Multilocus sequence analysis reveals a novel phylogroup of Xanthomonas euvesicatoria pv. perforans causing bacterial spot of tomato in Iran

A multilocus sequence analysis (MLSA) was performed on five housekeeping genes (fusA, gapA, gltA, lacF and lepA) of 22 Xanthomonas euvesicatoria strains recently isolated from alfalfa, pepper and tomato plants in Iran. In addition, 161 strains isolated worldwide from pepper, poinsettia, rose and tomato plants were included in the analysis. All X. euvesicatoria pv. perforans isolates from tomato plants in Iran clustered in a monophyletic group, although five MLSA haplotypes were detected among them. The Iranian tomato strains presented 10 nucleotide differences in the lepA gene sequences compared to the known worldwide population of X. euvesicatoria pv. perforans. Statistical analyses revealed a recombination event that had occurred in the lepA gene of the strains isolated from tomato in Iran. BOX‐PCR analysis confirmed the inclusion of Iranian tomato strains within X. euvesicatoria pv. perforans. Furthermore, X. euvesicatoria pv. euvesicatoria strains isolated from pepper in Iran differed in one nucleotide in the lepA gene sequence from the known worldwide population of the pathovar, and clustered in a group containing strains isolated in Nigeria. The strains isolated from alfalfa in Iran clustered with the type strain of X. euvesicatoria pv. alfalfae. Altogether, the results reveal the existence of a phylogenetically novel population of X. euvesicatoria pv. perforans in Iran which needs further in‐depth analysis to pinpoint the epidemiological impact of these strains.     

Resistance to Xanthomonas perforans race T4 causing bacterial spot in tomato breeding lines

Tomato (Solanum lycopersicum) is the second most important vegetable crop in the world. Bacterial spot (BS) of tomato, caused by four species of Xanthomonas: X. euvesicatoria, X. vesicatoria, X. perforans and X. gardneri, results in severe loss in yield and quality due to defoliation and formation of lesions on fruits, respectively. Currently management practices do not offer effective control under conditions of high disease pressure. Thus, developing BS resistance is a critical priority for tomato growers in order to minimize crop losses. Sixty‐three advanced tomato breeding lines, heirlooms and wild tomato lines with diverse genetic backgrounds were screened under greenhouse and field conditions for BS resistance using X. perforans race T4, which was found to be a prevalent race in North Carolina. Race T4 isolate 9 was used to inoculate the plants by spraying, and disease severity was measured using the Horsfall–Barratt scale. Tomato lines 74L‐1W(2008), NC2CELBR, 081‐12‐1X‐gsms, NC22L‐1 (2008) and 52LB‐1 showed resistance to BS in the field and/or greenhouse trials. These lines were derived from S. pimpinellifolium L3707. Screening L3707 followed by development of a mapping population and mapping resistance genes might be useful for breeding resistance against BS in future breeding programmes.     

Widespread distribution of Xanthomonas perforans and limited presence of X. gardneri in Brazil

Tomato bacterial spot is caused by Xanthomonas euvesicatoria, X. vesicatoria, X. perforans and X. gardneri. In order to determine the distribution, frequency of occurrence, and diversity of these species in the Brazilian commercial tomato fields, a survey was conducted between 2009 and 2012. In this period, 204 strains were obtained from 33 counties (22 with processing tomatoes and 11 with fresh‐market tomatoes). Pathogenicity tests, BOX‐PCR, PCR with species‐specific primers, and sequence analysis of the avirulence gene avrXv3 were performed in order to identify the strains at species and race level. Xanthomonas perforans predominated among the strains (92%) and was present in most counties. In addition, this species was prevalent in most areas of both fresh‐market tomatoes (63.6% of counties surveyed) and processing tomatoes (95.4% of counties surveyed). Fifteen strains (7.5%) were identified as X. gardneri, which was found mostly in fresh‐market fields located at regions with altitude higher than 900 m, and only one strain of X. euvesicatoria (0.5%) was found in a processing tomato field. High genetic diversity was observed within X. perforans, with 137 BOX‐PCR haplotypes. Race T3 prevailed (97.5%), but reported here for the first time is the occurrence of five strains identified as race T4 in fresh‐market fields in the state of São Paulo. The race T4 phenotype of these strains resulted from the presence of an 859 bp insertion in the avirulence gene avrXv3. This insertion is related to amino acid sequences of a transposase found in X. gardneri, and to amino acid sequences of X. campestris.     

Identification of <Emphasis Type="Italic">Xanthomonas</Emphasis> species associated with bacterial leaf spot of tomato,capsicum and chilli crops in eastern Australia

Several species of Xanthomonas cause bacterial leaf spot, a disease that affects solanaceous crops worldwide. The diversity of 64 Australian isolates of Xanthomonas spp. associated with bacterial leaf spot in tomato, capsicum and chilli crops in eastern Australia was determined using multi-locus sequence analysis of atpD, dnaK, efp and gyrB genes, species-specific PCR assays and biochemical analyses. At least five species of Xanthomonas associated with bacterial leaf spot were identified in Australian tomato, capsicum and chilli crops and their pathogenicity assessed. Phylogenetic and biochemical analyses identified X. euvesicatoria, X. perforans and X. vesicatoria as the most frequently recovered pathogenic species. Non-pathogenic and weakly pathogenic species were also identified. The suitability of the identification methods used and the implications of the detection of these species will be discussed.     

Tomato can be a latent carrier of ‘Candidatus Liberibacter solanacearum’, the causal agent of potato zebra chip disease

‘Candidatus Liberibacter solanacearum’ was recently described as the causal agent of potato zebra chip disease. This pathogen occurs in North America, New Zealand, and Northern Europe on various crops, and may spread to other potato growing regions. Observation on ‘Ca. L. solanacearum’‐infected tomato and potato plants propagated in growth chambers over 5 years indicated that tomato plants (cvs Moneymaker and Roma) can be a latent carrier of ‘Ca. L. solanacearum’. Tomato plants graft‐inoculated with scions from latently infected tomato plants remained symptomless, but tested positive in a species specific PCR assay. ‘Ca. L. solanacearum’ was consistently detected in the top, middle and bottom portion of the symptomless tomato plants, including stem, petiole, midrib, vein, flowers and fruits. In tomato fruits, ‘Ca. L. solanacearum’ was evenly distributed in the tissues at the peduncle and style ends, as well as in the pericarp, and columella placenta tissues. This is the first report that ‘Ca. L. solanacearum’ is present in a plant reproductive organ. In contrast, potato plants (cvs. Jemseg, Atlantic, Shepody, Frontier Russet, Russet Burbank, Red Pontiac, and Russet Norkotah) grafted with scions from the same latently infected tomato plants resulted in typical symptoms of purple top, leaf scorch, and other disease symptoms in plants and brown discoloration in the vascular ring and medullary rays in tubers.     

Occurrence of azoxystrobin‐resistant isolates in Passalora fulva,the pathogen of tomato leaf mould disease

Since 2007, serious damage to tomato from leaf mould caused by Passalora fulva has frequently been observed in commercial greenhouses in Gifu Prefecture, Japan. One of the factors relating to this damage was suspected to be a decrease in azoxystrobin sensitivity of the pathogen. Biological and molecular studies were conducted to characterize fungicide resistance. In in vitro sensitivity tests using mycelial homogenate placed on fungicide‐amended medium, the minimum inhibitory concentrations (MIC) of azoxystrobin for mycelial growth of the isolates divided into two ranges, 0.031–0.5 mg L^?1 and 8–32 mg L^?1. Isolates with MICs within the two ranges were considered as sensitive and resistant, respectively, to azoxystrobin because, in in vivo tests, the percentage protection conferred by this fungicide (100 mg a.i. L^?1) against these isolates was 89.7–100% and 4.5–31.1%, respectively. Resistant isolates had a replacement of phenylalanine with leucine at codon 129 (F129L) in cytochrome b. Forty‐five percent of the 271 isolates collected from 63 tomato greenhouses from 2007 to 2008 were resistant to azoxystrobin. In many greenhouses where the isolation frequency of resistant isolates was 80% or more, azoxystrobin had been used twice per crop for approximately 6 years. In 2012, 27% of the 405 isolates collected were resistant to azoxystrobin, and there was a marked difference in the frequency of occurrence of resistant isolates in the field populations between the three locations sampled. The occurrence of azoxystrobin‐resistant P. fulva isolates (F129L mutants) inflicted considerable damage on greenhouse tomatoes.     

PM 7/110 (1) Xanthomonas spp. (Xanthomonas euvesicatoria,Xanthomonas gardneri,Xanthomonas perforans,Xanthomonas vesicatoria) causing bacterial spot of tomato and sweet pepper

Specific scope

This standard describes a diagnostic protocol for Xanthomonas spp. causing bacterial spot of tomato and sweet pepper (Xanthomonas euvesicatoria, Xanthomonas gardneri, Xanthomonas perforans, Xanthomonas vesicatoria) ¹ .

Specific approval and amendment

Approved in 2012–09.     

A pith necrosis caused by Xanthomonas perforans on tomato plants

Pith necrosis is a common disease of tomato in Europe, mainly caused by Pseudomonas corrugata and other soil-borne species of Pseudomonas. During 2011–2012 a survey was conducted in soil-grown tomato crops in southeastern Sicily (Italy). Plants showed pith necrosis, brown discolouration of the vascular tissues, leaf chlorosis and sometimes wilting of leaves. Thirty bacterial isolates from symptomatic tissues, forming colonies on NA and KB, were identified by morphological, biochemical and physiological tests. Among them, seven isolates were analyzed for their 16S rDNA and 16S–23S spacer region sequence that resulted in 99 % identity to that of the Xanthomonas perforans type strain (GenBank accession number GQ46173over 2.085 bp.). Additional sequences of fusA, gapA, gltA, gyrB, lacF, and lepA from one selected isolate were 100% identical to sequences of the Xanthomonas perforans type strain. X. perforans local isolates showed similar genomic patterns with REP-PCR and fAFLP, and were clearly distinguished from other Xanthomonas spp. type strains. In stem-inoculation assays, bacteria isolated from symptomatic tomato plants identified as P. fluorescens, P. putida, P. marginalis, P. citronellolis, P. straminea, and Pantoea agglomerans induced discolouration of vascular tissues, while Pectobacterium carotovorum subsp. atrosepticum isolates induced soft rot. Conversely, the isolates here identified as Xanthomonas perforans were able to induce pith necrosis, vascular discolouration, longitudinal splits and external lesions on stems. This report of X. perforans causing pith necrosis on tomato represents a potentially serious problem that may limit the productivity of tomato crops.     

Baseline sensitivity of natural populations and resistance of mutants of Xanthomonas oryzae pv. oryzae to a novel bactericide,zinc thiazole

During 2009–2010, a total of 323 isolates of Xanthomonas oryzae pv. oryzae were obtained from rice with symptoms of bacterial leaf blight (BLB) in four provinces (Zhejiang, Jiangsu, Anhui and Hubei) in China. These isolates were tested for baseline sensitivity to zinc thiazole, a novel bactericide with strong antibacterial activity against Xanthomonas. The sampled pathogenic population had similar sensitivity to zinc thiazole (0·1–16·8 mg L^?1) in all four regions and over the whole two‐year study period. The baseline sensitivity was distributed as a unimodal curve with a mean EC₅₀ value of 6·79 ± 1·61 mg L^?1. The risk of mutation to resistance of zinc thiazole in X. oryzae pv. oryzae was further evaluated in vitro and in vivo. Twelve zinc thiazole‐resistant mutants were obtained through ultraviolet (UV) irradiation, culturing on zinc thiazole‐amended nutrient agar (NA) plates, and culturing on zinc thiazole‐treated rice plants. These zinc thiazole‐resistant mutants had resistance factors (RF = EC₅₀ value of a mutant / EC₅₀ value of the wildtype parent of this mutant) of 12·4 to 186·1 with a mean RF value of 44·1. Mutants obtained via UV irradiation, culturing on NA plates and culturing on rice plants had mean RF values of 51·8, 24·5 and 14·4, respectively. All mutants showed decreases in resistance to zinc thiazole after 20 successive transfers on bactericide‐free media or 10 successive inoculation–reisolations on bactericide‐free rice plants. No significant difference was found in bacterial growth and sensitivity to bismerthiazol between zinc thiazole‐resistant mutants and their parents. However, a significant decrease was observed in the pathogenicity of zinc thiazole‐resistant mutants compared with their parents, especially for mutants obtained via UV irradiation.     

Magnesium oxide nanoparticles induce systemic resistance in tomato against bacterial wilt disease

Bacterial wilt is a serious problem affecting many important food crops. Recent studies have indicated that treatment with biotic or abiotic stress factors may increase the resistance of plants to bacterial infection. This study investigated the effects of magnesium oxide nanoparticles (MgO NP) on disease resistance in tomato plants against Ralstonia solanacearum, as well as its antibacterial activity. The roots of tomato seedlings were inoculated with R. solanacearum and then immediately treated with MgO NP; the treated plants showed very little inhibition of bacterial wilt. In contrast, when roots were drenched with a MgO NP suspension prior to inoculation with the pathogen, the incidence of disease was significantly reduced. Rapid generation of reactive oxygen species such as O₂^?˙ radicals was observed in tomato roots treated with MgO NP. Further O₂^?˙ was rapidly generated when tomato plant extracts or polyphenols were added to the MgO NP suspension, suggesting that the generation of O₂^?˙ in tomato roots might be due to a reaction between MgO NP and polyphenols present in the roots. Salicylic acid‐inducible PR1, jasmonic acid‐inducible LoxA, ethylene‐inducible Osm, and systemic resistance‐related GluA were up‐regulated in both the roots and hypocotyls of tomato plants after treatment of the plant roots with MgO NP. Histochemical analyses showed that β‐1,3‐glucanase and tyloses accumulated in the xylem and apoplast of pith tissues of the hypocotyls after MgO NP treatment. These results indicate that MgO NP induces systemic resistance in tomato plants against R. solanacearum.     

The genetic characterization of Xanthomonas arboricola pv. juglandis,the causal agent of walnut blight in Poland

Leaves and fruits of walnut trees exhibiting symptoms of bacterial blight were collected from six locations in Poland. Isolations on agar media resulted in 18 bacterial isolates with colony morphology resembling that of the Xanthomonas genus. PCR using X1 and X2 primers specific for Xanthomonas confirmed that all isolates belonged to this genus. In pathogenicity tests on unripe walnut fruits, all isolates caused typical black necrotic lesions covering almost the entire pericarp. Results of selected phenotypic tests indicated that characteristics of all isolates were the same as described for the type strain of Xanthomonas arboricola pv. juglandis. Genetic analyses (PCR MP, ERIC‐, BOX‐PCR and MLSA) showed similarities between the studied isolates and the reference strain of X. arboricola pv. juglandis CFBP 7179 originating from France. However, reference strains I‐391 from Portugal and LMG 746 from the UK were different. MLSA analysis of partial sequences of the fyuA, gyrB and rpoD genes of studied isolates and respective sequences from GenBank of pathotype strains of other pathovars of X. arboricola showed that the X. arboricola pv. juglandis isolates consisted of different phylogenetic lineages. An incongruence among MLSA gene phylogenies and traces of intergenic recombination events were proved. These data suggest that the sequence analysis of several housekeeping genes is necessary for proper identification of X. arboricola pathovars.     

Identification of grapevine marker genes for early,non‐destructive Eutypa lata infection diagnosis

Eutypa lata is the causal agent of eutypa dieback, a highly damaging trunk disease affecting all grape‐growing areas, with currently neither an efficient curative treatment nor an early non‐destructive diagnostic method. The present work was carried out to discover grapevine genes expressed in response to the presence of E. lata that could be useful to develop an early (before visible foliar symptoms) and non‐destructive (using grapevine leaves) diagnostic tool. Microarray analyses were carried out from (i) infected plants showing characteristic E. lata foliar and vascular symptoms and positive pathogen recovery from vascular lesions (S⁺R⁺), (ii) infected plants showing no symptoms (S^?R⁺), and (iii) symptomless plants with negative pathogen recovery (S^?R^?). Vineyard and greenhouse‐grown plants, naturally or artificially infected respectively, and uninoculated controls were characterized and leaf RNA was hybridized with 15k operon grapevine oligonucleotide microarrays. Among the grapevine genes differentially expressed between S^?R⁺ and S^?R^? plants in greenhouse and vineyard conditions, 10 were highlighted as robust candidate genes for diagnosis: seven were specifically involved in response to infection and three were associated with symptom absence. Five were confirmed to be effective diagnostic marker genes usable in a qRT‐PCR‐based test performed on RNA extracted from grapevine leaves cultivated in either greenhouse or vineyard conditions. Furthermore, their expression profiles in response to infection with E. lata or other major grapevine fungi (Erysiphe necator, Plasmopara viticola, Botrytis cinerea) could be distinguished. The usefulness of these genes to develop an early and non‐destructive method for diagnosis of E. lata infection is discussed with regard to the advantages and drawbacks of previous E. lata diagnostic studies.     

Lack of host specificity of Colletotrichum spp. isolates associated with anthracnose symptoms on mango in Brazil

The aim of the present study was to analyse the genetic and pathogenic variability of Colletotrichum spp. isolates from various organs and cultivars of mango with anthracnose symptoms, collected from different municipalities of São Paulo State, Brazil. Colletotrichum gloeosporioides isolates from symptomless citrus leaves and C. acutatum isolates from citrus flowers with post‐bloom fruit drop symptoms were included as controls. Sequencing of the ITS region allowed the identification of 183 C. gloeosporioides isolates from mango; only one isolate was identified as C. acutatum. amova analysis of ITS sequences showed larger genetic variability among isolates from the same municipality than among those from different populations. fAFLP markers indicated high levels of genetic variability among the C. gloeosporioides isolates from mango and no correlation between genetic variability and isolate source. Only one C. gloeosporioides mango isolate had the same genotype as the C. gloeosporioides isolates from citrus leaves, as determined by ITS sequencing and fAFLP analysis. Pathogenicity tests revealed that C. gloeosporioides and C. acutatum isolates from either mango or citrus can cause anthracnose symptoms on leaves of mango cvs Palmer and Tommy Atkins and blossom blight symptoms in citrus flowers. These outcomes indicate a lack of host specificity of the Colletotrichum species and suggest the possibility of host migration.     

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.     

Arabidopsis thaliana,an experimental host for tomato yellow leaf curl disease‐associated begomoviruses by agroinoculation and whitefly transmission

Tomato yellow leaf curl disease is one of the most devastating viral diseases affecting tomato crops worldwide. This disease is caused by several begomoviruses (genus Begomovirus, family Geminiviridae), such as Tomato yellow leaf curl virus (TYLCV), that are transmitted in nature by the whitefly vector Bemisia tabaci. An efficient control of this vector‐transmitted disease requires a thorough knowledge of the plant–virus–vector triple interaction. The possibility of using Arabidopsis thaliana as an experimental host would provide the opportunity to use a wide variety of genetic resources and tools to understand interactions that are not feasible in agronomically important hosts. In this study, it is demonstrated that isolates of two strains (Israel, IL and Mild, Mld) of TYLCV can replicate and systemically infect A. thaliana ecotype Columbia plants either by Agrobacterium tumefaciens‐mediated inoculation or through the natural vector Bemisia tabaci. The virus can also be acquired from A. thaliana‐infected plants by B. tabaci and transmitted to either A. thaliana or tomato plants. Therefore, A. thaliana is a suitable host for TYLCV–insect vector–plant host interaction studies. Interestingly, an isolate of the Spain (ES) strain of a related begomovirus, Tomato yellow leaf curl Sardinia virus (TYLCSV‐ES), is unable to infect this ecotype of A. thaliana efficiently. Using infectious chimeric viral clones between TYLCV‐Mld and TYLCSV‐ES, candidate viral factors involved in an efficient infection of A. thaliana were identified.     

Development of a lateral flow device for in‐field detection and evaluation of PCR‐based diagnostic methods for Xanthomonas campestris pv. musacearum,the causal agent of banana xanthomonas wilt

Xanthomonas campestris pv. musacearum (Xcm) is the causal agent of banana xanthomonas wilt, a major threat to banana production in eastern and central Africa. The pathogen is present in very high levels within infected plants and can be transmitted by a broad range of mechanisms; therefore early specific detection is vital for effective disease management. In this study, a polyclonal antibody (pAb) was developed and deployed in a lateral flow device (LFD) format to allow rapid in‐field detection of Xcm. Published Xcm PCR assays were also independently assessed: only two assays gave specific amplification of Xcm, whilst others cross‐reacted with non‐target Xanthomonas species. Pure cultures of Xcm were used to immunize a rabbit, the IgG antibodies purified from the serum and the resulting polyclonal antibodies tested using ELISA and LFD. Testing against a wide range of bacterial species showed the pAb detected all strains of Xcm, representing isolates from seven countries and the known genetic diversity of Xcm. The pAb also detected the closely related Xanthomonas axonopodis pv. vasculorum (Xav), primarily a sugarcane pathogen. Detection was successful in both naturally and experimentally infected banana plants, and the LFD limit of detection was 10⁵ cells mL^?1. Whilst the pAb is not fully specific for Xcm, Xav has never been found in banana. Therefore the LFD can be used as a first‐line screening tool to detect Xcm in the field. Testing by LFD requires no equipment, can be performed by non‐scientists and is cost‐effective. Therefore this LFD provides a vital tool to aid in the management and control of Xcm.     

Population structure,genetic diversity,and sexual state of the rice brown spot pathogen Bipolaris oryzae from three Asian countries

Bipolaris oryzae causes brown spot in rice (Oryza sativa) inflicting substantial grain yield losses worldwide. Knowledge of the population structure, genetic diversity and sexual recombination of the fungal pathogen can help to implement effective disease management strategies. In this study, B. oryzae isolates sampled from Iran, the Philippines and Japan were analysed with 12 simple‐sequence repeat (SSR) markers, newly developed from the genome sequence of the fungus. Among the 288 B. oryzae isolates genotyped, 278 unique haplotypes were identified. High genotype numbers (richness) with even distribution (evenness) were found within the collection sites. Both mating types, MAT1‐1 and MAT1‐2, were present in each collection area, and the sexual state was induced under controlled conditions with production of viable ascospores. However, the tests of linkage disequilibrium rejected of the hypothesis of random mating. Discriminant analysis of principal components (DAPC) revealed that the B. oryzae collection formed three clusters, each consisting of isolates from different collection sites. Analysis of molecular variance (amova ) showed that genetic variation among clusters was 18.7%, with the rest of the variation distributed within clusters (R_ST = 0.187, P < 0.001). Statistically significant pairwise genetic differentiation was found between the clusters. These results show that Asian B. oryzae isolates are genetically diverse, and, overall, distributed in three groups. These findings will be helpful in managing the disease and guide the use of representative isolates needed for selection of resistant rice varieties.     

Genomics‐informed design of loop‐mediated isothermal amplification for detection of phytopathogenic Xanthomonas arboricola pv. pruni at the intraspecific level

The objective of this study was to develop a rapid, sensitive detection assay for the quarantine pathogen Xanthomonas arboricola pv. pruni, causal agent of stone fruit bacterial spot, an economically important disease of Prunus spp. Unique targets were identified from X. arboricola pv. pruni genomes using a comparative genomics pipeline of other Xanthomonas species, subspecies and pathovars, and used to identify specific diagnostic markers. Loop‐mediated isothermal amplification (LAMP) was then applied to these markers to provide rapid, sensitive and specific detection. The method developed showed unrivalled specificity with the 79 tested strains and, in contrast to previously established techniques, distinguished between phylogenetically close subspecies such as X. arboricola pv. corylina. The sensitivity of this test is comparable to that of a previously reported TaqMan^? assay at 10³ CFU mL^?1, while the unrivalled speed of LAMP technology enables a positive result to be obtained in <15 min. The developed assay can be used with real‐time fluorescent detectors for quantitative results as well as with DNA‐staining dyes to function as a simplified strategy for on‐site pathogen detection.     

Development of a tomato plant resistant to Clavibacter michiganensis using the endolysin gene of bacteriophage CMP1 as a transgene

The bacteriophage CMP1 endolysin gene (lys), encoding, a peptidase that was shown to effectively reduce Clavibacter michiganensis by specifically hydrolysing its murein, was transferred into tomato plants by Agrobacterium‐mediated transformation. The presence of the gene was verified by PCR and the gene product was confirmed in immunoblots and stably expressed over three generations. Transgenic tomato plants did not show disease symptoms after infection with C. michiganensis subsp. michiganensis, despite the fact that small amounts of bacteria could still be identified in xylem sap and leaf extracts, although in significantly reduced amounts.     

Morphological and molecular diversity of Colletotrichum spp. causing pepper spot and anthracnose of lychee (Litchi chinensis) in Australia

Since the 1980s a new disease has been affecting Australian lychee. Pepper spot appears as small, black superficial lesions on fruit, leaves, petioles and pedicels and is caused by Colletotrichum gloeosporioides, the same fungus that causes postharvest anthracnose of lychee fruit. The aim of this study was to determine if a new genotype of C. gloeosporioides is responsible for the pepper spot symptom. Morphological assessments, arbitrarily‐primed PCR (ap‐PCR) and DNA sequencing studies did not differentiate isolates of C. gloeosporioides from anthracnose and pepper spot lesions. The ap‐PCR identified 21 different genotypes of C. gloeosporioides, three of which were predominant. A specific genotype identified using ap‐PCR was associated with the production of the teleomorph in culture. Analysis of sequence data of ITS and β‐tubulin regions of representative isolates did not group the lychee isolates into a monophyletic clade; however, given the majority of the isolates were from one of three genotypes found using ap‐PCR, the possibility of a lychee specific group of C. gloeosporioides is discussed.