Dynamics in Secondary Metabolite Gene Clusters in Otherwise Highly Syntenic and Stable Genomes in the Fungal Genus Botrytis.

Fungi of the genus Botrytis infect >1,400 plant species and cause losses in many crops. Besides the broad host range pathogen Botrytis cinerea, most other species are restricted to a single host. Long-read technology was used to sequence genomes of eight Botrytis species, mostly pathogenic on Allium species, and the related onion white rot fungus, Sclerotium cepivorum. Most assemblies contained <100 contigs, with the Botrytis aclada genome assembled in 16 gapless chromosomes. The core genome and pan-genome of 16 Botrytis species were defined and the secretome, effector, and secondary metabolite repertoires analyzed. Among those genes, none is shared among all Allium pathogens and absent from non-Allium pathogens. The genome of each of the Allium pathogens contains 8-39 predicted effector genes that are unique for that single species, none stood out as potential determinant for host specificity. Chromosome configurations of common ancestors of the genus Botrytis and family Sclerotiniaceae were reconstructed. The genomes of B. cinerea and B. aclada were highly syntenic with only 19 rearrangements between them. Genomes of Allium pathogens were compared with ten other Botrytis species (nonpathogenic on Allium) and with 25 Leotiomycetes for their repertoire of secondary metabolite gene clusters. The pattern was complex, with several clusters displaying patchy distribution. Two clusters involved in the synthesis of phytotoxic metabolites are at distinct genomic locations in different Botrytis species. We provide evidence that the clusters for botcinic acid production in B. cinerea and Botrytis sinoallii were acquired by horizontal transfer from taxa within the same genus.

High nitrogen concentration alter microbial community in Allium fistulosum rhizosphere.

Welsh onion (Allium fistulosum L.) constitutes an important plant species cultivated in China due the benefits and applications in different areas. Moreover, nitrogen is an essential nutrient during the growth and development of plant. Here, we present the effects of nitrogen on soil microbiome in welsh onion plants. We used High-throughput sequencing analysis to determine the diversity and abundances of microbes associated to soil rhizosphere in welsh onion under the influence of nitrogen application. Nitrogen application significantly influenced in the diversity of fungal community. The relative abundance of Orbiliomycetes increased with the nitrogen concentration. Nitrogen application did not affect the diversity of bacterial community, whereas the relative abundance of Acidobacteria_Gp2, Verrucomicrobiae and Sphingobacteriia decreased with the nitrogen condition. In this work, we introduced evidences of the effect of nitrogen fertilization on microbial community in welsh onion rhizosphere, and the change of microbial community may interfere the growth and development of welsh onion.

Illumina-based Analysis of Endophytic Bacterial Diversity of four Allium species.

Allium species are popular vegetables in China and possess antifungal and antibacterial activities. This study aimed to compare the endophytic bacterial community in the four crucial Allium species in China, Chinese leek (CL), garlic (GA), onion (ON,) and Welsh onion (WO), using sequences of the V3-V4 region of the bacterial 16S rRNA gene. A total of 1,036,637 high-quality sequences and 719 operational taxonomic units (OTUs) were obtained across all libraries. A total of 20 phyla, 50 classes, 80 orders, 134 families, and 234 genera were identified. Among them, 18 OTUs and 19 genera were shared among the four Allium species. Proteobacteria (42.68%) and Bacteroidetes (20.18%) were the dominant phyla in CL, while one unclassified (>70%) was the dominant phyla in the other three Allium species. The alpha-diversity analysis showed the bacterial richness and diversity in CL were significantly higher than those in the other three Allium species. Principal coordinate analysis (PCA) showed endophytic bacterial communities in GA, WO, and ON were more similar than those in CL. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) analysis revealed endophytic bacteria mostly enriched in Membrane Transport, Amino Acid Metabolism and Carbohydrate Metabolism pathway. 17 of the 23 Kyoto Encyclopedia of Genes and Genomes (KEGG) categories and 159 of the 206 lower-level KEGG pathways in CL were significantly higher than those in the other three Allium species. Pearson's correlation indicated that KEGG pathways with significant differences among the Allium species were closely related to the bacterial genera with significant differences between the Allium species. The findings of our study provided insight into the complex endophytic microbial communities in Allium species.

Microbial basis of Fusarium wilt suppression by Allium cultivation.

Crop rotation and intercropping with Allium plants suppresses Fusarium wilt in various crops. However, the mechanisms underlying this phenomenon have not been fully elucidated. This study was designed to assess the role of microorganisms inhabiting Allium rhizospheres and antifungal compounds produced by Allium roots in Fusarium wilt suppression by Allium cultivation. Suppression of cucumber Fusarium wilt and the pathogen multiplication by Allium (Welsh onion and/or onion)-cultivated soils were eliminated by heat treatment at 60 °C, whereas those by Welsh onion-root extract were lost at 40 °C. The addition of antibacterial antibiotics eliminated the suppressive effect of Welsh onion-cultivated soil on pathogen multiplication, suggesting the contribution of antagonistic gram-negative bacteria to the soil suppressiveness. The Illumina MiSeq sequencing of 16S rRNA gene amplicons revealed that genus Flavobacterium was the predominant group that preferentially accumulated in Allium rhizospheres. Flavobacterium species recovered from the rhizosphere soils of these Allium plants suppressed Fusarium wilt on cucumber seedlings. Furthermore, confocal laser scanning microscopy revealed that Flavobacterium isolates inhibited the multiplication of the pathogen in soil. Taken together, we infer that the accumulation of antagonistic Flavobacterium species plays a key role in Fusarium wilt suppression by Allium cultivation.

An empirical investigation of the possibility of adaptability of arbuscular mycorrhizal fungi to new hosts.

Little is known about the adaptive capacity of arbuscular mycorrhizal (AM) fungi to novel hosts. Here we assessed the possibility of two heterospecific AM fungal isolates to adaptively change, in terms of host biomass response, as a function of host plant identity, over the course of a growing season. First, we produced pure inocula of Rhizophagus clarus and Rhizophagus intraradices, each starting from a single spore. Second, we "trained" each isolate individually in a community with two plants, sudangrass (Sorgum bicolour subsp. drummondii) and leek (Aliium ampeloprasum var. porrum), using a dual-compartment system to allow the establishment of a common mycorrhizal network between the two hosts. Third, we conducted a greenhouse experiment to reciprocally test each "trained" clone, obtained from each compartment, either with the same (home), or the other host (away) under two contrasting phosphorus levels. Overall, results did not support adaptive responses of the AM fungi to their hosts (i.e., greater host biomass under "home" relative to "away" conditions), but the opposite (i.e., greater host biomass under "away" relative to "home" conditions) was more frequently observed. These changes in AM fungal symbiotic functioning open the possibility for relatively rapid genetic change of arbuscular mycorrhizal fungi in response to new hosts, which represents one step forward from in vitro experiments.

Talaromycolides A-C, Novel Phenyl-Substituted Phthalides Isolated from the Green Chinese Onion-Derived Fungus Talaromyces pinophilus AF-02.

The green Chinese onion (Allium fistulosum L.), which is widely cultivated and has been naturalized in many places, is an important spice and vegetable in East and Southeast Asia. It is used to treat the common cold in China. In the ongoing search for antibacterial activity in fungi derived from natural, pungently scented vegetables, the secondary metabolites of Talaromyces pinophilus AF-02, which was isolated from the stem of the green Chinese onion, were investigated. The genus Talaromyces (Trichocomaceae) is an important fungal genus because of its ubiquity and the role of many of its species in food and agriculture production. Three new phthalide derivatives, talaromycolides A-C, 1-3; a new long-chain dicarboxylic acid, 11; and 12 known compounds were isolated from methanolic extracts of this fungus. Their structures were determined via extensive NMR, HR-ESI-MS, and CD spectroscopic analyses. Compounds 1-3 are rare phthalide derivatives with a novel linkage position between the phenyl and phthalide moieties. The biological properties of 1-16 were evaluated using six different bacteria, and 1-3, 5, and 11 exhibited significant antibacterial activity in response to some of the tested strains.

Seasonal variation in mycorrhizal fungi colonizing roots of Allium tricoccum (wild leek) in a mature mixed hardwood forest.

The community of arbuscular mycorrhizal (AM) fungi colonizing roots of the forest herb Allium tricoccum Ait. (wild leek) was examined to assess whether colonization varied seasonally and spatially within the forest. Whole plants were collected to coincide with observed phenological stages, and the perennial tissue (i.e., the bulb) was used to analyze total C, N, and P over the growing season. AM fungal community composition, structure, and abundance were assessed in roots by terminal restriction fragment length polymorphism analysis and quantitative PCR. It was found that A. tricoccum rDNA co-amplified using the general AM primers NS31/AM1, and a new primer for qPCR was designed that discriminated against plant DNA to quantify AM colonization. Community structure of AM fungi did not vary seasonally, but did change spatially within the forest, and AM fungal communities were correlated with the presence of overstory tree species. Fungal colonization of roots, however, did change seasonally with a maximum observed in late winter and early spring following leaf emergence. Maximum AM fungal colonization was associated with declines in bulb N and P, suggesting that leaf emergence and growth were responsible for both declines in stored nutrients and increases in AM fungal colonization. Plant N and P contents increased between late summer and early spring while C contents remained unchanged. The observed increase in nutrient content during a time when A. tricoccum lacks leaves indicates that the roots or AM fungi are metabolically active and acquire nutrients during this time, despite an absence of photosynthesis and thus a direct supply of C from A. tricoccum.

IDENTIFICATION OF DIFFERENT FUSARIUM SPP. IN ALLIUM SPP. IN GERMANY.

In 2013 Allium cepa bulbs from different fields in Northern and Southern Germany, seeds and sets from onion breeders were analysed for infestation with Fusarium species. The same investigation was done in 2014 with different edible Allium spp. from local markets. Different Fusarium spp. were isolated and identified by morphological characterisation. 24 different Fusarium spp. were identified. The diversity of Fusarium spp. and the intensity of infestation was higher on edible bulbs compared to the younger sets and seeds. The analysed onions and other edible Allium spp. from local markets showed also high contents of different Fusarium species. The most prevalent identified Fusarium sp. in the analysed Allium spp. in Germany was Fusarium oxysporum which can cause the Fusarium Basal Rot, followed by Fusarium solani. Fusarium proliferatum, which can cause the Fusarium Salmon Blotch in onions, could be detected in about half of the sampled onion fields and in approximately 10% of all analysed onions from fields. Also in the onion sets, on the surface of the seeds and in other edible Allium spp. F. proliferatum could be identified. Besides F. proliferatum, further mycotoxin producing Fusarium spp. like Fusarium equiseti or Fusarium tricinctum were identified. Other Fusarium spp. like Fusarium sporotrichioides and Fusarium poae were first described in Allium sp. in this study. The two most prevalent Fusarium spp. F. oxysporum and F. solani are able to produce mycotoxins like enniatins, fumonisins, moniliformin and T-2 toxins. Fusarium sp. like F. proliferatum, F. equiseti and F. tricinctum are able to produce additional toxins like beauvericins, zearalenone and diacetoscirpenol. This high number of Fusarium spp., which are able to produce a broad spectrum of different mycotoxins, could be a potential health risk for human beings and livestock.

Shading decreases plant carbon preferential allocation towards the most beneficial mycorrhizal mutualist.

Preferential allocation towards the most beneficial mutualist could maintain mycorrhizal mutualism. Context dependence of preferential allocation could then determine environmental patterns in abundance of mycorrhizal mutualists. We assessed the preferential allocation of carbon (C) and differential phosphorus (P) uptake across four light treatments between the host plant Allium vineale and two arbuscular mycorrhizal (AM) fungi within a split-root system. The ratios of C allocation and P uptake between the beneficial and nonbeneficial AM fungi were measured using isotopic labelling. Allium vineale preferentially allocated more C towards roots infected with the most beneficial AM fungus in high light and, in return, received more P from the beneficial fungus. Preferential allocation declined with shading, as A. vineale allocated 25% of labelled C to roots infected with beneficial AM fungi in high light, but only 15% with shading, a similar percentage to that allocated to roots infected with nonbeneficial fungi regardless of shading. Our findings demonstrate that plant preferential allocation towards the most beneficial mycorrhizal mutualist depends upon above-ground resources, suggesting that the abundance of beneficial mycorrhizal fungi will increase with amount of above-ground resources, with implications for mycorrhizal mediation of plant productivity with anthropogenic change.

Utilization of Leek (Allium ampeloprasum var. porrum) for inulinase production.

Inulinase production by Rhodotorula glutinis was carried out in this study, using leek (Allium ampeloprasum var. porrum) as an alternative carbon source due to its high inulin content and easy availability. Taguchi orthogonal array (OA) design of experiment (DOE) was used to optimize fermentation conditions. For this purpose, five influential factors (leek concentration, pH, incubation temperature, agitation speed, and fermentation time) related to inulinase production were selected at four convenient levels. The results showed that maximum inulinase activity was obtained as 30.89 U/mL, which was close to the predicted result (30.24 U/mL). To validate the obtained results, analysis of variance (ANOVA) was employed. Consequently, leek has a great potential as an effective and economical carbon source for inulinase production, and the use of Taguchi DOE enhanced enzyme activity about 2.87-fold when compared with the unoptimized condition.

Cellular imaging of cadmium in resin sections of arbuscular mycorrhizas using synchrotron micro X-ray fluorescence.

Arbuscular mycorrhizal (AM) fungi function as extended roots and take an active part in plant acquisition of nutrients and also soil pollutants, such as heavy metals. The objective of this study was to establish a method to observe the localization of cadmium (Cd) Kα at subcellular levels using X-ray fluorescence (XRF) imaging with a synchrotron irradiation microbeam in resin-embedded sections of mycorrhizas. To evaluate the methodology, distributions of Cd in high-pressure-frozen Lotus japonicus-Rhizophagus irregularis mycorrhizal roots were compared between two treatments; Cd was exposed either to the roots or to the extraradical hyphae. Results showed that, in the latter treatment, Cd was restricted to fungal structures, whereas in the former, Cd was detected in cell walls of the two organisms. Plunge-frozen extraradical mycelium of Gigaspora margarita exposed to Cd showed high signals of Cd in the cell walls and vacuoles, and low in the cytoplasm. With selective staining and elemental mapping by electron-dispersive X-ray spectrometry (EDS), a positive correlation between distributions of Cd and P was revealed in the vacuole, which suggested polyP as a counter ion of Cd. These results indicated that there was no Cd relocation in rapidly frozen resin-embedded materials, therefore supporting the usefulness of this methodology.

In vitro evaluation method for screening of candidate prebiotic foods.

The aim of this work was to develop a simple and rapid in vitro evaluation method for screening and discovery of uncharacterised and untapped prebiotic foods. Using a NMR-based metabolomic approach coupled with multivariate statistical analysis, the metabolic profiles generated by intestinal microbiota after in vitro incubation with feces were examined. The viscous substances of Japanese bunching onion (JBOVS) were identified as one of the candidate prebiotic foods by this in vitro screening method. The JBOVS were primarily composed of sugar components, especially fructose-based carbohydrates. Our results suggested that ingestion of JBOVS contributed to lactate and acetate production by the intestinal microbiota, and were accompanied by an increase in the Lactobacillus murinus and Bacteroidetes sp. populations in the intestine and fluctuation of the host-microbial co-metabolic process. Therefore, our approach should be useful as a rapid and simple screening tool for potential prebiotic foods.

Antioxidant changes of leek (Allium ampeloprasum var. porrum) during spontaneous fermentation of the white shaft and green leaves.

BACKGROUND: Leek is grown for its thickened cylindrical white shaft made up of long leaf bases. Despite the potentially valuable nutritional profile of the green leaves, a large portion remains unused owing its restricted culinary applications. This large quantity of this plant biomass could be valorized given an adequate stabilization method. In this study, we examined leek fermentation with regard to antioxidant changes. RESULTS: The oxygen radical absorbance capacity (ORAC) increased by 62% when the green leaves were fermented for 21 days, while 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity did not increase significantly. Fermentation resulted in an increase of endogenous polyphenolic compounds such as ferulic acid, astragalin, luteolin and naringenin. Moreover, fermentation stimulated the production of a series of polyphenolic compounds that were not present in the fresh leek. The flavour precursors in leek, i.e. methiin and isoalliin, were reduced by 91-93% and 100%, respectively, when spontaneous fermentation was allowed to occur on the white shaft and green leaves. CONCLUSION: Our results demonstrated that application of fermentation resulted in a higher ORAC value and polyphenol content of the leek plant, especially in the green leaves. These results indicate the nutritional relevance of fermentation, which hold promise as a stabilization technique.

Discovery of a new source of resistance to Fusarium oxysporum, cause of Fusarium wilt in Allium fistulosum, located on chromosome 2 of Allium cepa Aggregatum group.

This study was carried out to evaluate the antifungal effect of Allium cepa Aggregatum group (shallot) metabolites on Fusarium oxysporum and to determine the shallot chromosome(s) related to Fusarium wilt resistance using a complete set of eight Allium fistulosum - shallot monosomic addition lines. The antifungal effects of hexane, butanol, and water extraction fractions from bulbs of shallot on 35 isolates of F. oxysporum were examined using the disc diffusion method. Only hexane and butanol fractions showed high antifungal activity. Shallot showed no symptom of disease after inoculation with F. oxysporum f. sp. cepae. The phenolic content of the roots and the saponin content of root exudates of inoculated shallot increased to much higher levels than those of the control at 3 days after inoculation. Application of freeze-dried shallot root exudates to seeds of A. fistulosum soaked in a spore suspension of F. oxysporum resulted in protection of seedlings against infection. Among eight monosomic addition lines and A. fistulosum, FF+2A showed the highest resistance to Fusarium wilt. This monosomic addition line also showed a specific saponin band derived from shallot on the thin layer chromatography profile of saponins in the eight monosomic addition lines. The chromosome 2A of shallot might possess some of the genes related to Fusarium wilt resistance.

Inoculations with arbuscular mycorrhizal fungi increase vegetable yields and decrease phoxim concentrations in carrot and green onion and their soils.

BACKGROUND: As one of the most widely used organophosphate insecticides in vegetable production, phoxim (C(12)H(15)N(2)O(3)PS) is often found as residues in crops and soils and thus poses a potential threat to public health and environment. Arbuscular mycorrhizal (AM) fungi may make a contribution to the decrease of organophosphate residues in crops and/or the degradation in soils, but such effects remain unknown. METHODOLOGY/PRINCIPAL FINDINGS: A greenhouse pot experiment studied the influence of AM fungi and phoxim application on the growth of carrot and green onion, and phoxim concentrations in the two vegetables and their soil media. Treatments included three AM fungal inoculations with Glomus intraradices BEG 141, G. mosseae BEG 167, and a nonmycorrhizal control, and four phoxim application rates (0, 200, 400, 800 mg l(-1), while 400 mg l(-1) rate is the recommended dose in the vegetable production system). Carrot and green onion were grown in a greenhouse for 130 d and 150 d. Phoxim solution (100 ml) was poured into each pot around the roots 14d before plant harvest. Results showed that mycorrhizal colonization was higher than 70%, and phoxim application inhibited AM colonization on carrot but not on green onion. Compared with the nonmycorrhizal controls, both shoot and root fresh weights of these two vegetables were significantly increased by AM inoculations irrespective of phoxim application rates. Phoxim concentrations in shoots, roots and soils were increased with the increase of phoxim application rate, but significantly decreased by the AM inoculations. Soil phosphatase activity was enhanced by both AM inocula, but not affected by phoxim application rate. In general, G. intraradices BEG 141 had more pronounced effects than G. mosseae BEG 167 on the increase of fresh weight production in both carrot and green onion, and the decrease of phoxim concentrations in plants and soils. CONCLUSIONS/SIGNIFICANCE: Our results indicate a promising potential of AM fungi for enhancing vegetable production and reducing organophosphorus pesticide residues in plant tissues and their growth media, as well as for the phytoremediation of organophosphorus pesticide-contaminated soils.

Real-time PCR detection of Holophagae (Acidobacteria) and Verrucomicrobia subdivision 1 groups in bulk and leek (Allium porrum) rhizosphere soils.

In the light of the poor culturability of Acidobacteria and Verrucomicrobia species, group-specific real-time (qPCR) systems were developed based on the 16S rRNA gene sequences from culturable representatives of both groups. The number of DNA targets from three different groups, i.e. Holophagae (Acidobacteria group 8) and Luteolibacter/Prosthecobacter and unclassified Verrucomicrobiaceae subdivision 1, was determined in DNA extracts from different leek (Allium porrum) rhizosphere soil compartments and from bulk soil with the aim to determine the distribution of the three bacterial groups in the plant-soil ecosystem. The specificity of the designed primers was evaluated in three steps. First, in silico tests were performed which demonstrated that all designed primers 100% matched with database sequences of their respective groups, whereas lower matches with other non-target bacterial groups were found. Second, PCR amplification with the different primer sets was performed on genomic DNA extracts from target and from non-target bacteria. This test demonstrated specificity of the designed primers for the target groups, as single amplicons of expected sizes were found only for the target bacteria. Third, the qPCR systems were tested for specific amplifications from soil DNA extracts and 48 amplicons from each primer system were sequenced. All sequences were >97% similar to database sequences of the respective target groups. Estimated cell numbers based on Holophagae-, Luteolibacter/Prosthecobacter- and unclassified Verrucomicrobiaceae subdivision 1-specific qPCRs from leek rhizosphere compartments and bulk soils demonstrated higher preference for one or both rhizosphere compartments above bulk soil for all three bacterial groups.

Fumonisin B(1) production by Fusarium proliferatum strains isolated from Allium fistulosum plants and seeds in Japan.

AIMS: To test the fumonisin B(1)-producing ability of Fusarium proliferatum strains isolated from Welsh onion (Allium fistulosum) plants and seeds of commercial cultivars in Japan and to examine the applicability of PCR-based assays to discriminate between fumonisin B(1)-producing and nonproducing isolates. METHODS AND RESULTS: Fumonisin B(1) levels in 20 Fusarium isolates obtained from Welsh onion plants and seeds of seven commercial cultivars were determined by HPLC. Thirteen of the 20 isolates produced fumonisin B(1). PCR assay with FUM1 gene-specific primers amplified a DNA fragment (700 bp) only from fumonisin-producing isolates. CONCLUSIONS: Fusarium proliferatum isolates that can produce fumonisin B(1) were often associated with wilted Welsh onion plants and seeds of some commercial cultivars. The PCR assay with FUM1 gene-specific primers has the potential to discriminate between fumonisin B(1)-producing and nonproducing isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: This study revealed that F. proliferatum producing fumonisin B(1) is associated with Welsh onion plants and that commercial cultivar seeds may be contaminated with the fungus. PCR amplification of FUM1 gene can be a useful tool for the rapid identification of fumonisin B(1)-producing F. proliferatum isolates.

Preferential allocation to beneficial symbiont with spatial structure maintains mycorrhizal mutualism.

Mutualisms, beneficial interactions between species, are expected to be unstable because delivery of benefit likely involves fitness costs and selection should favour partners that deliver less benefit. Yet, mutualisms are common and persistent, even in the largely promiscuous associations between plants and soil microorganisms such as arbuscular mycorrhizal fungi. In two different systems, we demonstrate preferential allocation of photosynthate by host plants to the more beneficial of two AM fungal symbionts. This preferential allocation could allow the persistence of the mutualism if it confers sufficient advantage to the beneficial symbiont that it overcomes the cost of mutualism. We find that the beneficial fungus does increase in biomass when the fungi are spatially separated within the root system. However, in well-mixed fungal communities, non-beneficial fungi proliferate as expected from their reduced cost of mutualism. Our findings suggest that preferential allocation within spatially structured microbial communities can stabilize mutualisms between plants and root symbionts.

Polyphasic characterization of Xanthomonas axonopodis pv. allii associated with outbreaks of bacterial blight on three Allium species in the Mascarene archipelago.

Based on the number of new reports during the last two decades, bacterial blight of onion (Allium cepa) is considered an emerging disease. The causal agent, Xanthomonas axonopodis pv. allii, is pathogenic to several Allium species after inoculation, but outbreaks worldwide have been primarily reported on onion. We describe a unique epidemiological situation in Réunion Island, France, with concomitant outbreaks on three Allium species, onion, leek (A. porrum), and garlic (A. sativum). There was no host specialization within Allium spp. among strains associated with the three host species. Based on amplified fragment length polymorphism (AFLP) and restriction fragment length polymorphism, strains associated with these outbreaks in Réunion Island were highly related genetically to strains isolated from diseased plant samples and contaminated seed lots in the neighboring island of Mauritius, where the disease has occurred since 1984. All AFLP haplotypes were identified as X. axonopodis pv. allii based on polymerase chain reaction analysis using specific primers, biochemical tests, and/or pathogenicity tests. Two genetically related groups of strains (A and B) that can be distinguished by AFLP, differential utilization of three carbon sources, and xanthomonadin pigment production were detected initially after establishment of the pathogen. In less than 10 years after the establishment of the pathogen there was nearly an extinction of group A strains in Réunion Island, suggesting differences in fitness between strains in the two groups.