Differentiation of three species of Xanthomonas and Stenotrophomonas maltophilia using cellular fatty acid analyses

Five hundred eighty-eight strains, representing Xanthomonas albilineans, X. fragariae, ten pathovars of X. campestris, and Stenotrophomonas maltophilia from ornamentals, were subjected to fatty acid methyl ester (FAME) analyses. Quantitative variance among FAME profiles enabled identification of the four species with 100% accuracy. Dendrogram cluster analysis placed strains of X. albilineans remotely from those of the other two Xanthomonas species and S. maltophilia. Whereas some profiles of pathovars of X. campestris were distinct, strains within X. albilineans, X. fragariae, and S. maltophilia were homogeneous by their conserved FAME ratios. Pathovars of X. campestris that had conserved profiles were fittonia, hederae, malvacearum, pelargonii, and zinniae. FAME profiles of X. campestris pathovars begoniae, dieffenbachiae, fici, maculifoliigardeniae, and poinsettiicola were, however, quantitatively diverse. These pathovars did not form discrete subgroups, and intercalated randomly with one another on the dendrogram. Certain species or pathovars of X. campestris which have homogeneous FAME profiles can easily be identified with fatty acid analysis; however, pathovars of X. campestris with heterogeneous profiles are not readily identified by fatty acid analysis.     

Specific Oligonucleotide Primers for the Rapid Identification and Detection of the Agent of Tomato Pith Necrosis, Pseudomonas corrugata, by PCR Amplification: Evidence for two Distinct Genomic Groups

Unique DNA bands from strains representative of two groups of Pseudomonas corrugata, as shown by amplification of their genomic DNA by polymerase chain reaction using short random sequence oligonucleotide primers (RAPD-PCR), were isolated, cloned and sequenced. Two pairs of specific primer sequences, based on the ends of the cloned unique DNA bands from strains IPVCT10.3 and IPVCT8.1, were used in multiplex PCR with a range of P. corrugata strains. All strains produced one of the two specific bands, 1100bp (from the IPVCT10.3-based primers) and 600bp (from the IPVCT8.1-based primers), representing groups designated I and II, respectively. The primers were also tested on a wider range of Pseudomonas species, including the closely-related fluorescent Pseudomonas genomospecies FP1, FP2 and FP3: none of these bacteria produced any bands following amplification by PCR with these primers. The primer sets detected P. corrugata in tomato pith necrosis-infected plants providing a useful tool for rapid identification and epidemiological studies.     

Variability in aggressiveness,cultural characteristics,cercosporin production and fatty acid profile of Cercospora piaropi,a biocontrol agent of water hyacinth

The extent of variation in aggressiveness, growth and pigmentation in culture, phytotoxin production and fatty acid profile were determined in a population of 55 isolates of Cercospora piaropi, a fungus used as a biocontrol agent of the aquatic weed water hyacinth (Eichhornia crassipes). Besides differences in the colour of mycelium and diffusible pigments in culture, isolates of C. piaropi grown under standard conditions differed significantly in their ability to produce the phytotoxin cercosporin, as well as in aggressiveness and growth rate. A positive correlation existed between the ability of the isolates to produce cercosporin and their aggressiveness, and a negative correlation between growth rate and cercosporin production or growth rate and aggressiveness. Based on thin‐layer chromatographic separation of extracts and comparison with beticolin‐1, used as a standard, there was no evidence of production of beticolins. In discriminant analysis, fatty acid methyl ester (FAME) profiles had low resolution for differentiating populations among isolates of the fungus, and the level of resolution was influenced by the age of the colonies. Diffusible pigments in culture and cercosporin production are useful adjuncts to aggressiveness screening for choosing the most effective isolate of C. piaropi for biological control.     

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.     

浙江西兰花花球头腐病的病原鉴定

为明确近年来浙江省宁波市部分地区发生的西兰花花球头腐病的病原,通过田间症状观察、致病性测定、Biolog和脂肪酸(fatty acid methyl ester,FAME)分析将分离到的病原菌鉴定为荧光假单胞菌Pseudomonas fluorescens.利用16S rDNA和核糖体DNA内转录间隔区(internal transcribed spacer,ITS)序列构建的系统发育树也都显示分离的西兰花菌株与假单胞菌聚合成群,其中与荧光假单胞菌的亲缘关系最近,序列同源性分别为99%和98%.表明浙江省西兰花花球上发生的细菌性头腐病由荧光假单胞菌引起.     

Use of Fatty Acid Methyl Ester Profiles to Compare Copper-Tolerant and Copper-Sensitive Strains of Pantoea ananatis

ABSTRACT A survey was conducted to evaluate differences in fatty acid methyl ester (FAME) profiles among strains of Pantoea ananatis, causal agent of center rot of onion (Allium cepa), isolated from 15 different onion cultivars in three different sites in Georgia. Differences in FAME composition were determined by plotting principal components (PCs) in two-dimensional plots. Euclidean distance squared (ED(2)) values indicated a high degree of similarity among strains. Plotting of PCs calculated from P. ananatis strains capable of growing on media amended with copper sulfate pentahydrate (200 mug/ml) indicated that copper-tolerant strains grouped into tight clusters separate from clusters formed by wild-type strains. However, unlike copper-sensitive strains, the copper-tolerant strains tended to cluster by location. A total of 80, 60, and 73% of the strains from Tift1, Tift2, and Tattnall, respectively, exhibited either confluent growth or partial growth on copper-amended medium. However, all strains were sensitive to a mixture of copper sulfate pentahydrate (200 mug/ml) and maneb (40 mug/ml). When copper-tolerant clones were analyzed and compared with their wild-type parents, in all cases the plotting of PCs developed from copper-tolerant clones formed tight clusters separate from clusters formed by the parents. Eigenvalues generated from these tests indicated that two components provided a good summary of the data, accounting for 98, 98, and 96% of the standardized variance for strains Pna 1-15B, Pna 1-12B, and Pna 2-5A, respectively. Furthermore, feature 4 (cis-9-hexadecenoic acid/2-hydroxy-13-methyltetradecanoic acid) and feature 7 (cis-9/trans-12/cis-7-octadecenoic acid) were the highest or second highest absolute values for PC1 in all three strains of the parents versus copper-tolerant clones, and hexadecanoic acid was the highest absolute value for PC2 in all three strains. Along with those fatty acids, dodecanoic acid and feature 3 (3-hydroxytetradecanoic acid/14-methylpentadecenoic acid) also had an impact on the differences observed between copper-sensitive parents and copper-resistant mutants. Finding these changes in bacterial fatty acid composition could lead to the development of a laboratory assay to identify copper-tolerant strains using gas chromatography as well as providing clues to further elucidate the mode of action of copper tolerance.     

First report of Rhizomonas sp. causing corky root of lettuce in Europe

Typical symptoms of corky root were observed on iceberg lettuce (Lactuca sativa L.) in the Netherlands and England, on prickly lettuce (Lactuca serriola L.) in Spain, and on sowthistle (Sonchus oleraceus L.) in Greece. Slow-growing bacteria with similar colonies as strains ofRhizomonas suberifaciens or otherRhizomonas species were isolated from soil surrounding plants with corky root symptoms using lettuce seedlings as bait. Crude lysate from all strains was tested for DNA homology with DNA fromR. suberifaciens strain CA1 (R. sub. CA1) andRhizomonas sp. strain WI4 (R. sp. WI4). Strains that had homology values higher than that ofR. sp. WI4 orR. sub. CA1 were tested for pathogenicity on 1-wk-old lettuce seedlings, cv. Salinas, Two strains from the Netherlands induced typical symptoms of corky root on lettuce. These strains tested negative with monoclonal antibody MAb-Rs1 specific forRhizomonas suberifaciens in an enzyme-linked immunosorbent assay (ELISA). They had low DNA homology withR. sub. CA1 (4–9%) and low to moderate DNA homology withR. sp. WI4 (4–17%). Some nonpathogenic strains had moderate to high levels of DNA homology withR. sub. CA1 orR. sp. WI4 (19–84%). All strains had fatty acid profiles similar to those ofRhizomonas species. This is the first report ofRhizomonas sp. causing corky root of lettuce in Europe.     

Evaluation of plant growth-promoting rhizobacteria for biological control of pythium root rot of cucumbers grown in rockwool and effects on yield

Three strains ofPseudomonas fluorescens (63-49, 63-28, and 15), one strain ofPseudomonas corrugata (13) and one strain ofSerratia plymuthica (R1GC4) were tested on rockwool-grown cucumbers for their ability to reduce Pythium root-rot caused byPythium aphanidermatum. These strains were previously selected for biocontrol ability from collections of >4000 bacteria. Strains 63-49 and 63-28 were tested on cucumber plants grown in rockwool in two replicatedPythium-inoculated trials conducted in British Columbia (B.C). Another inoculated, replicated trial was conducted in Quebec with all five strains. Cucumber yields (fruit number and weight) were measured over a ten-week harvest period. Strain 63-49 caused an early promotion of plant growth and increased cucumber yields at early harvests. No measurable effect ofPythium inoculation on disease development was observed in the Quebec trial, due to unfavourable cool weather. However, 63-49 significantly increased the total number of cucumbers (12%) and cucumber weight (18%), compared to the non-treated control. Strains 13, 15 and R1GC4 slightly increased the cumulative cucumber yields, but strain 63-28 had no effect. In the B.C. trial, inoculation withP. aphanidermatum reduced the number and weight of cucumbers by 27%. Treatments ofPythium-inoculated cucumbers with 63-49 significantly increased fruit number and weight by 18%, compared to thePythium-inoculated control. Strain 63-28 increased the cumulative number of cucumbers over time, compared to thePythium-inoculated control, but the increase was less than with 63-49. The use ofPseudomonas spp. in rockwool-grown cucumbers can increase yields, both in the presence and absence of Pythium root rot, and with variable seasonal conditions and disease pressures.     

Identification of the causal agent of pistachio dieback in Australia

Symptoms associated with pistachio dieback in Australia include decline (little or no current season growth), xylem staining in shoots two or more years old, trunk and limb lesions (often covered by black, superficial fungal growth), excessive exudation of resin, dieback and death of the tree. Bacteria belonging to the genus Xanthomonas have been suggested as the causal agent. To confirm the constant association between these bacteria and the disease syndrome, the absence of other pathogens and the identity of the pathogen, we performed a series of isolations and pathogenicity tests. The only microorganism consistently isolated from diseased tissue was a bacterium that produced yellow, mucoid colonies and displayed morphological and cultural characteristics typical of the genus Xanthomonas. Database comparisons of the fatty acid and whole-cell protein profiles of five representative pistachio isolates indicated that they all belonged to X. translucens, but it was not possible to allocate the isolates to pathovar. Pathogenicity tests on cereals and grasses supported this identification. However, Kochs postulates have been only partially fulfilled because not all symptoms associated with pistachio dieback were reproduced on inoculated two-year-old pistachio trees. While discolouration was observed, dieback, excessive resinous exudate and trunk and limb lesions were not produced; expression of these symptoms may be delayed, and long-term monitoring of a small number of inoculated trees is in progress.     

A strain ofPseudomonas syringae which does not belong to pathovarphaseolicola produces phaseolotoxin

A bacterial strain, CFBP 3388, isolated from Vetch (Vicia sativa, L.) was identified asP. s. pv.syringae on the basis of nutritional and biochemical patterns which were obtained with classical tests and the Biolog system. It caused necrotic symptoms typical ofP. s. pv.syringae on bean leaves and pods after artificial inoculation. However, the isolate caused a citrulline-reversible inhibition ofE. coli in phaseolotoxin bioassay. Furthermore, with CFBP 3388 DNA as template a 1900 bp DNA fragment, specific for the phaseolotoxin DNA cluster ofP. s. pv.phaseolicola, was amplified by PCR. This is the first demonstration that an isolate ofP. syringae that is not pv.phaseolicola can produce phaseolotoxinAbbreviations bp base pair - kb kilobase - OCT Ornithine Carbamoyl Transferase     

吡虫啉胁迫对褐飞虱共生解脂假丝酵母抗性及敏感菌株生长的影响

为明确吡虫啉对褐飞虱共生解脂假丝酵母Candida lipolytica抗性及敏感菌株生长的影响,通过菌株培养、菌落观察和菌丝镜检等方法,比较了褐飞虱共生解脂假丝酵母不同菌株在含不同质量浓度吡虫啉的固体培养基上的菌落数,以及在液体培养基中的生长量差异。结果发现:不同质量浓度吡虫啉对褐飞虱共生解脂假丝酵母敏感和抗性菌株菌落生长均有抑制作用,且吡虫啉浓度越高,抑制作用越强。经500、1 000和2 000 mg/L吡虫啉处理4 d后,敏感菌株的菌落数量分别为对照的46.61%、27.58%和6.25%,均与对照差异显著;500和1 000 mg/L处理组抗性菌株菌落数量与对照无显著差异,而2 000 mg/L处理组与对照差异显著。经吡虫啉处理后,敏感菌株假菌丝形态变得不规则,部分假菌丝不舒展、萎缩或弯曲、顶端出现膨大,酵母出现空泡等,且吡虫啉浓度越高,不规则程度越明显;抗性菌株的假菌丝形态也有类似变化,但与敏感菌株相比,其菌丝体不规则形态的比例明显下降。500 mg/L吡虫啉处理对敏感和抗性菌株的生长量及菌丝干重均无明显影响,1 000和2 000 mg/L吡虫啉对不同菌株前期生长的抑制作用明显,但对后期生长影响不明显。研究表明,吡虫啉对褐飞虱共生解脂假丝酵母抗性菌株生长的影响显著小于对敏感菌株的影响。     

Effect ofCuscuta infection on chloroplast lipid composition ofBrassica leaves

Brassica campestris was infected with the angiosperm parasite,Cuscuta reflexa, on 24 DAS (days after sowing). A significant reduction (43%) in carbon dioxide assimilation in leaves was found compared to normal plant, when both were assayed on 38 DAS. At this stage, the chloroplast lipid in infected leaves had decreased by 34%, with a reduction of 13, 38 and 55%, respectively, in neutral, glyco- and phospholipids. Among the glycolipids, the decrease in the content of MGDG, DGDG and SQDG was 21, 67 and 44%, respectively, with 2.4-fold increase in the MGDG/DGDG ratio. The chlorophyll content was reduced by 24%, while the carotenoid level increased by 44%. The free fatty acid content was enhanced by 47% which may be attributed to an increase in lipase activity. The contents of saturated fatty acids (capric, lauric, myristic, palmitic and stearic) increased while the contents of unsaturated fatty acids (palmitoleic, oleic, linoleic and linolenic) decreased. The ratio of unsaturated to saturated fatty acids decreased over 50%. These findings may suggest chloroplasts as reactive sites of the host-parasite interaction in case of the angiosperm parasite.Abbreviations DAS day after sowing - DGDG digalactosyl diglyceride - FAME fatty acid methyl esters - FFA free fatty acids - MGDG monogalactosyl diglyceride - SQDG sulfoquinovosyl diglyceride     

Phytophthora root and crown rot of raspberry in Bulgaria

Root and crown rot of raspberry (Rubus idaeus L.) was observed in a plantation at the experimental station of small fruits in Kostinbrod, Bulgaria. Isolates ofPhytophthora spp. were obtained from diseased plants. Colony morphology, growth rates, features of asexual and sexual structures were studied and as a result twoPhytophthora species were identified:Phytophthora citricola Saw. andPhytophora citrophthora (R.E. Sm. & E.H. Sm.) Leonian. Their pathogenicity was confirmed in artificial inoculation experiments. The isozyme (-esterase) patterns ofP. citrophthora andP. citricola isolates from raspberry and from the collection of the CBS, Baarn the Netherlands were compared, using micro-gel electrophoresis. Both species are reported for the first time as pathogens of raspberry in Bulgaria. This is only the second report in phytopathological literature ofP. citrophthora on raspberry, the first being from Chile [Latorre and Munoz, 1993].     

Cloning of a specific DNA region of white top pathogen of pea and its detection by polymerase chain reaction

White top strain (WT strain) of Pseudomonas syringae pv. pisi (Ppi) is a variant strain causing white top disease of peas. The WT strain is distinguishable from common Ppi strains only by symptom expression chlorosis and whitening of apical shoots. To develop a specific detection method for the WT strain, we cloned a specific DNA region of the WT strain using transposon tagging. Five mutants defective in white top symptom expression were obtained. A part of the Tn5-flanking region was cloned and labeled as a hybridization probe. One clone, pAY3, gave two signal bands, one of which was detected from the genomic DNA of all the WT and the common Ppi strains; another was specific to WT strains. A restriction map of pAY3 showed that it contains two BamHI fragments; one is 5.0kb in length involving a part of Tn5, and the other is 1.5kb, did not carry Tn5, and may have been accidentally ligated into pAY3. The 1.5-kb band was subcloned as pAY13 and was used as a probe. It hybridized specifically to WT strains. These results suggested that the WT strains have a specific DNA region and that part of the region was successfully cloned. Sequence analysis of pAY13 showed that it is similar to part of nonribosomal peptide synthetases (NRPSs) genes. The deduced amino acid sequence of pAY13 suggested the existence of eight conserved motifs of NRPSs. WT strain-specific PCR primers, PS1 and PS2, were designed from the DNA sequence. These primers gave a specific amplification product of 981bp from both the genomic DNA and a direct cell preparation of WT strains. No specific amplicon was produced from Ppi strains that caused only water-soaked lesions or from strains of other P. syringae pathovars. A specific amplicon was not produced from four strains of the pea pathogen: P. marginalis pv. marginalis, P. viridiflava, Erwinia carotovora ssp. carotovora, Xanthomonas campestris pv. pisi. Using the primers, WT strain was detected from water-soaked lesions and green and white tissues without water soaking.The sequence reported in this paper has been deposited in the DDBJ database under accession no. AB117755     

Phytophthora root rot of sweet pepper

Phytophthora capsici proved to be the causal agent of a root and crown rot of sweet pepper in the Netherlands.P. capsici was pathogenic on sweet pepper, tomato and sometimes on eggplant but not on tobacco Xanthi. Of these test plants only tomato was infected byP. nicotianae.No different symptoms in plants infected with eitherP. capsici orP. nicotianae were found. Dipping the roots of tomato and sweet pepper plants in a suspension ofP. capsici resulted in a more severe attack than pouring the suspension on the stem base.Resistance in tomato toP. nicotianae did not include resistance toP. capsici. A method to distinguishP. capsici fromP. nicotianae after isolation from soil is described. Both species were able to infect green fruits of tomato and sweet pepper.p. capsici survived in moist soil in the absence of a host for at least 15 months.Samenvatting Phytophthora capsici bleek de oorzaak te zijn van een voet-en wortelrot in paprika op twee bedrijven in 1977 in Nederland.P. capsici was pathogeen op paprika, tomaat en soms op aubergine maar niet op tabak Xanthi.P. nicotianae tastte van deze toetsplanten alleen tomaat aan. Verschillen in symptomen tussenP. nicotianae enP. capsici werden bij tomaat niet waargenomen.Het dompelen van de wortels in eenP. capsici suspensie gaf een ernstiger aantasting dan het begieten van de wortelhals met deze suspensie.Resistentie in tomaat tegenP. nicotianae bleek geen resistentie tegenP. capsici in te houden. P. capsici kan in grond worden aangetoond door groene paprikavruchten als vangsubstraat te gebruiken.P. capsici enP. nicotianae kunnen beide zowel vruchten van tomaat als paprika aantasten. P. capsici overleefde een periode van 15 maan den in vochtige grond waarop geen waardplant werd geteeld.     

Characterization and differentiation of <Emphasis Type="Italic">Erwinia carotovora</Emphasis> strains from mulberry trees

We recently reported that two diverse types (types 1 and 2) were identified among strains of Erwinia carotovora from mulberry trees. Type 1 strains were similar to E. carotovora subsp. carotovora (Ecc), whereas type 2 strains were distinct from Ecc and other E. carotovora strains. In this study, seven more mulberry strains of type 2 and reference strains were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and randomly amplified of polymorphic DNA (RAPD). On the basis of SDS-PAGE profiles of whole-cell proteins, type 2 strains had high similarity with one another. In addition, they had an unique peptide band with a molecular mass of approximately 28kDa. RAPD analysis showed that they were also effectively differentiated by a strong, specific RAPD fragment for type 2 strains. Based on these two approaches, we have confirmed that the present type 2 strains from mulberry can be discriminated clearly from other soft rot Erwinia species.     

Transmission efficiency of five tobravirus strains byParatrichodorus teres

Nematodes from a population of virus-freeParatrichodorus teres were allowed to feed on roots ofPetunia hybrida andNicotiana tabacum cv. White Burley plants infected with five strains of tobacco rattle or pea early-browning tobraviruses and were subsequently tested for virus transmission to roots of virus-freeP. hybrida andN. tabacum cv. White Burley plants. Virus transmission was not correlated with serotypes. Two strains of TRV with different serotypes were efficiently transmitted, whereas two other strains of TRV and one strain of PEBV with serotypes similar to transmitted ones were not transmitted at all. The two efficiently transmitted strains had originally been obtained fromP. hybrida roots which had been exposed to viruliferousP. teres in laboratory bait tests, whereas the three non-transmitted strains had been obtained from infected plant material collected in the field.     

Differentiation of Three Homogeneous Groups of Rhizoctonia solani Anastomosis Group 4 by Analysis of Fatty Acids

ABSTRACT Profiles of fatty acids from 70 isolates of Rhizoctonia solani anastomosis group (AG)-4 clustered into three groups, corresponding to homogeneous group (HG)-I, HG-II, and a newly described HG-III. Isolates from Georgia peanuts exhibiting limb rot were characterized as gas chromatography (GC) subgroup 1 (GC-1) and contained HG-I isolates. Isolates from diseased soybean hypocotyls grown in North Dakota and sugar beet seedlings, taproots, and tare soil in Minnesota and North Dakota were characterized as GC subgroup 2 (GC-2) and contained predominantly HG-II isolates but also included three distinct isolates based on fatty acid methyl ester (FAME) analysis and morphological features. Selected isolates from North Carolina cucumbers clustered into three distinct groups that corresponded to HG-I, HG-II, and the newly described HG-III. Distinct isolates from the soybean and sugar beet populations clustered with HG-III. Fatty acid profiles of AG-4 were compared with FAME library profiles of AG-1, AG-2 type 2, and AG-3, which were developed in previous studies and were sufficiently different that they could be used to support speciation of this group from R. solani. It is suggested that binomial R. practicola may be appropriate for the portion of AG-4 identified as HG-II.     

Phenotypic and Genotypic Characterization of Xanthomonas campestris pv. zinniae Strains

During 1997 and 1998, serious outbreaks of bacterial leaf spot disease were observed on zinnia plants grown in home and commercial gardens in Ohio, USA. Twenty-two strains of Xanthomonas campestris pv. zinniae, isolated from diseased zinnia plants and contaminated seeds, were identified based on morphological, physiological and biochemical tests, fatty acid methyl ester analyses and pathogenicity tests on zinnia cv. Scarlet. Host range studies indicated that all of the X. campestris pv. zinniae strains were pathogenic on zinnia and tomato, but not on cabbage, lettuce, pepper and radish. The phenotypic and genotypic relationships among the strains determined based on serological reaction pattern, fatty acid profiles, repetitive extragenic palindromic-polymerase chain reaction (rep-PCR) fingerprints and sequence analysis of the 16S–23S rDNA spacer region suggested that X. campestris pv. zinniae strains were closely related to each other, but clearly distinct from other Xanthomonas species including X. campestris pv. campestris, X. axonopodis pv. vesicatoria, X. vesicatoria and X. hortorum pv. vitians tested in this study. The results also demonstrated that rep-PCR fingerprinting is rapid, reliable and the most practical method for routine detection and identification of X. campestris pv. zinniae strains.     

Induction of hypericins and hyperforin in Hypericum perforatum L. in response to biotic and chemical elicitors

Hypericum perforatum L. produces hyperforins, a family of antimicrobial acylphloroglucinols; and hypericins, a family of phototoxic anthraquinones exhibiting anti-microbial, anti-viral, and anti-herbivore properties in vitro. To determine whether these secondary metabolites are part of the specific plant defense systems that are mediated by methyl jasmonate or salicylic acid, we used meristem cultures to assess the effects of exposure to exogenous application of these chemical elicitors. Levels of hypericins in plant tissue increased in response to both elicitor treatments; total hypericin levels increased as much as 3.3 times control levels when treated with 200 μ methyl jasmonate for 14 days. Increased hyperforin concentrations were detected when plantlets were treated with 1 m salicylic acid or 50 μ methyl jasmonate. For assessing responses to a biotic elicitor, greenhouse-grown plant materials were inoculated with the plant pathogen, Colletotrichum gloeosporioides. Levels of hypericins increased twice as much as the control when inoculated with 1 × 10⁴ spores per ml; higher doses of spores overwhelmed the plant defenses. The elevation of hypericins and hyperforin in response to chemical and biotic elicitors suggests that these secondary metabolites are components in the inducible plant defense responses of H. perforatum.