JOURNAL OF PLANT PATHOLOGY

Online ISSN: 1125-4653
Publications
Southern hybridization of the nusA gene in clover phyllody phytoplasma genome. Genomic DNA was probed with a digoxigenin (DIG)-labeled 0.8 kbp DNA fragment amplified, in the polymerase chain reaction primed by primer pair PhNusF1/PhNusR5, from CPh phytoplasma DNA. M, DIG-labeled molecular weight marker. H!, undigested healthy plant DNA control. H*, digested healthy plant DNA control. CPh!, undigested CPh phytoplasma genomic DNA. CPh*, digested CPh DNA. *, DNA digested with HindIII.
Properties of ORFs in the clone CPh163 DNA and their deduced products.
Phylogenetic trees constructed by the Neighbor-Joining method of 16S rRNA gene sequences (left) and NusA amino acid sequences (right) from clover phyllody (CPh), onion yellows (OY-M), aster yellows (AY-WB), blueberry stunt (BBS3), tomato big bud (BB), paulownia witches' broom (PaWB), and maize bushy stunt (MBS) phytoplasmas; Acholeplasma palmae; Spiroplasma kunkelii, five Mycoplasma and one Ureaplasma species, and 28 walled bacteria. Numbers indicate numbers of amino acids in full length NusA proteins where known.
Amplification of 1 Kbp nusA gene sequences from phytoplasmas belonging to group 16SrI, subgroups 16SrI-A (tomato big bud phytoplasma, BB), 16SrI-B (aster yellows, AY [AY1]; chrysanthemum yellows, CY; maize bushy stunt, MBS), and 16SrI-C (CPh, clover phyllody phytoplasma). M, 1 Kb DNA Ladder. W, polymerase chain reaction (PCR) mixture devoid of template DNA. Polymerase chain reactions were primed by oligonucleotide pair PhNusF2/PhNusR5.
RFLP analysis of nusA gene sequences amplified in polymerase chain reactions (PCRs) primed by oligonucleotide pair PhNusF2/PhNusR5 from phytoplasma strains, BB, AY (AY1), CY, MBS and CPh. Products from PCRs were digested with restriction endonucleases MseI and Tsp509I. M, φX174 RFI DNA HaeIII digest size standard, fragment sizes (bp) from top to bottom: 1353, 1078, 872, 603, 310, 281, 271, 234, 194, 118, 72.
Article
Phytoplasmas are cell wall-less prokaryotes that exist as obligate parasites and pathogens of insect vectors and plants. In their descent from walled ancestors in the Bacillus/Clostridium group, phytoplasmas evolved some of the smallest known bacterial genomes. In this study, we cloned and sequenced nusA transcription factor gene sequences from clover phyllody (CPh) and other phytoplasmas and from Acholeplasma palmae, a nonphytopathogenic wall-less bacterium. The CPh nusA gene was flanked at its 5'-end by a hypothetical protein gene and smpB (small protein B), and at its 3'-end by a hypothetical protein gene that may be coordinately regulated with nusA and infB (translation initiation factor). The predicted 357-aa NusA protein of CPh phytoplasma was significantly smaller than those of Mycoplasma spp. and similar in size to NusA of Clostridium spp., Bacillus spp., and A. palmae. A phylogenetic tree based on NusA proteins indicated that phytoplasmal and acholeplasmal nusA genes diverged from a common ancestor. Amplification and RFLP analysis of nusA gene sequences, and phylogenetic analysis of NusA proteins indicated that use of nusA may assist in detection and differentiation of distinct lineages within group 16SrI, 'Candidatus Phytoplasma asteris'-related strains.
 
Article
European law provides that, in organic farming, organically produced seed should be used. Therefore new sanitation treatments need to be developed which do not use classical fungicides but still produce seed free from pathogens which can strongly affect yield of the crop. Greenhouse trials were carried out in order to test the efficacy of different seed treatments alternative to chemicals against Colletotrichum lindemuthianum causing anthracnose on bean and Ascochyta pisi causing Ascochyta blight on pea, respectively. Resistance inducers, commercially formulated micro-organisms, non-formulated selected strains of different micro- organisms (fungi, bacteria and yeasts) and plant extracts were applied as dry or liquid seed treatments on naturally infested seeds. Seedling emergence and disease incidence and/or severity were recorded. Possible suppression of Ascochyta blight in peas was masked by the high rate of infection (around 20%), so almost all seed treatments turned out to be ineffective in controlling infection, with the exception of treatments with thyme oil and a strain of Clonostachys rosea. C. lindemuthianum infection was successfully controlled by all resistance inducers. However, they also caused a significant reduction of plant emergence. Among the micro-organism formulations, Bacillus subtilis-based formulations provided the best protection to anthrachnose. Some bacterial strains, a disease-suppressive saprophytic strain of Fusarium oxysporum and the mustard powder-based product Tillecur™ proved to be effective against bean anthrachnose. Tillecur™ (Schaette AG, Bald Waldsee, Germany) and thyme oil are promising for application in integrated pest management and could possibly be used in organic farming
 
Article
Soft rot is a major disease of calla lily (Zantedeschia spp.) and other important crops worldwide. In this report, the bacterial isolate ZT0505 proved to be a soft rot pathogen of calla lily growing around Kunming (subtropical China) and was identified as Pectobacterium carotovorum subsp. carotovorum. The weight of macerated tuber tissue caused by inoculation of the isolate and incubation for 36 h at 22, 25, 28 and 32°C, was 0.21, 0.62, 0.67, and 0.60 g/tuber, respectively. The extent of tuber maceration was significantly less (65.0-68.7%) at 22°C than at 25- 32°C, while 28°C yielded the highest amount of macerated tissue. The bacterium grew faster at 32°C than at 22- 28°C. The highest pectate lyase (PL) activity of isolate ZT0505 was found at 28°C, a value much higher than the 14-17°C range at which P. carotovorum subsp. carotovorum strains from more temperate regions are usually found to have the highest PL activity. Temperature effect on PL activity tallied with that on tuber maceration. After 24 h incubation at pH 6.3, 7.0, 7.3, the weight of macerated tissue was 0.17, 0.61, 0.61 g/tuber, respectively. The extent of maceration increased when lengthening the incubation time. Tuber maceration at pH 6.3 was significantly less (54.9-72.1%) than at pH 7.0 and 7.3 after 24- 36 h incubation. Bacterial growth at pH 6.3 was significant slower than at pH 7.0 and 7.3. A good correlation between PL activity and extent of tuber maceration at the various pHs was found only when PL activity assays were conducted at similar pHs.
 
A. Mild diffuse spots in a leaf of C5 inoculated with PPV-Rec (second year after inoculation, June 2004). B. Severe yellow spots and rings in a leaf emerged from a non-transgenic bud (IB) infected with PPV-Rec (second year after inoculation, June 2004). C. Very mild difuse spots in a leaf of C5 inoculated with PPV-Rec (fifth year after inoculation, July 2007). D. Severe yellow spots and rings in a leaf g emerged from a non-transgenic bud (IB) infected with PPV-Rec (fifth year after inoculation, July 2007).  
Article
Transgenic plum (Prunus domestica L.) clone C5 was grafted on virus-free St. Julien rootstocks and inoculated by bud grafting with the recombinant strain (PPVRec) of the Plum pox virus (PPV). Non inoculated C5 plums were used as controls. Transgenic plums infected with PPV-Rec and control plants were evaluated for six years in the open field. PPV symptoms, mild diffuse spots and rings, appeared on the basal leaves the second year after inoculation. The presence of PPV was confirmed by DAS-ELISA, ISEM, and RT-PCR. Severe PPV symptoms appeared in the leaves of shoots grown from non-transgenic inoculum buds. A reduction of symptoms and a decrease of the relative PPV concentration, as determined by DAS-ELISA in transgenic C5 plants, were observed from the third to the fifth year after virus inoculation. Very mild symptoms in older leaves of the basal branches of trees subsided by the end of the vegetative period, and were difficult to observe. The level of PPV detection by DAS-ELISA dropped from June to August together with a reduction of symptoms. PPV was undetectable by DAS-ELISA in more than half of the trees by the end of August. PPV-Rec was detected by RT-PCR in the basal and middle parts of the C5 scions. The upper leaves did not contain a detectable level of the virus. These results show the high level and stability of PPV resistance in graft-inoculated C5 trees.
 
Map of Chile showing the regions where grapevines were surveyed for the presence of the most economically important viruses. III: Región Atacama; IV: Región Coquimbo; V: Región Valparaíso; VI: Región Libertador General Bernardo O'Higgins; VII: Región Maule; RM: Región Metropolitana.
Results of virus testing by ELISA.
RT-PCR results of ELISA-negative samples.
RT-PCR results of the assay for GLRaV-2-RG and GRSPaV.
Article
Grapevines from six Chilean regions were surveyed for virus diseases and tested for the presence of the most important viruses. ELISA testing of 2535 samples and confirmatory RT-PCR of some ELISA-negative samples from symptomatic and symptomless vines gave the following infection rates: 6.36% for Grapevine fanleaf virus (GFLV); 4.67% for Grapevine leafroll associated virus 1 (GLRaV-1); 16.05% for Grapevine leafroll associated virus 2 (GLRaV-2); 6.41 % for Grapevine leafroll associated virus 3 (GLRaV-3); 0.26% for Grapevine leafroll associated virus 7 (GLRaV-7); 14.99% for Grapevine fleck virus (GFkV); 5.57% for Grapevine virus A (GVA) and 0.78% for Grapevine virus B (GVB). Strawberry latent ringspot virus (SLRSV) Tomato ringspot virus (ToRSV) and Arabis mosaic virus (ArMV) were not detected. Overall infection in the surveyed Chilean grapevines, considering ELISA and RT-PCR, was 32.35%. Virus infection ratio obtained from ELISA analysis in the six regions, varied between 21.19% (Region Metropolitana) and 74.26% (Coquimbo). RT-PCR was used for detection of the Red Globe strain of GLRaV-2 and Grapevine rupestris stem pitting associated virus (GRSPaV), and to confirm and extend ELISA results. GVB, GFkV, GRSPaV, GLRaV-2 RG and GLRaV-7 are new records for Chile.
 
Article
Pre-treatment of pearl millet seedlings with a suboptimal concentration of the downy mildew pathogen Sclerospora graminicola inoculum induced resistance that protected plants (62%) from subsequent infection by the optimal concentration of the pathogen. Prior inoculation apart from offering protection, enhanced vegetative and reproductive growth parameters of induced resistant plants. Studies on resistance induction in different cultivars of pearl millet suggested that the resistance achieved was independent of the cultivar and not related to its constitutive resistance. Increase of β-1,3-glucanase and peroxidase activity was associated with resistance induction in pearl millet.
 
Article
The carbohydrate-containing polymer 1,3;1,6-β-Dglucan was obtained by transformation of laminaran from the alga Laminaria cichorioides with endo-β-1,3- glucanase from marine mollusks. In electron microscope observations of phosphotungstic acid-stained preparations from Nicotiana tabacum cv. Samsun leaves inoculated with a mixture of Tobacco mosaic virus (TMV) (1 μg/ml) and glucan (1 mg/ml) or with TMV alone, we found that such preparations contained, along with virus particles of normal diameter (about 18 nm), abnormal (swollen and thinner) particles. The highest number of thin viral particles was found in dips from leaves inoculated with TMV together with glucan. It is suggested that this may be caused by a glucan-mediated increase of TMV particle proteolysis in infected leaves.
 
Article
Phytophthora cactorum was identified as the causal agent of crown and root rot of apple varieties grafted onto MM. 106 rootstock in the orchards of Fars province, Iran. In a survey conducted in 2012, 3–5% of apple trees at 3- to 6-year-old were found dead and 10–12% of older ones showed declining symptoms. Field trials were carried out to investigate the effect of post-plant soil solarization on the reclamation of apple trees on infected rootstocks. Experiments were conducted in drip-irrigated Golden Delicious apple orchards in Soghad and Kodian districts for 1 to 4 wk. in the July 2013 and 2015 using transparent polyethylene sheets for soil tarping. In both trials soil temperature raised by 10 to 16 °C in the upper 25 cm layer of top soil and resulted in the apparent eradication of P. cactorum population and reclamation of apple trees in the subsequent year. Except in 1 wk. soil tarping treatment, apple trees showed significantly increased growth responses in 2, 3 and 4 wk. tarping treatments as compared to untreated control. The best results were obtained in the 4 wk. soil tarping treatment. Attempts to isolate P. cactorum from soil and roots of 2–4 wk. tarped plants were unsuccessful.
 
Article
A strain of Bacillus amyloliquefaciens denoted Lx-11 was found to possess biocontrol activity against rice bacterial leaf streak (BLS) caused by Xanthomonas oryzae pv. oryzicola. Lx-11 secreted three kinds of lipopeptides including surfactin, bacillomycin D and fengycin, which exhibted antibacterial activity against X. oryzae pv. oryzicola. The antibacterial activity could be associated with surfactin-lipopeptides, and was practically abolished in surfacin-deficient mutants. In addition, the defense-related genes PR-1a, PR-1b, NPR1 and PAL were concurrently expressed in the leaves of rice after treatment with Lx-11. Results suggest that Lx-11 triggers a systemic immunization activity. Lx-11 significantly reduced the incidence of BLS, control efficacy ranging from 60.2% to 70.6%, i.e. better than that afforded by bismerthlazol in field experiments. We conclude that B. amyloliquefaciens Lx-11 might be a promising biocontrol agent and should be further studied.
 
PCR-RFLP analysis of 16S-23S rDNA of representative Xanthomonas strains with AluI. М: DNA ladder, 100 bp; lanes 7, 15: X. gardneri representative strains forming profile I; lanes 14, 19: X. euvesicatoria representative strains forming profile I; lanes 1-6, 8-13, 16-18: X. vesicatoria representative strains forming profile II; lane 20: X. vesicatoria NBIMCC 2427 (type strain). 
PCR-RFLP analysis of 16S-23S rDNA of representative Xanthomonas strains with AluI. М: DNA ladder, 100 bp; lanes 1, 2, 5, 8-11, 13, 16, 18-20: X. euvesicatoria representative strains forming profile I; lanes 3, 4, 6, 7, 12, 14, 15, 17: X. vesicatoria representative strains forming profile II. 
Article
Bacterial spot of pepper and tomato plants is one of the most important constraints limiting crops yield in Bulgaria and Macedonia. Therefore,early identification of the pathogen is necessary for the control and prevention of the disease. In order to explore the strength of 16S-23S ITS rDNA PCR-RFLP as an approach for identification and differentiation of the causative agents of bacterial spot 262 Bulgarian and Macedonian strains pathogenic of pepper and tomato plants were used. The strains were previously identified as Xanthomonas euvesicatoria (132 strains),Xanthomonas vesicatoria (115 strains),and Xanthomonas gardneri (15 strains). Each restriction analysis resulted in two profiles that grouped the used strains. The restriction with AluI endonuclease differentiated X. vesicatoria isolates from the other three species. The enzyme HpaII separated the X. euvesicatoria strains. A combination of three restriction endonucleases (AluI,MboI,HpaII) successfully differentiated the four species described as causative agents of bacterial spot. Full text available from http://sipav.org/main/jpp/index.php/jpp/article/view/3713
 
Article
The full-length 16S rRNA gene region of a new "Candidatus Liberibacter" strain was PCR- amplified from tubers of potato plants showing zebra chip (ZC) disease symptoms and also from the potato psyllid [Bactericera (= Paratrioza) cockerelli Sulc], the presumptive vector of the ZC disease causal agent. Sequences of the amplicons from ZC diseased potato and potato psyllids were 100% identical but differed from "Ca. L. asiaticus" (Las) and "Ca. L. africanus" (Laf) by 4% and from "Ca. L. americanus" (Lam) by 6.0-6.3%. Neighbor-joining analysis placed the ZC disease-associated sequences in a monophyletic clade consisting of all known "Ca. Liberibacter" spp., positioned in the tree basal to a node shared exclusively by Las and Laf but proximal to a node shared by Lam and all other "Ca. Liberibacter" spp. Here we present evidence for the association between a new "Candidatus Liberibacter" strain and ZC disease in field samples. The relationship of this Liberibacter to the other Liberibacter bacteria detected in psyllids, potato and other solanaceous crops as well as in citrus remains to be determined.
 
Article
During 2011-2013, 7-55% incidence of sesame phyllody and witches' broom symptoms were observed on sesame plants in nine states of India. Seventeen sesame symptomatic samples were analyzed by polymerase chain reaction with phytoplasma specific primers, amplifying 16S rRNA and secA genes. Pairwise sequence comparison and phylogenetic analyses of 16S rRNA and secA gene sequences classified them with aster yellows (16SrI) and peanut witches' broom (16SrII) phytoplasma groups. Further virtual RFLP analysis of 16S rDNA sequences allowed finer classification of the sesame phytoplasma strains into 16SrI-B, 16SrII-C and 16SrII-D subgroups. 'Candidatus Phytoplasma asteris' of the subgroup 16SrI-B was present in sesame plants in two states viz., Uttar Pradesh and West Bengal, while 16SrII-C subgroup phytoplasma was found in sesame plants grown in four states viz., Uttar Pradesh, Madhya Pradesh, Chhattisgarh and Rajasthan. Phytoplasma belonging to 16SrII-D subgroup was found as the most widely distributed phytoplasma strain on sesame from five states viz., Delhi, Rajasthan, Gujarat, Tamil Nadu and Maharashtra. This is the first report of subgroup level classification of phytoplasma strains from sesame in India.
 
Article
Mexico, which is considered the origin and the biodiversity center of the family Agavaceae, hosts 117 of the 155 known species. Agave tequilana Weber cv. Azul is a national icon and the most widely cultivated species for the production of tequila, a widely known alcoholic beverage. In spring 2013, a disease of A. tequilana was observed in Tala, (Jalisco, Mexico) which greatly reduced its availability for tequila production. Affected plants showed extensive chlorosis, followed by yellowing and necrosis of stem and leaf tissues. As the disease progressed, leaves collapsed and hung downwards around the central head. To investigate the possibility of a phytoplasma infection, DNA was extracted according to Lee et al. (1993) from 25 symptomatic and 25 symptomless plants. A nested PCR was performed using two universal primer sets specific for the phytoplasma 16S rRNA gene, i.e. R16mF2/R16mR1 followed by R16F2n/R16R2 (Gundersen and Lee, 1996). The expected 1,200 bp product was obtained from 88% of the symptomatic plants. The PCR products were cleaned with a Wizard kit (Promega, USA) and cloned in Escherichia coli using a TOPO-TA cloning kit (Invitrogen, USA) in accordance with manufacturer’s instructions. When nucleotide sequences of the amplified products (accession Nos KJ156364, KJ156365) were compared with those available in GenBank, it was found that the agave phytoplasma was most similar (99.98%) to the Texas phoenix palm phytoplasma (USA, JF791816) and Sabal mexicana decline phytoplasma (Mexico, GU473588). Phylogenetic and putative restriction site analysis of 16Sr DNA indicated that the phytoplasma associated with A. tequilana is closely related to the lethal yellows 16SrIV group. To our knowledge, this is the first report of a 16SrIV group phytoplasma associated with a disease of A. tequilana.
 
Article
Lethal yellowing (LY) represents a serious threat to coconut in Mexico, Central America and the Caribbean islands. It is caused by a phytoplasma of group 16SrIV, and particularly subgroups -A and -D have been the major phytoplasmas affecting coconut palms grown in the southeast of Mexico. Therefore, is important to reliably detect the 16SrIV group of phytoplasmas not only for diagnosis purposes but also to improve the understanding of pathogen-plant-vector pathosystems. The 16S ribosomal operon genes have been particularly used as the targets for probes and primers for universal phytoplasma detection. However, its conservative nature has imposed the search of non-ribosomal genes to support a finer characterization and differentiation of phytoplasmas, particularly those closely related and one alternative has been the GroEL gene for the specific detection of subgroups 16SrIV-A and 16SrIV-D. Sensitive, rapid and reliable singleplex and duplex assays were developed based on real-time PCR (TaqMan) targeting both the GroEL gene and 16S rRNA. The assay performed with higher sensitivity compared to conventional nested-PCR when analysing DNA extracts obtained from LY symptom bearing palms. The LY 16S TaqMan probe assays amplified DNA from both subgroups, while the LY GroEL TaqMan probe assay only amplified DNA from subgroup 16SrIV-A. The duplex TaqMan probe assay is a novel option for the simultaneous detection and quantification of LY phytoplasmas of 16Sr subgroups -A and -D, which are the phytoplasmas that predominantly affect palms in Mexico.
 
Article
Potato (Solanum tuberosum L.) is a widely cultivated and commercially important vegetable crop in India where it is cultivated in an area of about 1.34 million ha with a total production of about 24.7 million tons. Phytoplasma diseases of potato have been reported from several parts of the world as associated with organisms belonging to different 16Sr DNA group/subgroups. In December, 2011, little leaf disease symptoms were observed in potato crops grown in the Kushinagar district of Uttar Pradesh (India). Phytoplasmas were detected by PCR using P1/P7 primers (Deng and Hiruki, 1991; Schneider et al., 1995) which yilded amplicons ca. 1.8 kb in size only from four symptomatic samples. All four amplicons were directly sequenced from both ends and the sequence was deposited in GenBank under the accession No. KC312703. The little leaf potato phytoplasma isolate shared high 16S rDNA sequence identity (99%) with several isolates of Candidatus Phytoplasma asteris (16SrI group) from different parts of the world. Virtual PCR analysis using the web tool iPhyClassifier (Zhao et al., 1999) assigned the isolate to subgroup 16SrI-B. A number of crop and non-crop species have been reported to be infected by Ca. P. asteris in India (Raj et al., 2011). To the best of our knowledge, this is the first report of the occurrence of Ca. P. asteris (16SrI-B) in association with the little leaf disease of potato in India.
 
16S rRNA gene sequences from 'Candidatus Phytoplasma'-related species used in this study.
Phylogenetic analysis of 16S rRNA gene sequences from 23 phytoplasma strains classified in diverse subgroups of group 16SrI ('Candidatus Phytoplasma asteris'-related strains), and Acholeplasma palmae, employing A. palmae as the outgroup. Phytoplasma classification is according to Lee et al. (1998), Marcone et al. (2000), and Samuitiene et al. (2007). Strains OnP1 and OnP2 were classified in the present study. Arrows indicate 16S rRNA gene sequences from strains, except AV2192, that are classified in subgroup 16SrI-L. Broken arrows indicate rRNA operon sequences of subgroup 16SrI-L strains that exhibited virtual RFLP patterns characteristic of subgroup 16SrI-B; rrnA and rrnB indicate the operons from which the analyzed 16S rDNA sequences were derived. For nucleotide sequences derived from intragenomic sequence-heterogeneous rRNA operons, the RFLP pattern type of the given nucleotide sequence is noted in parentheses. Subgroup classification of each strain is given as a letter not in parentheses. For strain AV2192 classified in subgroup 16SrI-L, nucleotide sequence is available for one of its two sequence-heterogeneous operons; classification is according to Marcone et al. (2000). Numbers at nodes are bootstrap (confidence) >500 values. *, unclassified. Phytoplasma strain names and GenBank accession numbers for rDNA sequences used in constructing the tree are given in Table 1.
Article
Onions (Allium cepa L.) grown for seed production in the Kaunas region of Lithuania exhibited mild yellowing of leaves and stems, stunting, phyllody, and proliferation of flowers. RFLP and sequence analysis of PCR-amplified 16S rRNA, ribosomal protein (rp), and secY genes revealed the presence of phytoplasmas belonging to subgroups 16SrI-A (rpI-A) and 16SrI-L (rpI-B, secYI-B). The results indicated that phytoplasma strains in subgroup 16SrI-A (rpI-A) have potential to damage onions in Europe, as well as in North America, and for the first time demonstrated onion as a host for subgroup 16SrI-L. Subgroup 16SrI-L was distinguished based on a composite HinfI RFLP pattern of 16S rDNA that revealed the presence of two sequence heterogeneous rRNA operons in this subgroup, thus showing the significance of composite RFLP patterns for phytoplasma identification and classification. A single nucleotide polymorphism (SNP) in the first base of one HinfI recognition site (5'-GANTC-3') marked the divergence of major phylogenetic branches, supporting the concept that SNPs provide powerful molecular markers of phytoplasma evolution.
 
Article
Sunflowers (Helianthus annuus L.) exhibiting symptoms of stem fasciation and cluster of multiple inflorescences were observed in Baicheng, Jilin province, China. Coincidently, cockleburs (Xanthium strumarium L.), a weed in the sunflower fields, showing similar symptoms were observed as well. Based on successful amplification of 16S rRNA and secY genes of phytoplasmas, the sunflower fasciation disease (SFD) and cocklebur fasciation disease (CFD) were determined to be associated with phytoplasmas. Based on virtual and actual RFLP analysis of the F2nR2 region of the 16S rRNA genes, the SFD and CFD associated phytoplasmas were classified into a novel 16SrI subgroup, designated 16SrI-AI. This was the first report of SFD associated phytoplasma in China, and the first evidence of phytoplasma infecting cocklebur worldwide.
 
Article
The article “First report of a 16SrII-A phytoplasma infecting Spermacoce exilis in China”, written by Yan Li and Wang Chen, was originally published electronically on the publisher’s internet portal (currently SpringerLink) on 7 May 2018 with open access.
 
Article
The aim of this study is the molecular characterization of a phytoplasma associated with cabbage stunt and spatial analyses of symptomatic plants in the field. Detection by nested PCR, sequencing of 16S rRNA gene and conventional and virtual RFLP, as well as phylogenetic analysis revealed that a 16SrIII-J phytoplasma was associated with cabbage plants that exhibited stunting, reduced size, malformation or failure to form head, reddening leaves, sprout proliferation and vessel necrosis. The spatial analyses demonstrated that the dispersion pattern of symptomatic plants was aggregated and the pathogen introduced from adjacent areas. Phytoplasmas of 16SrIII group were also detected in leafhoppers of the species Atanus nitidus collected from cabbage fields and adjacent areas, evidencing that these insects are possibly potential vectors of this fastidious bacteria found in cabbage plants. These findings confirmed that a subgroup 16SrIII-J phytoplasma is associated with cabbage stunt. In addition, the spatial analyses indicated that the disease emerges from primary foci located in external areas and progress toward the center of the fields. The discovery of the etiological agent of the cabbage stunt together with the type of dispersion of the disease represent a relevant contribution to improve the management strategies aiming harm reduction.
 
Article
Lethal yellowing (LY) is a fatal disease that affects coconut and other palm species in the Americas. Phytoplasmas associated with this disease belong to group 16SrIV. Reliable detection of group 16SrIV phytoplasmas is important for diagnostic purposes and to increase understanding of pathogen-plant-vector pathosystems. The present study describes the development of a TaqMan/real-time PCR assay for detection and quantification of selected 16SrIV subgroups affecting palms in the Americas. The specificity of the assay was assessed on DNA extracts from LY-infected palms in the Americas, coconut lethal phytoplasma-associated diseases in Africa and other phytoplasma-infected plants. Successful amplification was obtained only with DNA extracts from palms infected by LY phytoplasmas that were sampled in the Americas, belonging to the 16SrIV group, subgroups A, D and E. No amplification was obtained from DNA of palms sampled in Africa and phytoplasma-infected plants of other 16Sr groups. The assay was compared with conventional nestedPCR on DNA extracts from 36 palms. The real-time PCR assay showed higher sensitivity as phytoplasmas were detected in several nested-PCR negative and in all the nestedPCR positive samples. The assay was also used to evaluate accumulation of LY phytoplasma DNA in different tissues of palms showing LY symptoms. The highest concentration was found in the trunk, followed in decreasing order by primary root apex, mature inflorescences -1 and -2, inflorescences -3 to -7, spear leaf, flag leaf and mature leaf. The present TaqMan/real-time PCR assay represents a new alternative for LY phytoplasmas detection and quantification, offering high specificity and improvements in sensitivity.
 
Article
Conocarpus erectus, also known as buttonwood, is widely planted as an ornamental plant in southern Iran. In June 2020, symptoms of stem fasciation, shoot twisting and leaf rolling were observed in plants in Bandar Abbas (Hormozgan Province, Iran), with an incidence level of 10%. This study was carried out to identify the associated phytoplasma and its vector. Orosius albicinctus were collected from symptomatic C. erectus plants and 5 adults per plant were caged on ten periwinkle plants for 60-80 days. DNA was extracted from fifteen symptomatic, ten asymptomatic plants and periwinkle (at the end of transmission assays), followed by nested PCR using P1/P7 and R16F2n/R16R2 primers. The PCR products of expected sizes (1.8 and 1.25 kb, respectively) were detected from the symptomatic plants and periwinkles but not from the asymptomatic or negative control. Three sequences of nested PCR amplicons were deposited in GenBank under accession numbers MT712209-MT712211. BLAST search showed their 100% identity with the pigeon pea witches’ broom phytoplasma sequence (AF248957). Restriction fragment length polymorphism (RFLP) analysis with AluI, HhaI, HinfI, HpaII, MseI, RsaI, Sau3AI and TaqI enzymes of R16F2n/R16R2 amplicons and the comparison of the obtained profiles with those of the reference strains (Gundersen et al. 1996) showed that the buttonwood phytoplasma is a member of 16SrIX-A subgroup. Furthermore, this phytoplasma was experimentally transmitted to healthy periwinkle plants by the leafhopper Orosius albicinctus, with the 30% transmission efficiency. The association between 16SrII-D phytoplasma and diseased buttonwood has been reported from Iran (Azimi et al. 2017), however, this is the first report of the detection of 16SrIX-A phytoplasma and its insect vector in buttonwood in the world. Given that C. erectus is widely planted as ornamental species in southern Iran where the presence of O. albicinctus is common, this may present a high risk for an outbreak.
 
Article
Naturally diseased grapevines exhibiting symptoms of grapevine yellows disease were observed in Israel and Greece. Infection of the plants by phytoplasmas (formerly mycoplasmalike organisms, MLOs) was indicated by amplification of phytoplasma DNA in nested polymerase chain reactions (PCR) primed by phytoplasma-universal and group-specific oligonucleotide pairs. Restriction fragment length polymorphism (RFLP) analysis of amplified 16S rDNA and putative restriction site analysis of the nucleotide sequence of 16S rDNA from stolbur (STOL) phytoplasma indicated that the phytoplasmas in the diseased grapevines and STOL phytoplasma were affiliated with 16S rRNA group 16SrXII (stolbur phytoplasma group), subgroup A (formerly subgroup G in group 16SrI), a genetically distinct strain population unreported in North America.
 
Article
This study was conducted to assess the antimicrobial activity of the essential oils of Rosmarinus officinalis, Thymus daenensis, Foeniculum vulgare, Mentha spicata, Mentha piperita and Pelargonium graveolens against Pseudomonas syringae pv. syringe (Pss). The chemical composition of essential oils of six plant species grown in Iran was analyzed by GC-MS and their inhibitory effects were studied on Pss strains IVIA 773–1 and W1 and Pseudomonas fluorescens CHA0, using the disc-diffusion method. The essential oils showed various antimicrobial activities based on the size of zone of inhibition against pathogenic and nonpathogenic bacteria on Petri plates. The minimum inhibitory concentration was evaluated by the micro broth dilution method. The results showed that all essential oils have an inhibitory effect on multiplication of the three bacterial strains tested. According to the present findings the studied essential oils have a potential to be used as antimicrobial agents.
 
Article
The names of all plant pathogenic bacteria which have been effectively and validly published in terms of the International Code of Nomenclature of Bacteria and the Standards for Naming Pathovars are listed to provide an authoritative register of names for use by authors, journal editors and others who require access to currently correct nomenclature. Included are species, subspecies and pathovar names and details of type and pathotype strains reported from 1980 to 2007. An explanation of how to use this list is provided. In recent years the taxonomy of plant pathogenic bacteria has been extensively revised. For some taxa there are several valid synonyms. Unless otherwise stated, the most recently published name is used in this list as the reference name (in bold italic) to which all other synonyms are referred. This does not mean that the reference name is always to be preferred. A synonym, i.e., a previously published name for the same organism, may represent a classification considered by individual scientists or groups to give a more coherent taxonomy and may be used. This list is presented by the International Society of Plant Pathology Committee on the Taxonomy of Plant Pathogenic Bacteria, Carolee T. Bull Convener.
 
Article
PCR-based methods offer advantages over more traditional diagnostic tests, in that organisms do not need to be cultured prior to their detection and protocols are highly sensitive and rapid. Consequently, there is a shift in research towards DNA-based techniques. Although reports already exist on a variety of PCR-based fingerprinting assays used to analyse the genetic diversity of bacterial populations and define their relationships, this review focuses on the general use of PCR in phytobacteriology for detection and diagnosis purposes. An updated and detailed list of published PCR protocols for detection and identification of plant-pathogenic bacteria is presented and discussed, aimed at facilitating access to information that could be particularly useful for diagnostic laboratories. This compilation includes and discusses 246 articles published between 1989 and 2007 addressing 23 genera, more than 50 species, 10 subspecies and more than 40 pathovars.
 
Article
The disease brown rot of potato, caused by the bacterium Ralstonia (Pseudomonas) solanacearum (Rsol), Race 3, biovar 2 (R3b2), was found for the first time in 1992 in a potato field in The Netherlands and caused an outbreak in the warm summer of 1995 that appeared to be connected to use of contaminated irrigation water as in other outbreaks in western Europe at that time. The Dutch Plant Protection Service (PPS) immediately took action and started to test all traded seed for (latent) infections, applied strict control measures upon positive detections, and started an intensive survey of surface water contamination. In later years the control measures and testing procedures for Rsol in different substrates, laid down in an EU Directive, were followed. The PPS also conducted intensive applied research (in cooperation with the University of Wageningen, Plant Research International, and foreign research Institutions, also in the framework of an EU-SMT project on the EU brown rot testing method to unravel the complex epidemiology of the pathogen and to improve its detection and identification. These actions have led over a ca. 15-year period to a drastic reduction, actually a functional eradication (only one single finding in 2009/2010 and 2010/2011), of the disease from the production system. A main factor in the combat was a nation-wide irrigation ban for seed potatoes since 2005. The PPS coordinated two EU-funded projects, where the UK, The Netherlands, Belgium and France assisted Egypt in implementing a brown rot safe potato production system. In this project substantial epidemiological research was conducted. This article wants to be a reflection on the work done and the results obtained by the PPS and has a glance into the future, where it will be indicated that the persisting presence of Rsol in surface water necessitates an enduring alert and actively maintained control and survey system. The main lessons learned are: stay away, if possible, from surface water; use disease-free (tested) and certified seed; apply strict hygiene; handle/grade and store seed and ware/industry potatoes separately; compensate growers or enable them to insure against the disease; invest pro-actively in emergency plans and in up to date diagnostic expertise, education and advice; maintain an active and statistically meaningful survey and control system; perform a regular survey in ware and industry potatoes, greenhouse host crops and surface water.
 
Article
Genomospecie 8, sensu Gardan et al. (1999), includes Pseudomonas avellanae, P. syringae pv. theae and P. s. pv. actinidiae. To further characterize this genomospecies, 14 P. avellanae, three P. s. pv. theae and 18 P. s. pv. actinidiae strains were analysed by multilocus sequence typing (MLST) using gapA, gltA, gyrB and rpoD gene fragments. These strains were also checked for the presence/absence of 38 effector protein genes based on the corresponding sequences of P. syringae. pv. Tomato DC3000 and P. syringae pv. phaseolicola 1448A. Nutritional tests and a comparison of the 16S rDNA gene sequences deposited at NCBI database were also done to detect possible differences. MLST analysis, based on 2.5 kb sequences, revealed that P. s. pv. theae and P. s. pv. actinidiae are more closely-related to one another than to P. avellanae. This technique clearly revealed that the P. s. pv. actinidiae strains causing the current severe epidemics in Italy are different from those of past outbreaks in Japan and central Italy. Nine effector protein genes were displayed by all strains of genomospecies 8. However, each pathogen of this genomospecies displays some distinctive effector protein genes. HopA1 and hopH1 are unique to P. s. pv. actinidiae strains of the recent epidemics of bacterial canker on Actinidia chinensis and A. deliciosa in Italy. A triplet, in position 461-463 of the 16S rDNA gene, is different in P. s. pv. actinidiae, namely GAT, and in P. avellanae and P. s. pv. theae, namely ATC. Contrarily to P. s. pv. theae and P. s. pv. actinidiae, P. avellanae did not utilize sorbitol.
 
Article
Bacillus strains are known to produce cyclic lipopeptides that are capable of providing protection against plant pathogens. Such abilities could be utilized to protect greenhouse tomatoes against diseases including bacterial canker caused by Clavibacter michiganensis subsp. michiganensis (Cmm). In the present study, Bacillus velezensis strain 1B-23 and Bacillus sp. strain 1D-12 were assessed for their potential biocontrol abilities against Cmm strain 98–1 (Cmm98–1). Both Bacillus strains interfered with growth of Cmm98–1 in vitro, as determined by agar plate assays to screen for microbial antagonism. Inoculation of Cmm98–1 infected tomato plants with B. velezenis 1B-23 or Bacillus sp. 1D-12 lead to significantly reduced disease incidence in a greenhouse setting. Liquid Chromatography coupled to Mass Spectrometry (LC-MS) of 1B-23 and 1D-12 extracts identified [Leu7]surfactin C13 (often called surfactin A), [Leu7]surfactin C14 (often called surfactin B) and [Leu7]surfactin C15 (often called surfactin C) in fractions of extracts that inhibited growth of Cmm98–1.
 
Number of isolates of different races among isolates of Phytophthora infestans from Estonia (2004-2007).
Frequencies of specific compatibility (virulence) to potato R-genes in isolates of Phytophthora infestans from differ- ent locations and years in Estonia.
Article
A collection of 432 single-lesion isolates of Phytophthora infestans collected from blighted potato foliage during 2004-2007 in Estonia, were analyzed for virulence (all isolates), mating type (424 isolates) and response to metalaxyl (412 isolates). The samples came from 25 fields comprising conventional production in central, northern, southern, south-eastern and southwestern regions and from untreated experimental field trials at Jõgeva Plant Breeding Institute in eastern Estonia. Of the isolates 33% were A2 mating type. Both mating types were present in all fields; the frequency of A2 mating type varied from 3% to 71%. In the context of specific virulence, the Estonian population had a very low frequency of virulence against R5 (17%) and R9 (3%). The most common pathotype was 1.3.4.7.10.11. A subgroup of 57 isolates was assessed for mtDNA haplotype and RG57 fingerprints. Three mitochondrial DNA haplotypes, i.e. Ia (51%), IIa (42%) and IIb (7%), were found. Twenty-one RG57 fingerprints were detected. The four most common fingerprints represented more than half of the isolates (67%). On the basis of combined markers, thirty-three multilocus genotypes were identified, of which 81% were detected only once. Genotypic diversity measured by the normalized Shannon diversity index was 0.79. The data indicate that the Estonian population of P. infestans is diverse, having a large number of multilocus genotypes and both mating types within fields, with potential for sexual recombination and spread of fungicide resistance.
 
Article
The International Society of Plant Pathology Committee on the Taxonomy of Plant Pathogenic Bacteria has responsibility to evaluate the names of newly proposed pathovars for adherence to the International Standards for Naming Pathovars of Phytopathogenic Bacteria. Currently, the Comprehensive List of Names and the List of New Names of Plant Pathogenic Bacteria provide the authoritative register of names of bacterial plant pathogens. In this manuscript we up-date the list of names by cataloguing and evaluating names of plant pathogenic bacteria published in 2011 and 2012. We provide those names that have been validly and effectively published in this time frame, the proposed names that we judged to be invalid, and names published earlier that did not make the previous lists.
 
Article
The purpose of this study was to characterize Phytophthora infestans isolates obtained from diseased tomato tissue in Oman from 2013 to 2015. Late blight caused severe epidemics on tomatoes in Oman in 2013–2014 and 2014–2015. Prior to 2013, late blight had not previously been reported from Oman. Isolates collected from diseased tomato plants in commercial production fields in Oman were assayed for a series of traits. All fifty-five isolates tested for mating type were determined to be A1. A subset of nine isolates analysed for allozymes, sensitivity to mefenoxam, DNA fingerprint as detected with random genome probe fifty-seven (RG57), and microsatellite genotypes revealed a diverse population. A range of sensitivity to mefenoxam was observed and one multi-locus genotype (MLG) was detected in four of the nine isolates. The A2 strain Blue_13, which was recently identified in India and thought to be a possible source of the pathogen, was not detected.
 
Article
The development of plant virology as a science is, almost totally, a 20th century phenomenon. Early papers on plant viruses emphasize pathogenicity and ultra-filterability and report transmission by leafhopper and aphid vectors, and from soil or through seed. Successive advances were largely driven by access to new techniques. For example, biochemical approaches, and local-lesion and serological assays, led to purification and crystallization of virus particles, and their identification as nucleoproteins; electron microscopy and X-ray crystallography revealed their detailed structure. Biological tests revealed mite, nematode, chytrid and plasmodiophorid vectors. Viroids were discovered. Proof of the infectivity of viral RNA and DNA, followed by sequencing of complete viral genomes, allowed their genes and control elements to be defined. Although some viral genomes were monopartite and others multipartite, each typically encoded proteins involved in replication, movement within the plant, particle structure and, commonly, transmission by vectors. The processes involved in vector transmission, viral replication, disease induction and resistance, and the role of gene silencing, were then studied intensively. Genetic variability was analyzed and short-term evolution explored. A soundly-based virus taxonomy was devised and now includes over 70 plant virus genera containing some 800 species. Virus ecological and epidemiological research established patterns of virus spread and aided the development of control measures. These include schemes for producing virus-tested stocks of vegetatively propagated plants, quarantine regulations, application of vector-controlling chemicals to crops and use of virus-resistant cultivars that contain conventional or man-made resistance genes. These and other changes in agricultural practices have resulted in substantial increases in crop yields and food security. Since about 1950, results of research on viruses infecting the whole spectrum of host taxa have been integrated to constitute virology, with plant virology contributing many seminal discoveries. The biological phase of plant virology has progressed to molecular biological and molecular genetic phases, and plant viruses have become tools in molecular biology, cell biology and biotechnology.
 
Article
The bacterial blight gene locus Xa 27 in rice has two striking differences in the presumed promoter-raised resistant/susceptible allele expression. We used a functional marker targeting the 25 deletion at 18 bp upstream of the putative TATA box and assayed the allelic status at Xa 27 in 572 rice genotypes. The alleles evaluation obtained from 191 cultivars tested may contribute to little information of the resistance specificity associated with allele diversity. The allele frequency associated with complete resistance in the indica was different from that associated with complete resistance in the japonica.
 
Article
The biological and molecular characteristics of 8 strains of biovars 2A of Ralstonia solanacearum from the Iranian province of Isfahan and of 7 isolates of biovar 2T from the province of Khuzestan were investigated morphologically, physiologically, biochemically, and using pathogenicity tests, BOX-PCR, ERIC-PCR and RFLP. Biovar 2A strains showed irregular, creamy-white and highly fluid colonies with pink center on TZC medium, slow spreading necrosis in hypersensivite reaction (HR) after 48 h, produced melanin in tyrosin test, were less pectolytic and did not utilize ribose and tryptophan. By contrast, biovar 2T strains had small size, less fluid, round and deep red colonies with a thin white margin, did not produce melanin, had a higher pectolytic activity, rapid HR on tobacco leaves after 24 h and utilized ribose and tryptophan. Comparative analysis of the BOX-PCR and ERIC-PCR fingerprint of the 15 strains at the 45% similarity level, revealed two main clusters. comprising all biovar 2A strains (cluster 1) or all biovar 2T strains (cluster 2). RFLP profiles of both biovers did not show any polymorphism. Typically, biovar 2A isolates were more pathogenic and bacterial density was higher in the xylem of all hosts tested. There was no significant difference in biovar 2A and 2T survival in loamy-clay soil, which was up to 186 days under controlled conditions.
 
Top-cited authors
Joseph Bové
  • Université Victor Segalen Bordeaux 2
D. Boscia
  • Italian National Research Council
Franco Nigro
  • Università degli Studi di Bari Aldo Moro
Maria Saponari
  • Italian National Research Council
Carmine Marcone
  • University of Salerno