Bacterial Leaf Spot of Ivy Caused by Xanthomonas campestris pv. hederae

A bacterial leaf spot disease was observed on Hedera helix (English ivy) and H. canariensis (Algerian ivy) in Japan. The causal agent was identified as Xanthomonas campestris pv. hederae (Arnaud 1920) Dye 1978. Received 13 May 2002/ Accepted in revised form 3 July 2002     

Movement of Xanthomonas campestris pv. vitians in the stems of lettuce and seed contamination

Xanthomonas campestris pv. vitians , the causal agent of bacterial leaf spot of lettuce (BLS), can be seedborne, but the mechanism by which the bacteria contaminates and/or infects lettuce seed is not known. In this study, the capacity of X. campestris pv. vitians to enter and translocate within the vascular system of lettuce plants was examined. The stems of 8- to 11-week-old lettuce plants were stab-inoculated, and movement of X. campestris pv. vitians was monitored at various intervals. At 4, 8, 12 and 16 h post-inoculation (hpi), X. campestris pv. vitians was recovered from 2 to 10 cm above (depending on stem length) and 2 cm below the inoculation site. Xanthomonas campestris pv. vitians was also recovered from surface-disinfested stem sections of spray-inoculated plants. Together, these results are consistent with X. campestris pv. vitians invading and moving systemically within the vascular system of lettuce plants. To investigate the mechanism of seed contamination, lettuce plants at the vegetative stage of growth were spray-inoculated with X. campestris pv. vitians and allowed to develop BLS. Seed collected from these plants had a 2% incidence of X. campestris pv. vitians external colonization, but no bacteria were recovered from within the seed.     

Phenotypic Characteristics of Xanthomonas campestris pv. campestris from Nepal

Twenty strains of Xanthomonas campestris pv. campestris (Xcc) were isolated from two major crucifer-growing valleys, Chitwan and Kathmandu in Nepal and characterized by biochemical and pathogenicity tests. Strains were homogeneous in bacteriological characteristics. The ability of a strain to induce high or low disease severity index (DSI) on three host plants, broccoli, cabbage, and cauliflower, was interpreted as virulence. Strains that were associated with high or low virulence were significantly different (P>0.05). No relationship between virulence and biochemical characteristics was observed.     

Characterization and PCR-based Typing of Xanthomonas campestris pv. vesicatoria from Peppers and Tomatoes in Serbia

During the last two decades bacterial strains associated with necrotic leaf spots of pepper and tomato fruit spots were collected in Serbia. Twenty-eight strains isolated from pepper and six from tomato were characterized. A study of their physiological and pathological characteristics, and fatty acid composition analysis revealed that all of the strains belong to Xanthomonas campestris pv. vesicatoria. Being non-amylolytic and non-pectolytic, pathogenic on pepper but not on tomato, containing lower amounts of fatty acid 15 : 0 ante–iso, the pepper strains were designated as members of the A group of X. campestris pv. vesicatoria. However, the tomato strains hydrolyzed starch and pectate, caused compatible reactions on tomato but not on pepper, had higher percent of 15 : 0 ante–iso fatty acid, and were classified into B phenotypic group and identified as X. vesicatoria. PCR primers were developed which amplified conserved DNA regions related to the hrp genes of different strains of X. campestris pv. vesicatoria associated with pepper and tomato. Restriction analysis of the PCR product resulted in different patterns and enabled grouping of the strains into four groups. When xanthomonads isolated from pepper and tomato in Serbia were analyzed, they clustered into two groups corresponding to the grouping based on their physiological and pathological characteristics. According to the reaction of pepper and tomato differential varieties, the strains from pepper belong to races P7 and P8 and tomato strains belong to the race T2. All strains were sensitive to copper and streptomycin. Advantages and disadvantages of various bacterial spot management practices are discussed.     

芒果细菌性黑斑病菌在不同场所的存活期

选择芒果细菌性黑斑病菌(Xanthomonas campestris pv.mangiferaeindicae,Xcm)抗利福平菌株RifXcm,通过人工模拟接种试验,采用半选择性平板分离、菌落PCR和常规PCR方法,研究该病原细菌在芒果叶片、土壤和水中的存活期,并对其能否作为侵染源进行了评价。结果表明,芒果细菌性黑斑病菌在芒果叶片病斑内可存活5~6个月,是此病发生最主要的侵染源。病原菌在土壤和自然水中的存活期有限,其中以含芒果残体土壤中的病原菌存活期最长(49~63d),但也没有超过3个月,因此年前存在于这些场所的病原菌均不可能成为第2年的初侵染源。     

Rapd Pcr-based differentiation of Xanthomonas campestris pv. phaseoli and Xanthomonas campestris pv. phaseoli var. fuscans

A RAPD PCR-based method was used to differentiate between isolates of Xanthomonas campestris pv. phaseoli and Xanthomonas campestris pv. phaseoli var. fuscans. Using random primer OP-G11, a single, high intensity band of 820 bp was amplified from DNAs of all X. c. pv. phaseoli var. fuscans isolates, while multiple amplification products of varying sizes were generated from X. c. pv. phaseoli DNAs. Whereas RAPD PCR differentiation gave an unambiguous result in under 4 h, standard differentiation by recording the production of a brown pigment by X. c. pv. phaseoli var. fuscans isolates took up to 7 days and showed variation both between isolates and between media. The unequivocal nature of the RAPD PCR method was demonstrated when isolate 408, originally classified as X. c. pv. phaseoli var. fuscans, failed to produce the 820 bp band typical of X. c. pv. phaseoli var. fuscans isolates, and after also failing to produce a brown pigment, was re-classified as X. c. pv. phaseoli.     

Heterogeneity of Xanthomonas campestris pv. hederae Strains from Araliaceous Hosts

ABSTRACT Xanthomonas campestris pv. hederae (synonym X. hortorum pv. hederae) strains (59 total) were collected from plants in the araliaceae family. Strains were isolated from Hedera helix, Schefflera arboricola, Brassaia actinophylla, and Polyscias spp. from Florida, California, Hawaii, and New Zealand. All strains produced yellow mucoid growth; hydrolyzed esculin, starch, casein and gelatin; were pectolytic; produced urease; and grew on minimal media containing asparagine. All bacterial strains were pathogenic on H. helix (English ivy), B. actinophylla (dwarf schefflera), and Polyscias fruticosa (ming aralia). No differences in symptomatology were detected among strains; however, severity of symptoms usually was greatest on the host of origin. In planta growth rates of representative strains isolated from H. helix, B. actinophylla, and Polyscias spp. also were compared among these three hosts. In all cases, populations grew more rapidly when strains were inoculated to their original host species. All 59 bacterial strains were compared by 95-carbon source GN microplate, fatty acid methyl ester (FAME), and restriction fragment-length polymorphisms (RFLP), with the pulse-field gel electrophoresis method, analyses. All three analyses grouped strains into two distinct groups that correlated with the host of origin. Using metabolic profiles, 75% of the H. helix strains were separated from strains isolated from Brassaia and Schefflera and 95% of the Polyscias strains. FAME analysis separated strains into two distinct groups, with 96% of the H. helix strains placed in one group. RFLP analysis placed all of the H. helix and Schefflera strains in one group, as well as 33% of the Brassaia strains, whereas the other group contained all of the Polyscias strains and the remainder of the Brassaia strains. It is apparent that the pathovar hederae is made up of heterogeneous populations that can be separated by biochemical, pathological, genetic, and physiological analyses into two groups that are closely associated with the host of origin.     

Diversity Among Strains of Xanthomonas campestris pv. vitians from Lettuce

ABSTRACT Diversity in host range, pathogenicity, phenotypic characteristics, repetitive extragenic palindromic polymerase chain reaction (rep-PCR) profiles, and sequence of the 16S-23S rDNA spacer region was examined among 44 Xanthomonas strains isolated from lettuce. Forty-two of the strains were divided into two groups, designated A and B. Seventy percent were Group A, and most of the remaining strains including a reference strain (LMG 938) were Group B. Group A strains induced both local and systemic symptoms, whereas Group B strains caused only distinct necrotic spots. Two strains, including the X. campestris pv. vitians type strain, were distinct from the Group A and B strains and were not pathogenic on lettuce. Analysis of fatty acid profiles, serotype, carbon substrate utilization patterns, and protein fingerprints confirmed this grouping. The Group A and B strains also formed two unique clusters (I and II) by rep-PCR profiling that corresponded to the two groups. Direct sequencing of a PCR-amplified DNA fragment (680 bp) from the 16S-23S rDNA spacer region of four representative strains, however, did not differentiate these groups. Serology and rep-PCR fingerprinting can be used to diagnose and identify X. campestris pv. vitians strains, while the other analyses evaluated are useful for strain characterization.     

辣椒细菌性疮痂病病原菌分类、检测及综合防治研究进展

辣椒细菌性疮痂病是一种世界性分布的细菌性病害,该病能引起辣椒严重的产量损失和品质下降。国外特别是美国对该病害研究较早且较深入,国内相关研究几乎是空白。本文主要围绕病原菌的分类、检测和病害综合防治等研究进展做一概述。     

广东水稻白叶枯病菌遗传多样性和小种分化研究

 通过IS-PCR和rep-PCR指纹技术,分析了广东水稻白叶枯病菌(Xanthomonas oryzae pv.oryzae)群体遗传多样性。用2对特异性引物J3和ERIC对114个菌株基因组DNA进行了PCR扩增,分别呈现89和40种谱型,以彼此间的带位相似率达70%为界,J3扩增的谱型被分为11簇,ERIC的谱型被分为8簇。J3的簇1包含89个菌株,占总数的78.07%,ERIC的簇1包含52个菌株,占总数的45.61%,均为优势簇群。群体遗传多样性值J3为0.8919,ERIC为0.8278。上述结果表明,广东水稻白叶枯病菌的遗传多样性较高。全部参试菌株接种于含有不同抗性基因近等基因系及高感品种金刚30共6个鉴别品种,被划分为6个小种(X-gd1,X-gd2,X-gd3,X-gd4,X-gd5和X-gd6),X-gd4出现频率最高,为广东省的优势小种。     

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.     

Bacteriocin-Like Substances from Tomato Race 3 Strains of Xanthomonas campestris pv. vesicatoria

ABSTRACT Tomato race 3 (T3) strains of Xanthomonas campestris pv. vesicatoria are antagonistic in vitro to tomato race 1 (T1) strains of the bacterium. All T1 strains and 11 strains of other X. campestris pathovars tested were inhibited by T3 strains. Sensitivity of tomato race 2 (T2) strains was variable. No strains from other bacterial genera tested were inhibited. Cell-free filtrates from T3 strains were inhibitory to sensitive strains. The inhibitory activity of these filtrates was lost after treatment at temperatures above 80 degrees C and with selected protease enzymes. However, treatment with trypsin or DNase had no effect on their activity. Seven cosmid clones from a genomic library of a T3 strain were selected for their ability to consistently inhibit a sensitive indicator strain in plate assays. Southern hybridization analysis placed these into three bacteriocin (BCN)-producing groups designated BCN-A, BCN-B, and BCN-C. The BCN-like groups could be differentiated by variations in inhibitory spectra and levels of activity in plate assays. Mutations that inactivated expression of each BCN group individually in a wild-type T3 strain had inhibitory activity confirming that multiple BCNs are present in the T3 strain. T3 strains were inhibitory to a sensitive indicator strain in tomato leaf tissue, but this effect was observed only when T3 strains were applied in advance of the sensitive strain. BCN-A was the major BCN-like substance involved in the suppression of the sensitive indicator strain in tomato leaf tissues.     

Differentiation of Xanthomonas campestris pv. Phaseoli from Xanthomonas campestris pv. Phaseoli var. Fuscans by PFGE and RFLP

Xanthomonas campestris pv. phaseoli and X. campestris pv. phaseoli var. fuscans, the causal agents of the common and fuscous bacterial blight of beans, appear to be phenotypically identical except that the latter can produce a melanin-like pigment in culture. Ten isolates of X. campestris pv. phaseoli and 12 isolates of X. campestris pv. phaseoli var. fuscans were examined using pulsed-field gel electrophoresis (PFGE) and restriction fragment length polymorphism (RFLP). The average genome sizes for X. campestris pv. phaseoli and X. campestris pv. phaseoli var. fuscans were 3850.6±48.9 and 3584.3±68.1kb respectively. The genetic relatedness of the isolates was determined from macrorestriction patterns generated using XbaI. Cluster analysis indicated that the non-fuscous and fuscous strains are distinct. RFLP results, based on the highly conserved hrp genes and a pectate lyase gene from Xanthomonas, also indicated that the two bacteria are genetically different. The results obtained in this study suggest that this pathovar can be segregated into two subgroups under a recently proposed reclassification of the Xanthomonas genus.     

Symptomless Spread of Blight-inducing Strains of Xanthomonas campestris pv. campestris on Cabbage Seedlings in Misted Seedbeds

Xanthomonas campestris pv. campestris induces two types of symptoms, namely, black rot and blight. Black rot symptoms are V-shaped lesions and black veins on the leaf, and blight symptoms are sudden collapse of interveinal tissues following the lack of veinal necrosis at early stages of infection. These two symptoms can occur simultaneously. However, the tendency to induce either symptom type is strain-dependent. Six strains were evaluated for their rate and pattern of spread in misted seedbeds by using strain-specific monoclonal antibodies and miniplate enrichment/ELISA. Data on pathogen incidence was defined as the presence of the pathogen in or on plants rather than visual symptoms. The results indicated that blight-inducing strains spread to more seedlings than black rot-inducing strains. The high incidences of blight-inducing strains in experimental plots were associated with non-randomness of spatial pattern of pathogen spread, indicating that high incidence is primarily due to the spread from adjacent plants by leaf contact and water splash. Most ELISA-positive seedlings were symptomless, indicating that the sensitivity of the system used in this study was adequate for detection of latent or epiphytic spread.     

Survival and Extinction of Xanthomonas campestris pv. campestris in Soil

Carry-over of inoculum of X.c. pv. campestris in the soil from one cropping season to the next was studied in field experiments over three years. These studies were supported by laboratory and greenhouse experiments on quantitative assessment of bacteria by bioassay using the Most Probable Number technique, and on recovery rates of bacteria from the soil. The mean recovery rate from artificially infested soil was 58%. Extinction of X.c. pv. campestris in soil infested with infected plant debris proceeded exponentially and extinction rates depended on temperature, as did the decomposition of plant debris. In replicated field plots, over three years, infection foci of black rot disease were established. At harvest time, all plants were chopped and resulting plant debris was rotovated into the soil. The resulting soil infestation was sampled and showed clear infestation foci reflecting the original infection foci of the crop. These infestation foci decreased with time and disappeared after the winter. Follow-up crops remained virtually uninfected. The results show that in The Netherlands good crop and soil management impedes survival of inoculum from one year to the next, so that cabbage can be grown continuously. Polyetic carry-over of inoculum by debris in the soil can be avoided in The Netherlands.     

Molecular characterization of the incitant of cowpea bacterial blight and pustule, Xanthomonas campestris pv. vignicola

Strains of Xanthomonas campestris pv. vignicola (Xcv), isolated from cowpea leaves with blight or minute pustules and collected from various geographic areas, were selected on the basis of pathological and physiological features. All strains were analyzed for genotypic markers by two methods: ribotyping with EcoRI endonuclease, and RFLP analysis with a plasmid probe (pthB) containing a gene required for pathogenicity from Xanthomonas campestris pv. manihotis. Ribotyping revealed a unique pattern for all the strains that corresponded to the previously described ribotype rRNA7. Based on polymorphism detected by pthB among Xcv strains, nine haplotypes were defined. The observed genetic variation was independent of the geographic origin of the strains and of pathogenic variation. Some haplotypes were widely distributed, whereas others were localized. In some cases, we could differentiate strains isolated from blight symptoms and pustules according to haplotypic composition. However, in most cases, no significant differences were observed. Our results and the previous pathogenic and biochemical characterizations suggest that the strains isolated from leaves with blight symptoms or minute pustules belong to the same pathovar. We provide information on pathogen diversity that can be used to identify and characterize resistant germplasm.     

Pigment-associated proteins of Xanthomonas campestris pv. juglandis

Two pigment-protein complexes extracted from the cell membrane of Xanthomonas campestris pv. juglandis with 2% Triton X-100 were separated from other membrane proteins by electrophoresis on a 10%., non-denaturing discontinuous polyacrylamide gel. One pigment-protein complex band was distinct, while the other was diffuse. The apparent M_r of the protein from the distinct pigment-protein band was 16400, while the protein in the diffuse band had an apparent M_r of about 45000. The protein in the distinct band consisted of 13 amino acids of which 10% were aromatic, 12% hydroxy, 16% basic, 16% acidic and 46% non-polar. Polyclonal antibody, against the distinct protein, was used to assay for cross-reactivity with cell wall and membrane proteins of 23 bacterial species by the Ouchterlony double-diffusion assay. Seven of the bacteria, representing seven genera, cross-reacted with the antibody, suggesting that a serologically-related, pigment-associated protein is commonly distributed among bacteria and which, unlike the pigment, may limit its use as a chemotaxonomic marker for Xanthomonas .     

Bacterial Blight of Welsh Onion : A New Disease Caused by Xanthomonas campestris pv. allii pv. nov.

In June of 1998, a new bacterial disease was observed on Welsh onion in Okinawa Prefecture, Japan. Infected plants in nursery boxes were stunted with tip dieback, and heavily infected plants died. In fields, the disease appeared on leaves as irregular gray spots or elliptical spots with creases in the center. These spots enlarged and spread rapidly continued cloudy or rainy weather, and formed blight lesions on outer leaves. Yellow mucoid bacterial colonies were consistently isolated from these lesions. The causal bacterium was identified as a pathovar of Xanthomonas campestris on the basis of bacteriological properties. The bacterium was pathogenic to Welsh onion, onion, but nonpathogenic to chive, Chinese chive and hyacinth. Of Liliaceae plants, which contain Welsh onion and onion, only hyacinth has been reported as a host for the genus Xanthomonas, namely X. campestris pv. hyacinthi. However, strains of X. campestris pv. hyacinthi were not pathogenic against either Welsh onion or onion. From these results, the bacterium isolated from Welsh onion is considered to be a new pathovar of X. campestris, and the name of X. campestris pv. allii pv. nov. is proposed. A strain MAFF 311173 is designated as the pathotype strain. Received 29 March 2000/ Accepted in revised form 4 July 2000     

Identification and Origin of Xanthomonas campestris pv. campestris Races and Related Pathovars

ABSTRACT One hundred sixty-four isolates of Xanthomonas campestris pv. campestris and other X. campestris pathovars known to infect cruciferous hosts (X. campestris pvs. aberrans, raphani, armoraciae, and incanae) were inoculated onto a differential series of Brassica spp. to determine both pathogenicity to brassicas and race. Of these, 144 isolates were identified as X. campestris pv. campestris and grouped into six races, with races 1 (62%) and 4 (32%) being predominant. Other races were rare. The remaining 20 isolates from brassicas and other cruciferous hosts were either nonpathogenic or very weakly pathogenic on the differential series and could not be race-typed. Five of these isolates, from the ornamental crucifers wallflower (Cheiranthus cheiri), stock (Matthiola incana) and candytuft (Iberis sp.), showed clear evidence of pathovar-like specificity to the hosts of origin. A gene-for-gene model based on the interaction of four avirulence genes in X. campestris pv. campestris races and four matching resistance genes in the differential hosts is proposed. Knowledge of the race structure and worldwide distribution of races is fundamental to the search for sources of resistance and for the establishment of successful resistance breeding programs.