable temperatures

Quantitative data were collected to describe the relation between temperature and growth of the cabbage black rot pathogen,Xanthomonas campestris pv.campestris (Xcc). Relative growth rates derived from experiments at constant temperatures were used in dynamic simulation of bacterial population development. The relative growth rates were adequate to simulate growth ofXcc populations at constant temperatures but overestimated growth of populations at variable temperatures. This finding gives rise to the hypothesis, that under field conditions, disease development is slower than is expected on the basis of growth parameters obtained from studies with constant temperatures.     

Phenotypic and phylogenetic studies associated with the crucifer white leaf spot pathogen,Pseudocercosporella capsellae,in Western Australia

Pseudocercosporella capsellae (white leaf spot disease) is an important disease on crucifers. Fifty‐four single‐conidial isolates collected from Brassica juncea (Indian mustard), B. napus (oilseed rape), B. rapa (turnip), and Raphanus raphanistrum (wild radish) across Western Australia were investigated for differences in pathogenicity and virulence using cotyledon screening tests, genetic differences using internal transcribed spacer (ITS) sequencing and phylogenetic analysis, and growth rates on potato dextrose, V8 juice and malt extract agars. All isolates from the four crucifer hosts were pathogenic on the three test species: B. juncea, B. napus and R. raphanistrum, but showed differences in levels of virulence. Overall, isolates from B. juncea, B. napus and B. rapa showed greatest virulence on B. juncea, least on R. raphanistrum and intermediate virulence on B. napus. Isolates from R. raphanistrum showed greatest virulence on B. juncea, least on B. napus and intermediate virulence on R. raphanistrum. Growth and production of a purple‐pink pigment indicative of cercosporin was greatest on malt extract agar and cercosporin production on V8 juice agar was positively correlated with virulence of isolates on B. juncea and B. napus. ITS sequencing and phylogenetic analysis showed that isolates collected from B. napus, B. juncea and B. rapa, in general and with few exceptions, had a high degree of genetic similarity. In contrast, isolates from R. raphanistrum were clearly differentiated from isolate groups collected from Brassica hosts. Pseudocercosporella capsellae reference isolates from other countries generally grouped into a single separate cluster, highlighting the genetic distinctiveness of Western Australian isolates.     

Inference of the phylogenetic diversity and population structure of Xanthomonas campestris affecting Brassicaceae using a multilocus sequence typing‐based approach

Xanthomonas campestris pathovars are widely distributed throughout the globe and have a broad host range, causing severe economic losses in the food and ornamental crucifers markets. Using an approach based on multilocus sequence typing, phylogenetic diversity and population structure of a set of 75 Portuguese and other Xanthomonas campestris isolates from several cruciferous hosts were assessed. Although this population displayed a major clonal structure, neighbour‐net phylogenetic analysis highlighted the presence of recombinational events that may have driven the ecological specialization of X. campestris with different host ranges within the Brassicaceae family. A high level of genetic diversity within and among X. campestris pathovars was also revealed, through the establishment of 46 sequence types (STs). This approach provided a snapshot of the global X. campestris population structure in cruciferous host plants, correlating the existing pathovars with three distinct genetic lineages. Phylogenetic relationships between the founder genotype and remaining isolates that constitute the X. campestris pv. campestris population were further clarified using goeBURST algorithm. Identification of an intermediate link between X. campestris pv. campestris and X. campestris pv. raphani provided new insights into the mechanisms driving the differentiation of both pathovars. Wide geographic distribution of allelic variants suggests that evolution of X. campestris as a seedborne pathogen was not shaped by natural barriers. However, as Portuguese isolates encompass 26 unique STs and this country is an important centre of domestication of Brassica oleracea crops, a strong case is made for its role as a diversification reservoir, most probably through host–pathogen coevolution.     

Detection and identification of the crucifer pathogen, <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> pv. <Emphasis Type="Italic">campestris</Emphasis>, by PCR amplification of the conserved Hrp/type III secretion system gene <Emphasis Type="Italic">hrcC</Emphasis>

The development of a rapid detection method for Xanthomonas campestris pv. campestris (Xcc) in crucifer seeds and plants is essential for high-throughput certification purposes. Here we describe a diagnostic protocol for the identification/detection of Xcc by PCR amplification of fragments from the pathogenicity-associated gene hrcC. Under stringent conditions of amplification, a PCR product of 519 bp from hrcC was obtained from a collection of 46 isolates of Xcc, with the exception of two isolates from radish. No amplicons were obtained from 39 pure cultures of the phytopathogenic bacteria Xanthomonas campestris pv. cerealicola, X. campestris pv. juglandis, X. campestris pv. pelargonii, X. campestris pv. vitians, X. arboricola pv. pruni, X. axonopodis pv. phaseoli, X. axonopodis pv. vesicatoria, X. vesicatoria, Pseudomonas syringae pv. phaseolicola, P. syringae pv. syringae, P. syringae pv. tomato, P. fluorescens, P. marginalis, Pectobacterium atrosepticum, P. carotovorum subsp. carotovorum. In addition, PCR reactions were negative for fifty unidentified environmental isolates purified from the surface of crucifers. The PCR fragment was obtained from four strains previously classified as X. campestris pv. aberrans, X. campestris pv. armorociae, X. campestris pv. barbarae and X. campestris pv. incanae using pathogenicity assays. Our PCR protocol specifically detected Xcc in inoculated leaves, seeds and naturally infected leaves of crucifers.     

Evidence for an expanded host range ofFusarium oxysporum f.sp.raphani

The pathogenicity of four isolates ofFusarium oxysporum obtained from infected cultivated rocket (Eruca vesicaria) and wild (sand) rocket (Diplotaxis tenuifolia) was tested on the following cruciferous hosts: stock, radish, wild and cultivated rockets, and various species in the cabbage tribe: cabbage (Brassica oleracea var.sabauda), cauliflower (Brassica oleracea var.botrytis), Brussels sprouts (Brassica oleracea var.gemmifera), broccoli (Brassica oleracea var.italica), turnip (Brassica rapa var.rapa). The results indicated that isolates ofF. oxysporum from cultivated and wild rocket belong to theforma specialis raphani. The isolates from rocket were pathogenic on cabbage, Brussels sprouts, broccoli, turnip, radish and stock; isolates ofF. oxysporum conglutinans from cabbage and radish, and the isolate ofF. oxysporum f.sp.raphani from rape obtained from the ATCC collection, were pathogenic on both cultivated and wild rocket.     

Flower infection of Brassica oleracea with Xanthomonas campestris pv. campestris results in high levels of seed infection

During seed production, Brassica seed may become infected with Xanthomonas campestris pv. campestris after systemic colonization of plants upon leaf infection, or alternatively, after flower infection. Polytunnel experiments were conducted in 2007 and 2008 to study the relative importance of these colonization routes resulting in seed infection. Cauliflower plants (Brassica oleracea) were spray-inoculated at the 8-leaf stage, after formation of cauliflowers or during flowering, at which stage leaves or blossoms were inoculated. Inoculation at all stages resulted in a relatively high percentage of systemic infection; the average estimated infection incidences for stem base and peduncle infections were 16 % and 19 %, respectively. When seed samples were examined by dilution plating for deep-seated infection following hot water treatment, Xcc was detected in 61 % of the 23 seed samples harvested from plants with inoculated flowers. However, symptom development in seedlings raised from the seeds could not be confirmed in a grow-out test under favourable conditions for Xcc infection at a high RH (>95 %) and a relative high temperature (28 °C). Xcc was not detected in 59 seed samples harvested from leaf-inoculated plants with the exception of one sample from plants inoculated at peduncle formation. In a third polytunnel experiment carried out in 2009, the population dynamics of Xcc on inoculated flowers was investigated. Following spray-inoculation of flowers, 52 % of the flowers were infected with Xcc. During development of siliques, infection incidence decreased slowly and at 56 dpi, 20 % of the superficially disinfected siliques were infected with Xcc. It was estimated that 0.18 % of the seeds was infected and that 1–10 % of the infected siliques contained infected seeds. The implications of these results for control of Xcc in a seed production crop are discussed.     

Transmission and detection of toria [<Emphasis Type="Italic">Brassica rapa</Emphasis> L. subsp. <Emphasis Type="Italic">dichotoma</Emphasis> (Roxb.)] phyllody phytoplasma and identification of a potential vector

Phyllody disease associated with 16SrIX phytoplasma was observed in the range of 4.1–11% in 10 different lines of toria [Brassica rapa L. subsp. dichotoma (Roxb.)] in experimental fields of the Indian Agricultural Research Institute, New Delhi, India during 2008 and 2009. The toria phyllody (TP) phytoplasma was detected in all the symptomatic and 13.3% of asymptomatic toria plants by nested PCR. The phytoplasma was detected in midrib, flower part, siliquae, stem, and root of infected plants as well as seeds. TP was transmitted by grafting and by dodder to toria and nine other rapeseed/mustard species as confirmed by nested PCR. However, symptoms of phytoplasma infection were induced only in toria, yellow sarson [Brassica rapa L. subsp. trilocularis (Roxb.)], brown sarson [Brassica rapa L. subsp. sarson (Prain)], rapeseed (B. napus subsp. oleifera), and rocket or taramira (Eruca sativa) but not in mustard (B. juncea), black mustard (B. nigra), Ethiopian mustard (B. carinata), B. tournefortii and white mustard (Sinapis alba). Transmission of TP phytoplasma to periwinkle (Catharanthus roseus) was successful only through dodder, but no transmission to tomato (Lycopersicon esculentum) or brinjal (Solanum melongena) was found. TP phytoplasma was detected in Laodelpax striatellus, an abundant planthopper in toria fields, which indicates that this planthopper may be a potential vector for TP phytoplasma.