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.     

Genetic relationships among strains of <Emphasis Type="Italic">Xanthomonas campestris</Emphasis>pv. <Emphasis Type="Italic">campestris</Emphasis>revealed by novel rep-PCR primers

Novel primers for rep-PCR were developed with the original software and based on `ancient diverged periodical sequences'. Rep-PCR with these primers was applied to study genetic relationships among 51 Xanthomonas campestris strains. The strains were collected from different countries including Russia, Japan, UK, Germany and Hungary. Reference strains of three X. campestrispathovars and five other Xanthomonas species were included. Based on qualitative differences in amplification profiles, the strains were divided into four major groups. Two subgroups recognised within X. campestrispopulation were similar to RFLP haplotypes. The third subgroup included strains of two other pathovariants and Japanese isolates of X. campestris pv. campestriswhile the fourth group comprised the other species of Xanthomonas. The analysis of the diversity within X. campestris resulted in the conclusion that isolates belong to distinct clonal populations (subgroups). The differences between the subgroups of X. campestris were only slightly smaller than between species of Xanthomonas. A PCR fragment about 600 bp amplified by primer KRPN2 was found in nearly all tested strains of X. campestris.SCAR primers designed for this marker produced a single specific band for strains of X. campestris, but not for other Xanthomonas, Pseudomonas and Erwiniastrains tested. Application of the new primer set for rep-PCR offers a rapid, simple and reproducible method for identification of bacterial strains. The X. campestris-specific SCAR primers may be used in diagnostics of this important plant pathogen.     

Development of a specific molecular tool for detecting Xanthomonas campestris pv. musacearum

A specific and rapid diagnostic tool has been developed to detect Xanthomonas campestris pv. musacearum, the causal agent of bacterial wilt of banana. PCR primers were developed from intergenic regions of X. campestris pv. musacearum following its partial sequence. A total of 48 primers were tested for specificity to X. campestris pv. musacearum strains collected from various regions in Uganda. These were also tested for specificity against related Xanthomonas species from the vasicola group, Xanthomonas species pathogenic to other crops, and against those existing saprophytically on banana plants. Seven primer sets (Xcm12, Xcm35, Xcm36, Xcm38, Xcm44, Xcm47 and Xcm48) were found to be very specific to X. campestris pv. musacearum. These primer sets directed the amplification of the expected product for all 52 strains of X. campestris pv. musacearum collected from different locations in Uganda. No amplification products were obtained with unrelated phytopathogenic bacteria or endophytic/epiphytic bacteria from banana. A detection limit of 10³ CFU mL^?1 corresponding to about four cells per PCR reaction was observed when X. campestris pv. musacearum cells were used for all the seven primer sets. The DNA samples from symptomless plant tissues also tested positive with primer set Xcm38. The specific PCR method described here is a valuable diagnostic tool which can be used to detect the pathogen at early stages of infection and monitor disease.     

Differentiation of Xanthomonas species Pathogenic to Sugarcane by PCR-RFLP Analysis

The PCR-RFLP of the 16S-23S rDNA spacer region was used to differentiate Xanthomonas species pathogenic to sugarcane. Strains of X. albilineans, X. campestris pv. vasculorum Types A and B, X. sacchari and Xanthomonas sp. from Trinidad, South Africa and India were examined. The amplification products were digested with Alu I, Hae III, Hpa II and Mbo I and the results showed that the different groups of bacterial strains exhibited distinct RFLP patterns for each tested endonuclease, except X. albilineans and X. sacchari which could only be differentiated from each other by the digestion with Hpa II. The results also allowed the separation of X.c. pv. vasculorum Type A from X.c. pv. vasculorum Type B and strongly suggested that the analyzed Xanthomonas sp. strains belong to X. sacchari. Nine X. campestris (pv. not determined) strains included in this study showed identical profiles to X.c. pv. vasculorum Type A group and DNA–DNA hybridization experiments confirmed these results. PCR-RFLP of the 16S-23S rDNA spacer region could be applied as a reliable method for differentiating the xanthomonads pathogenic to sugarcane.     

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.     

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.     

Metabolic Diversity of Xanthomonas axonopodis pv. vignicola, Causal Agent of Cowpea Bacterial Blight and Pustule

Fifty-five strains of Xanthomonas axonopodis pv. vignicola, isolated from blight and pustule symptoms of cowpea leaves, originating from 11 countries, were characterized for their carbon-source metabolization pattern using the Biolog GN microplate system. Great variation was found between strains according to origin. Dextrin, glycogen and succinamic acid were not used by strains from Benin, Uganda or Thailand, but by all the other strains (excluding two strains from Mozambique), whereas N-acetyl-D-glucosamine and malonic acid were used by the strains from Benin, Uganda and Thailand, but generally not by the other strains. The strains from Benin, Uganda and Thailand, as well as strains from Venezuela, Brazil and Mozambique, clustered separately from the others in multivariate analysis. Nineteen substrates were used by all the strains, 47 not by any strain and 29 only by some strains. No considerable differences were found between strains isolated from blight symptoms and from pustules. Virulence of strains was not related to the metabolic pattern. The Biolog database was not representative of the diversity of X. axonopodis pv. vignicola, since all strains were identified as Xanthomonas campestris, although belonging to eight pathovars, while only eight of nine strains from Benin and both strains from Thailand were identified as X. campestris pv. vignicola. The Biolog system appeared to be useful for characterizing the diversity of X. axonopodis pv. vignicola strains. A set of representative strains based on metabolic and molecular diversity, virulence and geographic origin is suggested for screening for resistant cowpea cultivars.     

Consequences of the taxonomic revision of Xanthomonas axonopodis pv. dieffenbachiae and of the analysis of its associated pathogenicity for the recommendation of regulation (EPPO A2 List)

Xanthomonas axonopodis pv. dieffenbachiae, the causal agent of bacterial blight of Araceae (aroids), is a regulated pest in several countries and is included in the EPPO A2 List. Reference strains of Xanthomonas axonopodis pv. dieffenbachiae have recently been reclassified into the species Xanthomonas phaseoli, Xanthomonas citri and Xanthomonas euvesicatoria on the basis of different features, including multilocus sequence analysis, average nucleotide identity and homology in DNA–DNA hybridization analyses. Based on pathogenicity tests, Constantin et al. (2017) proposed naming the pathogens on aroids as X. phaseoli pv. dieffenbachiae, X. phaseoli pv. syngonii and X. citri pv. aracearum. Recommendations are made on how to deal with these changes for the group of pathogenic bacteria for Araceae. The name Xanthomonas axonopodis pv. dieffenbachiae on the EPPO List should be adjusted to the names proposed in the taxonomic study by Constantin et al. (2016). The current EPPO Diagnostic Standard is directed at strains pathogenic on Anthurium. They mainly belong to X. phaseoli pv. dieffenbachiae, but some also to X. citri pv. aracearum that are not detected by the EPPO Diagnostic Standard. Xanthomonas phaseoli pv. syngonii strains are also aggressive, but with a host range restricted to Syngonium. The pathogenicity specific to aroids of the bacterial isolates reclassified as Xanthomonas euvesicatoria was not confirmed and no pathovar epithet has been retained for these strains.     

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.     

A functional XopAG homologue in Xanthomonas fuscans pv. aurantifolii strain C limits host range

Citrus canker is caused by two Xanthomonas species, Xanthomonas citri, which has become the primary pathogen where citrus canker occurs (type A citrus canker, Xc‐A), and X. fuscans pv. aurantifolii (Xfa), which consists of strains B and C. The B strain is less pathogenic than the A strain, but produces symptoms in all citrus species. The C‐type cankers only infect Key lime (Citrus aurantifolia) and produce a hypersensitive reaction (HR) in grapefruit (Citrus paradisi) leaves. An avirulence gene, avrGf2, was identified in a C strain that was responsible for the HR in grapefruit. AvrGf2 is a member of XopAG effector family and shares 45% identity at amino acid level with another member of the same family, AvrGf1 from strain Xc‐A^w, which was previously shown to elicit an HR in grapefruit. AvrGf2 shares sequence identity with other XopAG effectors present in Xanthomonas vasculorum, Xanthomonas campestris pv. musacearum and Pseudomonas syringae pv. tomato. Mutagenesis of avrGf2 in C strain resulted in a compatible reaction in grapefruit. There was no observable effect on virulence when Xc‐A transconjugants containing either avirulence gene were inoculated on Key lime. Expression of avrGf1 or avrGf2 in Xc‐A resulted in a similar phenotype following infiltration into grapefruit leaves, although the avrGf2 transconjugant elicited a faster HR and lower populations than the transconjugant containing avrGf1. Also, it was shown that all Xfa‐B strains tested contain a transposon in avrGf2 that helps to explain the differences in host range between B and C strains.     

Protein electrophoresis and DNA:DNA hybridizations of xanthomonads from grasses and cereals1

More than 120 Xanthomonas campestris strains pathogenic for grasses and cereals were compared by polyacrylamide gel electrophoresis (SDS-PAGE) of their whole-cell proteins. Genotypic relationships between representative strains of the electrophoretic groups were determined by DNA:DNA hybridizations. Two major groups of bacteria were delineated. The first included X. campestris pv. graminis, pv. arrhenatheri and some isolates from Bromus, which could be differentiated from each other by their protein fingerprints, and also the following pathovars which it was impossible to differentiate by SDS-PAGE: cerealis, hordei, poae, secalis, translucens and undulosa. DNA:DNA hybridizations indicated that significant degrees of DNA-binding (>60% D) exist between all these pathovars. In the second group, strains of X. campestris pv. holcicola, pv. vasculorum and pv. oryzae were related at 40–45% DNA-binding, while strains of pv. oryzae and pv. oryzicola were genotypically highly related (85% D). All the pathovars of this second group could be differentiated from each other by their protein electrophoretic fingerprints.     

Bacterial pathogens of Gramineae: systematic review and assessment of quarantine status for the EPPO region1

Bacterial pathogens of Gramineae principally belong to the genera Clavibacter, Erwinia, Pseudomonas and Xanthomonas, the last being the most important. A general survey of these pathogens is given, with details on nomenclature, symptoms, natural host range, geographical distribution and potential quarantine significance for the EPPO region. The status of Xanthomonas campestris pathovars with overlapping broad and narrow host ranges on Gramineae is discussed. It is proposed to adopt a broad concept of X. campestris pv. translucens and evaluate it as a potential quarantine hazard for the EPPO region.     

Investigation of Genomic Variability of Xanthomonas Arboricola Pv. juglandis by AFLP Analysis

Xanthomonas arboricola pv. juglandis is the causal agent of walnut blight, one of the most important and widespread diseases of Persian (English) walnut (Juglans regiaL.), causing severe damage to leaves, twigs and nuts. To investigate the genomic variability of X. arboricolapv. juglandis, 66 isolates obtained from different countries (England, France, Italy, The Netherlands, Romania, Spain, USA, and New Zealand) were analysed using the Amplified Fragment Length Polymorphism (AFLP) technique. EcoRI and MseI were used as restriction endonucleases. Primers with a core sequence including endonuclease recognition sites and a 3^prime-terminal cytosine selective base for MseI primer, or no selective base for EcoRI primer, were used. Data were analysed by means of a multiple correspondence analysis. A total of 76 amplified polymorphic DNA fragments were used to compute relationships among isolates. The AFLP profiles of X. arboricola pv. juglandis isolates appeared to be reliably distinguishable from X. arboricola pv. pruni and X. arboricola pv. corylina, and from other Xanthomonas species, i.e. X. campestris pv. campestris, X. fragariae, X. hortorum, X. axonopodis pv. vesicatoria. Though this pathogen is associated with one single host genus, a high level of genomic diversity was found. This diversity might be partly explained by the geographic origin. Nevertheless, isolates with different patterns were collected within one country, and similar molecular patterns were found in isolates collected at different sites. However, genetic diversity might be influenced by exchanging vegetative material from different countries. Mixing of X. arboricola pv. juglandis isolates might have partly concealed the influence of the geographic location from which the bacteria were isolated.     

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.     

Biofilm formation and motility of Xanthomonas strains with different citrus host range

Xanthomonas citri subsp. citri (Xcc) strain A is the causal agent of citrus bacterial canker (CBC) on most Citrus spp. and close relatives. Two restricted host range strains of CBC, A^w and A*, from Florida and southwest Asia, respectively, infect Mexican lime. Several studies have linked biofilm formation by Xcc to bacterial colonization prior to and after plant ingress, but none have evaluated connections between biofilm formation and the behaviour of different strains of Xcc on citrus hosts and non‐hosts. In this study biofilm formation and swimming motility were evaluated for citrus pathogenic xanthomonads including wide and restricted host range strains of Xcc, X. alfalfae subsp. citrumelonis (Xac) (the causal agent of citrus bacterial spot) and X. campestris pv. campestris (Xc). Differential biofilm formation was observed in vitro and in planta among the Xanthomonas strains assayed. Minimal medium XVM2 increased biofilm formation, especially for those strains with a host range restricted to Mexican lime. In planta, strains produced more biofilm on leaves or fruits of their host than on non‐hosts. Scanning electron microscopy of biofilms on leaf and fruit surfaces revealed differences in structure of bacterial aggregates with respect to the strain's host range. In addition, swimming motility varied widely depending on the host range of the strain. It was concluded that biofilm formation in vitro and in planta for strains of Xcc and Xac was related to their host range, as these processes affect colonization at the early stages of the infection process.     

Xanthomonas campestris pv. populi,the causal agent of bark necrosis in poplar

A bacterium was isolated from superficial bark necroses on young poplars and its pathogenicity demonstrated by inoculation experiments. The organism was identified asXanthomonas campestris. Cross-inoculations showed that a previously undescribed pathovar was involved. It is suggested to designate this organismX. campestris pv.populi.Samenvatting Uit een oppervlakkige bastnecrose bij jonge populieren werd massaal een bepaalde bacterie geïsoleerd. Met deze bacterie werden gezonde populieren in het veld geïnoculeerd via verwonding van de bast. Als gevolg van de inoculaties ontwikkelden zich bij ongeveer 40% van de geïnoculeerde bomen hetzelfde type bastnecrosen, terwijl bij de controleplanten geen enkele reactie optrad. Uit de kunstmatig verkregen necrosen werd dezelfde bacterie geïsoleerd.Identificatie met biochemische en serologische methoden toonde aan dat de bacterieXanthomonas campestris was.Vervolgens werden in de kas kruisinoculaties uitgevoerd met verschillende xanthomonaden op populier, wilg, kool en geranium. DeX. campestris isolaten uit populier tastten behalve populier ook wilg aan. De andere gebruikte stammen waren waardplant-specifiek, al bleven sommigen ervan minstens acht maanden in leven in een niet-waardplant, evenwel zonder symptomen te veroorzaken. Geconcludeerd wordt, dat de bastnecrosen zijn veroorzaakt door een nog niet beschreven pathovar vanX. campestris. Voorgesteld wordt om deze bacterieXanthomonas campestris pv.populi te noemen.     

Development of a semi-selective medium for isolation of <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> pv<Emphasis Type="Italic">. musacearum</Emphasis> from banana plants

Banana Xanthomonas wilt, caused by Xanthomonas campestris pv. musacearum, is a new threat to banana cultivation in eastern Africa. The causal bacterium grows slowly in culture and is easily overgrown by contaminants. A selective culture medium for isolation of X. c. pv. musacearum will facilitate disease study. A medium that suppressed saprophytic growth and possessed diagnostic characters for the pathogen was developed. Various carbon sources were tested with two isolates of X. c. pv. musacearum, and sucrose was selected as main carbon source. The susceptibility of X. c. pv. musacearum and other bacterial strains was tested with 29 different antibiotics. Cephalexin and cycloheximide had no effect on X. c. pv. musacearum but cephalexin inhibited most of the saprophytes and cycloheximide inhibited the fungal contaminants. Based on these studies, we have developed a semi-selective medium YTSA-CC containing yeast extract (1%), tryptone (1%), sucrose (1%), agar (1.5%), cephalexin (50 mg l⁻¹) and cycloheximide (150 mg l⁻¹), pH 7.0. The pathogen X. c. pv. musacearum was easily identified as yellowish, mucoid and circular colonies on YTSA-CC medium. This simple semi-selective medium was effective for isolation of X. c. pv. musacearum from infected banana tissues and soil, and it should be a valuable tool in ecological and epidemiological studies.     

Identification of Resistance to Common Bacterial Blight Disease on Bean Genotypes Grown in Turkey

Common bacterial blight (CBB) in edible beans (Phaseolus vulgaris), incited Xanthomonas campestris pv. phaseoli, reduces bean yields and seed quality. The main objective of this study was to determine resistance to common bacterial blight in bean genotypes. Twenty-two bean genotypes grown in Turkey including common and snap bean cultivars/lines were collected from different parts of Turkey and tested for resistance against to Xanthomonas campestris pv. phaseoli strain MFD-11. All the common and snap bean lines/cultivars tested were moderately susceptible, susceptible or highly susceptible, except AG-7117 which was found resistant to Xanthomonas campestris pv. phaseoli. This is the first report of a resistance source in a common bean line (AG-7117) against Xanthomonas campestris pv. phaseoli.     

Copper resistance in Xanthomonas campestris pv. campestris affecting crucifers in Trinidad

Fifty-six native isolates collected in 12 farming districts of Trinidad and seven reference strains of Xanthomonas campestris pv. campestris were evaluated for resistance to copper in buffered (pH 7.0) and unbuffered (pH 5.6) nutrient agar media. All isolates and reference strains were pathogenic and elicited typical black rot symptoms on a susceptible variety of Brassica olearceae, ‘Copenhagen Market’. Thirty-four and thirty-three native isolates were highly resistant to copper (growth on?≥?200 ppm copper) in buffered and unbuffered media, respectively; however, all the reference strains were highly susceptible to copper. The mean minimum inhibition concentration for the 56 native isolates was 224.6 ppm copper indicating that high levels of copper resistance are present in X. campestris pv. campestris in Trinidad. The association between growth of the 56 isolates and seven reference stains on buffered and unbuffered media was strong (Pearson’s and Spearman’s r?=?0.93; P?<?0.01) suggesting that either medium can be used to evaluate resistance to copper in X. campestris pv. campestris. There was also a strong association between length of time of continuous applications of copper formulations to treat black rot disease and proportion of the native X. campestris pv. campestris with resistance to copper (Pearson’s r?=?0.96; Spearman’s r?=?0.93); however, there was no association between resistance to copper and aggressiveness at 10 days after inoculation.