Resistance of Common Bean (Phaseolus vulgaris) to Bacterial Wilt Caused by Curtobacterium flaccumfaciens pv. flaccumfaciens

Bacterial wilt caused by Curtobacterium flaccumfaciens pv. flaccumfaciens is an important new disease of common bean (Phaseolus vulgaris) in western Canada. Both yellow and orange variants of the pathogen were found in the region. A controlled environment study was conducted to assess 124 common bean cultivars and lines from eight market classes for resistance to the yellow and orange variants of the pathogen, using the hilum injury/seed inoculation method. Results of the screening tests showed significant (P < 0.05) differences in resistance to bacterial wilt among the cultivars or lines. The great northern line L02E317, the great northern cultivar Resolute and pinto lines L02B662 and 999S‐2A, were highly resistant to both variants of the pathogen, with disease severity indices of 0 on a rating scale of 0 (no wilt symptoms) to 5 (dead seedling). Resistant cultivars or lines were found among black, great northern, pink, pinto, small red and Flor de Mayo bean market classes. The study concludes that new bacterial wilt‐resistant germplasm exists among Canadian bean cultivars and lines, and constitutes a valuable resource for breeding common beans for resistance to both yellow and orange variants of C. flaccumfaciens pv. flaccumfaciens.     

First Report of Curtobacterium flaccumfaciens pv. flaccumfaciens Causing Cowpea Bacterial Wilt in Iran

During surveys in cowpea fields of Marand County, East Azerbaijan province, Iran, in the summer of 2013, a suspected bacterial disease was observed on cowpea leaves as tan spots and interveinal necrotic lesions surrounded by chlorotic margins. The disease was of high incidence where some fields had been fully destroyed and severity of the disease in some fields had reached up to 70%. Gram‐positive, yellow‐pigmented, coryneform bacteria were isolated from infected leaves. Pathogenicity of isolates was confirmed on 20‐day‐old cowpea (cv. Khoy) plants, and they were identified as Curtobacterium flaccumfaciens pv. flaccumfaciens based on biochemical test results confirmed using specific PCR primers. This is the first report of C. flaccumfaciens pv. flaccumfaciens, the causal agent of cowpea bacterial wilt in Iran.     

Biological Control of Bacterial Wilt of Bean Using a Bacterial Endophyte, Pantoea agglomerans

Indoor studies were conducted to determine the potential use of Pantoea agglomerans isolate LRC 8311 as a biocontrol agent for control of bacterial wilt of bean caused by Curtobacterium flaccumfaciens pv. flaccumfaciens. Soaking seeds of great northern bean cv. US1140 in a suspension of 3 × 10⁸ cfu/ml P. agglomerans resulted in thorough endophytic colonization of the entire bean seedling from root to apical stem after 7 days, regardless of whether the inoculated seeds were hilum injured or not. Colonization of seedlings by P. agglomerans increased seedling height after 10 days, and had no negative effect on seedling emergence. Treatment of hilum‐injured bean seeds of great northern bean cv. US1140 or navy bean cv. Morden003 with a mixture of P. agglomerans + C. flaccumfaciens pv. flaccumfaciens resulted in a high rate of colonization of seedlings by P. agglomerans, reduced frequency of infection by C. flaccumfaciens pv. flaccumfaciens, improved seedling emergence and height, and reduced disease severity, compared with seeds treated with the wilt pathogen alone. Application of P. agglomerans as a soil drench 24 h after planting was also effective in suppressing bacterial wilt in some instances, but was generally not as effective as seed treatment. The study suggests that seed treatment with P. agglomerans may be an effective and practical method for control of bacterial wilt of bean.     

Curtobacterium flaccumfaciens pv. flaccumfaciens on Soybean in Germany – A Threat for Farming

Plant diseases caused by Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff) are distributed in North and South America as well as in South and East Europe and occur mostly on beans (Phaseolus vulgaris). This is the first report of Cff on soybean in Germany. Cff was detected in complex with Pseudomonas syringae pv. glycinea on field‐grown soybeans that were not treated with pesticides. Cff, the causal agent of bacterial tan spot disease, was identified by 16S rDNA sequencing and by artificial infection and re‐isolation from the host plants soybean (Glycine max) and bush bean (Phaseolus vulgaris).     

Development of a PCR test for the detection of Curtobacterium flaccumfaciens pv. flaccumfaciens

A chromosomal DNA library of the bacterial pathogen of bean, Curtobacterium flaccumfaciens pv. flaccumfaciens NCPPB 559 was constructed in the plasmid pGEM-7Zf(+). Several clones were identified that hybridised to all Curtobacterium flaccumfaciens pathovars including: C. f. betae, C. f. flaccumfaciens, C. f. oortii, C. f. poinsettiae and, in addition, to some strains of Clavibacter michiganensis subsp. insidiosus and Clavibacter michiganensis subsp. One of these clones (pPMP-26), after subsequent digestion with restriction endonucleases EcoRI/SacI, yielded a fragment of approximately 0.2 Kb (pPMP-26D) that hybridised specifically to C. f. flaccumfaciens and not to any of the other plant pathogenic members of the order Actinomycetales or any of the other prokaryotic bean pathogens tested. This fragment was subcloned and sequenced, analysis of the resultant 198 bp sequence showed that no significant homology existed with any other sequence currently deposited in public databases. Further analysis of these data facilitated the design of PCR primers which were subsequently tested against a wide range of plant pathogenic actinomycetes and other prokaryotic bean pathogens. Results show that these primers are highly specific for all strains of C. f. flaccumfaciens with no cross-reaction to strains from any other bacterial taxa tested.     

Age-stage,two-sex life table of the tomato looper,Chrysodeixis chalcites (Lepidoptera: Noctuidae), on different bean cultivars

The tomato looper, Chrysodeixis chalcites (Esper), is one of the polyphagous pests of several economically crops worldwide. Two-sex life table parameters of C. chalcites reared on eight bean cultivars including white kidney bean (cultivars Daneshkadeh and Dehghan), red kidney bean (cultivars Goli and Naz), common bean (cultivars Khomein, Talash and Sadra) and cowpea (cultivar Mashhad) were studied under laboratory conditions (25 ± 1 °C, 65 ± 5 % RH, a 16:8-h light–dark photoperiod). The shortest larval period of C. chalcites was 14.15 days on common bean Sadra. The longest and shortest development time of total preadult was on white kidney bean Daneshkadeh and common bean Sadra (25.77 and 23.42 days, respectively). The highest total fecundity was on common bean Sadra (674.4 eggs), and the lowest was observed on white kidney bean Daneshkadeh (136.7 eggs). The intrinsic rate of increase (r _m ) ranged from 0.0976 to 0.1599 female/female/day, which was lowest on white kidney bean Daneshkadeh and highest on common bean Sadra. The net reproductive rate (R ₀) was highest on common bean Sadra (265.82 offspring) and lowest on white kidney bean Daneshkadeh (46.88 offspring). The results revealed that the cultivar Daneshkadeh was unsuitable host to C. chalcites in comparison to the other cultivars tested.     

Phaseolus lunatus,a New Host of Xanthomonas axonopodis pv. phaseoli in Iran

Common bacterial blight caused by Xanthomonas axonopodis pv. phaseoli (Xap) is one of the most destructive diseases limiting the production of common bean (Phaseolus vulgaris) in Iran. The disease has previously been described on common bean and mung bean from several regions of Iran, including the central plain and south‐western provinces. In this study, lima bean (Phaseolus lunatus cv. Christmas) plants are being reported as a new natural host of Xap in East Azerbaijan and West Azerbaijan Provinces, northwestern Iran. Disease symptoms consisted initially of water‐soaked spots that progressed to irregular necrotic lesions with chlorotic margins. Infection was observed to affect up to 40% of plants in the field. Identification of the pathogen was based on the biochemical and molecular characteristics, as well as the pathogenicity tests. To our knowledge, this is the first report of X. axonopodis pv. phaseoli causing common bacterial blight on lima bean plants in Iran.     

Virulence and type III effector diversities of Xanthomonas citri pv. fuscans and X. phaseoli pv. phaseoli in Brazil

Common bacterial blight (CBB) is caused by four genetic lineages belonging to two species of Xanthomonas, namely Xanthomonas citri pv. fuscans (includes fuscans, NF2 and NF3 lineages) and X. phaseoli pv. phaseoli (lineage NF1). A collection of 117 strains of Xanthomonas isolated from common bean plants grown in several producing regions of Brazil, between 2007 and 2016 was established. For species and lineage identification, the following tests were performed: multiplex PCR with a set of four specific primer pairs, pathogenicity tests on susceptible cultivar BRS Artico and phylogenetic analysis based on housekeeping gene sequences. The presence of the two species were confirmed among the 117 strains, being 62 non-fuscans strains (NF1, NF2 and NF3) and 55 fuscans strains of X. citri pv. fuscans. To select a set of representative strains for the virulence assay, a PCR-based analysis of effector diversity was performed with 42 strains belonging to the two species. PCR with primers for xopL, avrBsT, xopE2 and xopE1 genes were positive for all strains, while for the other six effectors there was variation. Six distinct effector profiles were detected, and one strain representing each type was inoculated in 15 common bean cultivars with varying levels of resistance to CBB. The fuscans strains showed uniformity in their effector profiles and were the most virulent. The phylogenetic analyses of our strain collection revealed that all genetic variants of CBB pathogens (NF1, NF2, NF3 and fuscans) are present in Brazil, with significant variability in virulence to common bean cultivars.     

Mixed Infection and Natural Spread of ‘Candidatus Phytoplasma asteris’ and Mungbean Yellow Mosaic India Virus Affecting Soya Bean Crop in India

Suspected phytoplasma and virus‐like symptoms of little leaf, yellow mosaic and witches’ broom were recorded on soya bean and two weed species (Digitaria sanguinalis and Parthenium hysterophorus), at experimental fields of Indian Agricultural Research Institute, New Delhi, India, in August–September 2013. The phytoplasma aetiology was confirmed in symptomatic soya bean and both the weed species by direct and nested PCR assays with phytoplasma‐specific universal primer pairs (P1/P6 and R16F2n/R16R2n). One major leafhopper species viz. Empoasca motti Pruthi feeding on symptomatic soya bean plants was also found phytoplasma positive in nested PCR assays. Sequencing BLASTn search analysis and phylogenetic analysis revealed that 16Sr DNA sequences of phytoplasma isolates of soya bean, weeds and leafhoppers had 99% sequence identity among themselves and were related to strains of ‘Candidatus Phytoplasma asteris’. PCR assays with Mungbean yellow mosaic India virus (MYMIV) coat‐protein‐specific primers yielded an amplicon of approximately 770 bp both from symptomatic soya bean and from whiteflies (Bemisia tabaci) feeding on soya bean, confirmed the presence of MYMIV in soya bean and whitefly. Hence, this study suggested the mixed infection of MYMIV and ‘Ca. P. asteris’ with soya bean yellow leaf and witches’ broom syndrome. The two weed species (D. sanguinalis and P. hysterophorus) were recorded as putative alternative hosts for ‘Ca. P. asteris’ soya bean Indian strain. However, the leafhopper E. motti was recorded as putative vector for the identified soya bean phytoplasma isolate, and the whitefly (B. tabaci) was identified as vector of MYMIV which belonged to Asia‐II‐1 genotype.     

Occurrence and Characterization of the Bacterial Spot Pathogen Xanthomonas euvesicatoria on Pepper in Iran

We report in this study for the first time the occurrence of bacterial spot of pepper in Iran and both phenotypic and genetic characterization of its causal agent, Xanthomonas euvesicatoria. Pepper plants grown in 15 of 30 surveyed private gardens and commercial fields were infected by the pathogen in Marand County, East Azerbaijan Province, north‐western Iran. The obtained strains of X. euvesicatoria had different amylolytic and pectolytic activities compared with those reported for this species elsewhere. Pathogenicity tests showed that strains isolated from diseased pepper are able to infect tomato, in addition to pepper. Host range of the pathogen was assessed on eight annual plant species including crops and weeds by measuring the population dynamics. The host range assessment showed that in addition to pepper and tomato, known hosts of X. euvesicatoria, the Iranian strains were able to colonize a number of new hosts such as nightshade and common bean. In contrast, none of them were able to build up their population on cowpea, eggplant, bindweed and zucchini. All X. euvesicatoria strains obtained in this study were sensitive to copper sulphate and streptomycin at concentrations higher than 20 and 50 mg/l, respectively. Phylogenetic analyses of the strains using the sequences of gyrB and hrpB genes confirmed their species as X. euvesicatoria. Given a direct commercial trade of fresh solanaceous vegetables between Iran and Turkey, it is hypothesized that the pathogen entered north‐western Iran from eastern parts of Turkey through infected plant materials. Finally, the role of prevention – based on the use of healthy planting materials and resistant and/or tolerant plant varieties – to contain the potential disease epidemics is discussed.     

Production and Application of Syringomycin E as an Organic Fungicide Seed Protectant against Pythium Damping‐off

Syringomycin E (SRE) is a cyclic lipodepsinonapeptide with potent antifungal activity and is produced by certain strains of Pseudomonas syringae pv. syringae. In this study, its potential as an organic‐compatible agrofungicide and vegetable seed treatment against the soilborne pathogen Pythium ultimum var. ultimum was examined. A variant of P. syringae pv. syringae strain B301D with enhanced SRE‐producing capabilities was isolated and grown in a bioreactor with SRE yields averaging 50 mg/l in 40 h. SRE was extracted and purified through a large‐scale chromatography system using organic‐compatible processes and reagents. The minimum concentrations of the purified product required to inhibit 50 and 90% of P. ultimum oospore germination were determined as 31.3 and 250 μg/ml, respectively. Drench treatment of cucumber seeds in P. ultimum‐infested potting medium (500 oospores/g) with 50 μg/ml SRE or water with no SRE resulted in 90.2 ± 4.5% and 65.7 ± 4.6% germination rates, respectively. Seed coating with 0.03% (w/w) SRE allowed 65.7 ± 4.6% seedlings to germinate on naturally infested soil while 100.0 ± 0.0% of non‐coated seeds were unable to germinate due to Pythium infection. Organic‐compatible and scalably produced SRE is potentially a novel organic fungicide seed protectant.     

Inferring climatic controls of rice stem borers’ spatial distributions using maximum entropy modelling

Accurate assessment of pest potential distributions is needed to identify their establishment risks that play a key role in pest management in agricultural ecosystems. We used a correlative niche modelling method (Maxent) to predict and map the spatial distributions of two important rice stem borers, Chilo suppressalis and Sesamia cretica, in paddy fields of Iran. In total, 195 presence occurrence records (101 records for C. suppressalis and 94 records for S. cretica) were compiled. A set of environmental and topographic variables, with the highest effects on the species distributions and the lowest correlations among themselves, were used. The results showed that mainly the northern parts of Iran were the most suitable areas for C. suppressalis, and north, north‐east and south‐west of Iran as the most suitable areas for S. cretica. Both models performed well, with an area under the receiver operating characteristic curve (AUC) of 0.983 and 0.786 for C. suppressalis and S. cretica, respectively. The Maxent models showed higher accuracy for predicting the distribution of the specialist pest with the small range sizes compared to the generalist species. Assessing the importance of environmental variables, which were derived from the jackknife test, showed the precipitation as the variable with the highest contribution (66%) in explaining the spatial distribution of C. suppressalis compared to the other variables. The distribution of S. cretica was influenced by a set of variables derived from both the precipitation and temperature. The Maxent predictions were useful to map the geographical distributions of the risk for both rice stem borers that is needed to develop effective management strategies.     

Comparative Molecular Analyses of Phytoplasmas infecting Sophora japonica cv. golden and Robinia pseudoacacia

Phytoplasmas were detected in Sophora japonica cv. golden and Robinia pseudoacacia with diseased branches of witches'‐broom collected in Haidian district, Beijing, China. Phytoplasma cells were observed in phloem sieve elements of symptomatic S. japonica cv. golden by transmission electron microscopy. The presence of phytoplasmas was further confirmed by sequence determination of partial gene sequences of 16S rDNA, rp (ribosomal protein) and secY. Phylogenetic trees and virtual restriction fragment length polymorphism (RFLP) analyses indicated that the phytoplasmas causing S. japonica cv. golden witches'‐broom (SJGWB) and R. pseudoacacia witches'‐broom (RPWB) belong to the 16SrV (elm yellows) group, and they are most closely related to subgroup 16SrV‐B, rpV‐C and secYV‐C jujube witches'‐broom (JWB) phytoplasma. Comparative analyses indicated that the phytoplasma of RPWB was closer to the JWB and that R. pseudoacacia might serve as an alternative host plant of JWB phytoplasma.     

Defence‐Related Enzymes in Soybean Resistance to Target Spot

Target spot, caused by the fungus Corynespora cassiicola, has become a serious foliar disease in soybean production in the Brazilian Cerrado. Information in the literature regarding the biochemical defence responses of soybean to C. cassiicola infection is rare. Therefore, the objective of this study was to determine the biochemical features associated with soybean resistance to target spot. The activities of chitinases (CHI), β‐1‐3‐glucanases (GLU), phenylalanine ammonia‐lyases (PAL), peroxidases (POX), polyphenol oxidases (PPO) and lipoxygenases (LOX), as well as the concentrations of total soluble phenolics (TSP) and lignin‐thioglycolic acid (LTGA) derivatives, were determined in soybean leaves from both a resistant (FUNDACEP 59) and a susceptible (TMG 132) cultivar. The target spot severity, number of lesions per cm² of leaflet and area under the disease progress curve were significantly lower for plants from cv. FUNDACEP 59 compared to plants from cv. TMG 132. The GLU, CHI, PAL, POX and PPO activities and the concentration of LTGA derivatives increased significantly, whereas LOX activity decreased significantly on the leaves infected by C. cassiicola. Inoculated plants from cv. FUNDACEP 59 showed a higher PPO activity and concentrations of TSP and LTGA derivatives at 4 and 6 days after inoculation compared to plants from cv. TMG 132. In conclusion, the results of this study demonstrated that the defence‐related enzyme activities increased upon C. cassiicola infection, regardless of the basal level of resistance of the cultivar studied. The increases in PPO activity and concentrations of TSP and LTGA derivatives, but lower LOX activity, at early stages of C. cassiicola infection were highly associated with soybean resistance to target spot.     

Susceptibility of six cultivars of bean to different isolates of two species of Fusariumin greenhouse

In greenhouse, susceptibility of 6 cultivars of bean (b1, green bean cv. contender, b2, Pinto b. cv. Cos-16, b3, green b. Sun Ray, b4, white b. cv. Daneshkadeh, b5, red b. cv. Naz, b6, red b. cv. Goli) evaluated to 7 isolates of 2 species of Fusarium (6 isolates of F. oxysporum and 1 isolate of F. solani from Varamin, Iran) in 4 pots (as replications) in factoriel experiment. The seedlings of each bean cultivars soaked in a suspension of the fungi (106 spores/ml.) for 5 minutes and transplanted into the pots contained sterilized soil, for control, the roots soaked in distilled water. After 35 days of inoculating and transplanting in greenhouse conditions (with 18-26 degrees C and 12 h light & 12 h dark) the percentage of necrotic roots and stems of the seedlings affected to the isolates of the fungi, also height and weight of them in different treatments datermined and calculated. The results showed that, the average of necrotic percent of the cultivars of bean seedlings grouped in 3 category (b1, b2, b3, b4 with 25.15% in group a and b5, red bean cv. Naz with 16.29% in group b and b6 red bean cv. Goli with 10.02% in group c) based on cluster analysis. But by Dunken method the infection set on 5 groups as respectively b4, 28,72% (a), b2, 25.83% (ab), b1 & b3, 23,18% & 22.84% (b), b5, 16.29% (c) and b6, 10.02% (d). So, red bean cv. Goli was more resistant and white bean cv. Daneshkadeh was more susceptible than of another cultivars, but averse the effect of the fungi on growth factors were different.     

Rapid Diagnosis of Soya Bean Root Rot Caused by Fusarium culmorum Using a Loop‐Mediated Isothermal Amplification Assay

We report a rapid diagnosis of soya bean (Glycine max L.) root rot caused by Fusarium culmorum, using a loop‐mediated isothermal amplification (LAMP) assay. We used the CYP51C gene sequence to design LAMP assay primers specific for F. culmorum. The LAMP assay amplified the target gene efficiently in 60 min at 63°C. The sensitivity of the assay was 100 pg/μl of genomic DNA. Among the tested soya bean pathogens, a positive colour (sky blue) was only observed in the presence of F. culmorum with the addition of hydroxynaphthol blue (HNB) dye prior to amplification, whereas other species isolates showed no colour change. Suspected diseased soya bean samples collected in the field from Jiangsu, Shandong and Anhui provinces and Beijing were diagnosed successfully using the LAMP assay reported here. This study provides a new and readily available method for rapid diagnosis of soya bean root rot caused by F. culmorum.     

Evaluating the Qualitative Effectiveness of a Novel Pollinator: a Case Study of Two Endemic Hawaiian Plants

In situations where native mutualists have become extinct, non‐native species may partner with remnant native species. However, non‐native mutualists may differ behaviorally from extinct native mutualists. In the case of pollination, novel relationships between natives and non‐natives could differ both quantitatively and qualitatively from native–native relationships. In Hawai'i, the non‐native Japanese White‐eye (Zosterops japonicus) has largely replaced endemic birds as pollinator of the endemic Clermontia parviflora and C. montis‐loa. We surveyed Clermontia patches and found that they ranged from 106 to 1198 m in diameter. We performed manual pollination of flowers with pollen taken from plants at five distance categories, ranging from 0 (self‐fertilization) to 20 km, and examined the germination of resulting seeds. We used radiotelemetry to estimate daily Japanese White‐eye movement distances. Percent germination of seeds after short‐ to intermediate‐distance pollination crosses (i.e., 20–1200 m, or intra‐patch pollen transfer distances) significantly exceeded germination of seeds from selfed trials for C. parviflora. No significant differences in germination rates among treatments were detected for C. montis‐loa. The maximum daily movement distances of radio‐tracked birds were generally <1 km. Together, these results suggest that this novel pollinator may be an effective mutualist for both Clermontia species. This study serves as an example of research examining qualitative components of novel mutualism, which are generally neglected relative to quantitative components.