Plant Growth‐Promoting Bacteria from Solarized Soil with the Ability to Protect Melon Against Root Rot and Vine Decline Caused by Monosporascus cannonballus

This study was undertaken to isolate indigenous plant growth‐promoting (PGP) bacteria from solarized soil effective in the biocontrol of Monosporascus cannonballus, the cause of root rot and vine decline of melon, which is one of the most destructive soilborne diseases of this crop worldwide. The screening strategy resulted in the selection of two interesting PGP bacteria as biocontrol candidates against M. cannonballus belonging to the same microbial community. The two bacterial species, identified according to phenotypic, physiological tests and analysis of the 16S rDNA sequence as Bacillus subtilis/amyloliquefaciens (BsCR) and Pseudomonas putida (PpF4), showed PGP traits and in vitro antagonistic activity towards M. cannonballus. Antagonism by BsCR was characterized by a consistent inhibition of the pathogen in vitro growth; PpF4 strongly inhibited the development of perithecia of the pathogen. Under greenhouse conditions, the selected bacteria were tested for their biocontrol activity in the pathosystem melon‐M. cannonballus. BsCR alone and in combination with PpF4 determined a consistent decrease in the disease symptoms. BsCR and the combination of the bacterial strains significantly increased root biomass in both inoculated and un‐inoculated plant. Upon seed treatment with BsCR, the accumulation and isoenzyme induction of peroxidase in roots as biochemical marker for induction of resistance were found, thus indicating that BsCR may reduce the disease severity also by the activation of the plant defence responses. The study highlights the synergistic biocontrol potential of B. subtilis BsCR and P. putida PpF4 in the integrated management of root rot and vine decline of melon caused by M. cannonballus.     

Role of Pseudomonas fluorescens and INA against Black Rot of Cabbage

Cabbage (Brassica oleracea var. capitata) is an important vegetable crop among crucifers. It is affected by a bacterial disease known as black rot. Black rot is caused by Xanthomonas campestris pv. campestris a disease of worldwide importance. The present study highlights the effect of biotic inducer—Pseudomonas fluorescens—and an abiotic inducer—2,6‐dichloro‐isonicotinic acid—in combating black rot, followed by their effect on the seed treatment and disease incidence, role of antioxidant enzymes followed by validation of the defence‐related genes by quantitative real‐time PCR. The resistant (Pusa mukta) and the highly susceptible (NBH boss) cabbage cultivars were analysed for defence‐related enzymes such as peroxidase and superoxide dismutase. An increase in total peroxidase and superoxide dismutase activity was observed upon inoculation with X. campestris pv. campestris. The activity was greater in resistant cultivar when compared to susceptible ones. Both enzyme activity assays and qPCR analyses for the expression of the defence genes in susceptible and resistant cultivars demonstrated that the peroxidase gene was up‐regulated in resistant cultivar compared to susceptible cultivar. The present study proved that P. fluorescens‐induced resistance against X. campestris pv. campestris in cabbage seedlings is more efficient as compared to the use of INA—abiotic inducer.     

Differences in the Induction of Defence Responses in Resistant and Susceptible Muskmelon Plants Infected with Colletotrichum lagenarium

Defence reactions occurring in resistant (cv. Gankezaomi) and susceptible (cv. Ganmibao) muskmelon leaves were investigated after inoculating with Colletotrichum lagenarium. Lesion restriction in resistant cultivars was associated with the accumulation of hydrogen peroxide (H₂O₂). The activity of antioxidants catalase (CAT) and peroxidase (POD) significantly increased in both cultivars after inoculation, while levels of both CAT and POD activity were significantly higher in the resistant cultivar. Ascorbate peroxidase (APX) increased in both cultivars after inoculation, and level of APX activity was significantly higher in the resistant cultivar. Glutathione reductase (GR) activity significantly increased in both cultivars following inoculation, but was higher in the resistant cultivar, resulting in higher levels of ascorbic acid (AsA) and reduced glutathione (GSH). Phenylalanine ammonia lyase (PAL) significantly increased in inoculated leaves of both cultivars, resulting in higher levels of total phenolic compounds and flavonoids. The pathogenesis‐related proteins chitinase (CHT) and β‐1, 3‐glucanase (GLU) significantly increased following inoculation with higher activity in the resistant cultivar. These findings show that resistance of muskmelon plants against C. lagenarium is associated with the rapid accumulation of H₂O₂, resulting in altered cellular redox status, accumulation of pathogenesis‐related proteins, activation of phenylpropanoid pathway to accumulation of phenolic compounds and flavonoids.     

Assessing cultivar resistance to Sitodiplosis mosellana (Géhin) (Diptera: Cecidomyiidae) using a phenotyping method under semi‐field conditions

The orange wheat blossom midge, Sitodiplosis mosellana (Géhin), can significantly reduce wheat yield. Growing resistant wheat cultivars is an effective way of managing this pest. The assessment of cultivar resistance in field trials is difficult because of unequal pressure of S. mosellana caused by differences in cultivar heading dates relative to the flight period of S. mosellana adult females and huge variations of egg laying conditions from 1 day to another. To overcome these hurdles and to expose all cultivars homogeneously to the pest, an assessment method of cultivar resistance was developed under semi‐field conditions. In 2015, the resistance of 64 winter wheat cultivars to S. mosellana was assessed. Few or no larvae developed in the ears of resistant cultivars, but in susceptible cultivars, large numbers of larvae developed. Seventeen cultivars proved to be resistant, whereas 47 were susceptible. The identification of new resistant cultivars offers more opportunities to manage S. mosellana. The phenotyping method is easy, cheap, efficient and reliable. It can be used to guide the breeding of new resistant wheat cultivars. Using specific midge populations, this method could also be used in research on new resistance mechanisms in winter wheat or in other cereal species.     

Reactive Oxygen Species are not Increased in Resistant Oat Genotypes Challenged by Crown Rust Isolates

Crown rust (Puccinia coronata Corda f.sp. avenae) can devastate oats (Avena sativa). Oxidative stress is part of the resistance mechanism in several pathosystems, but in the oat–crown rust system, it is unclear, especially regarding partial resistance. We evaluated the effects of P. coronata on oxidative stress in oat cultivars: URS 21 (partially resistant), Leggett (race‐specific resistant), URS22 and Clintland 64 (susceptibles). Seedlings and plants were inoculated with P. coronata uredospores. Cultivars were assessed for antioxidant enzyme activity and the reactive oxygen species (ROS) hydrogen peroxide and superoxide. Due to the importance of the partial resistance of URS21, this cultivar and URS 22 were also appraised for total phenolics and the relative expression of oxidative stress genes. Postinoculation, Leggett and URS 21 showed no increased peroxide levels. The susceptible cultivars increased ROS and ascorbate peroxidase activity. Clintland 64 increased also catalase activity, whereas URS 22 increased glutathione reductase and the expression of genes encoding antioxidant enzymes. URS 21 showed almost no antioxidant enzyme induction. Shortly after inoculation, URS 21 showed increased expression of genes encoding lipoxygenase and peroxidase. Cultivars URS 21 and Leggett accumulated cell wall fluorescent compounds, phenolics being detected in the former. Oxidative stress appears not to cause the hypersensitive response in this pathosystem, but late ROS accumulation did occur in the susceptible cultivars. Cultivar URS 21 may, differently from other known mechanism to date, reduce ROS accumulation by increasing the level of phenolics, resulting in later pathogen and cell death, showing non‐specific resistance to races of the pathogen also at seedling stage.     

Cotton,cowpea and sesame are alternative crops to cucurbits in soils naturally infested with Monosporascus cannonballus

Monosporascus cannonballus is an important cucurbit root pathogen, which has been reported in the main production areas of melon and watermelon in Brazil and worldwide and potentially capable to colonize roots of different species. Crop rotation is considered an effective management strategy to prevent this disease. The aim of this study was to evaluate the response of different crops, pumpkin, cotton, cowpea, sesame, watermelon, melon, corn, cucumber, sorghum and tomato, to the infection of this pathogen. Seedlings were transplanted into plastic containers with an inoculum concentration of 20 colony‐forming units (CFU) g^?1 of M. cannonballus. Fifty days after transplanting, the variables analysed were the degree of disease severity on the root system and the frequency of reisolation. On cucurbits, the results demonstrated different degrees of susceptibility among crops and cultivars, being melon and watermelon the most sensitive species. In contrast, Cucurbita cultivars were the most tolerant. Regarding non‐cucurbit crops, maize, sorghum and tomato presented root discoloration and M. cannonballus was reisolated from roots. Cotton, cowpea and sesame cultivars were not affected by the pathogen, so they can be considered as alternative crops to be cultivated, or in rotation with cucurbits, in M. cannonballus infested soils.     

Cytology of infection,development and expression of resistance to Plasmodiophora brassicae in canola

The timing and expression of resistance to four isolates of Plasmodiophora brassicae, collected from research sites where pathotypes 2, 3, 5 and 6 (Williams' system) had been dominant when characterised in 2006, were assessed in four new commercial cultivars of canola (Brassica napus) with resistance to clubroot. Each of the resistant cultivars was highly resistant to all four of the isolates, and there was no difference in their response to infection. Root hair infection occurred at high levels, but pathogen development occurred more slowly than in a susceptible cultivar (control). Secondary infection and development in cortical cells was severely inhibited in each of the resistant cultivars; only a few bi‐nucleated plasmodia were observed at 12 days after inoculation (DAI), and plasmodia were rarely observed at 18 and 24 DAI. In contrast, development in the susceptible cultivar had progressed to resting spores by 24 DAI. A dense ring of accumulated reactive oxygen species (ROS) was observed in the endodermis, pericycle and vascular cambium of non‐inoculated controls and inoculated plants of the resistant cultivars. However, the ROS ring disappeared rapidly in infected plants of the susceptible cultivar. Plasmodia invaded the stele of susceptible roots by preferentially colonising the xylem parenchyma cells. Expansion and enlargement of lignified xylem cells was observed by 35 DAI. The absence of any specific points of ROS accumulation or lignification of epidermal or cortical cells in the resistant cultivars indicates that a hypersensitive response is not the main mechanism of resistance in these lines. The uniform response of these resistant cultivars to the four isolates of P. brassicae indicates that the resistance in each cultivar may be conditioned by a gene(s) from a single source that confers broad resistance, because most sources of resistance to P. brassicae are pathotype specific.     

Effects of Preconditioning Through Mycorrhizal Inoculation on the Control of Melon Root Rot and Vine Decline Caused by Monosporascus cannonballus

The use of inoculum of arbuscular mycorrhizal fungi (AMF) in nursery represents a promising field in horticulture because of its known benefits in terms of plant growth and bioprotection. The present work was undertaken to determine the effect of mycorrhizal inoculation with Rhizophagus irregularis in a nursery medium on the containment of melon root rot and vine decline (MRRVD) caused by the soil‐borne pathogen Monosporascus cannonballus. The percentage of mycorrhization, biomass and yield following mycorrhizal inoculation were also evaluated. Biocontrol activity was assessed in greenhouse pot experiments upon artificial inoculation of M. cannonballus and in a two‐season field experiment under production conditions in an unheated greenhouse with a history of MRRVD. On the basis of the mycorrhization parameters, the interaction appeared to be established within 30 days after inoculation. The total shoot growth in the mycorrhized plants was significantly higher when compared to the control, while the root growth was unaffected. Upon artificial inoculation of M. cannonballus, mycorrhization provided complete protection against the pathogen. Greenhouse experiments under production conditions during spring cropping season showed that pretransplanting inoculation with R. irregularis significantly decreased the severity of the disease. Also, the average fruit weight of mycorrhized plants was significantly higher than the untreated control. Nevertheless, in summer crop, the bioprotection activity of AMF failed. Present results indicate that the use of AMF in a nursery setting can contribute to the prevention of the onset of this problematic soil‐borne disease within a sustainable and integrated soil‐borne disease management.     

Feeding behavior and performance of Nasonovia ribisnigri on grafts,detached leaves,and leaf disks of resistant and susceptible lettuce

Aphids are dependent on the phloem sap of plants as their only source of nutrients. Host‐plant resistance in lettuce, Lactuca sativa L. (Asteraceae), mediated by the Nr gene is used to control the lettuce aphid Nasonovia ribisnigri (Mosely) (Hemiptera: Aphididae). The resistance is located in the phloem; however, the exact mechanism of resistance is unknown. In this study, we investigated whether the resistance factor (or factors) is synthesized in the root or in the shoot. The feeding behavior and performance of avirulent N. ribisnigri were studied on grafts of resistant and susceptible lettuce. In addition, the persistence of resistance in excised lettuce tissue was measured, by studying the feeding behavior and performance of N. ribisnigri on detached leaves and leaf disks of resistant lettuce. It appears that the resistance factor encoded by the Nr gene is produced in the shoots: aphid feeding was reduced on resistant shoots grafted on susceptible roots, whereas aphids were able to feed on grafts of susceptible shoots on resistant roots. Partial loss of resistance was observed after detachment of leaves and excision of leaf disks from resistant plants. Aphids fed longer on excised resistant plant tissue compared with intact resistant plants; however, compared with excised plant tissue of the susceptible cultivar, the time spent on feeding was shorter, indicating resistance was not completely lost. Our findings caution against the use of excised leaf material for aphid resistance bioassays.     

Influence of Soil Temperature on Meloidogyne incognita Resistant and Susceptible Cotton,Gossypium hirsutum

The degree of resistance by a cotton plant to Meloidogyne incognita is affected by soil temperature, particularly in moderately resistant cultivars, The total number of nematodes in the resistant and moderately resistant rools at 35 C was equal to, or greater than, the number in susceptible roots at 20, 25, or 30 C. A shift in numbers to developing and egg-bearing forms of nematodes in the susceptible cultivar as tentperature increased indicates development was affected by temperature rather than by genetic resistance mechanisms. However, the nematode resistant cultivar did not support maturation of nematodes until a soil tempurature of 35 C was attained. This indicated that resistance mechanisms are partially repressed at 35 C and differences in nematode development cannot be explained in terms of accumulated heat units. The moderately resistant cultivar was significantly more sensitive to the effects of high temperature than was the resistant cultivar.     

Screen of safflower (Carthamus tinctorius L.) genotypes against Phytophthora drechsleri and Fusarium solani,the causal agents of root rot disease

Safflower (Carthamus tinctorius L.) plants were affected by a severe root rot disease caused by Phytophthora drechsleri and Fusarium solani in Isfahan province of Iran during 2005–2007. Disease incidence was more than 30% in severely infected fields. Twenty-one safflower genotypes, including six local cultivars and 15 internal pure lines were evaluated for their resistance to root rot disease in laboratory and greenhouse conditions. Safflower seedlings were evaluated for lesion length on infected roots in laboratory, as well percentage of live seedlings in greenhouse. The results indicated a high negative correlation between lesion length on roots and percentage of live seedlings. The most resistant and susceptible genotypes to P. drechsleri were identified as pure line Karaj row 12 (KW12) and cultivar Koseh with lesion lengths of 10.01 and 15.51?mm on roots and 45.60 and 18.00% live seedlings, respectively. The most resistant genotype to F. solani was identified as pure line KW11 with a lesion length of 9.31?mm on roots as well 62.80% live seedlings. The most susceptible genotypes were identified as cultivar Koseh and pure lines KW2 and KW3 with lesion lengths of 13.29, 12.72 and 12.13?mm on roots and 25.60, 28.40 and 28.40% live seedlings, respectively. The most resistant genotypes to both P. drechsleri and F. solani were identified as pure lines KW15 and KW9 with a 55.40% live seedlings. The most susceptible genotypes were cultivars Koseh, Goldasht and pure lines KW6, KW3 and KW2 with 35.40, 35.40, 35.40, 37.60 and 37.60% live seedlings in greenhouse, respectively.     

Components of Resistance to Early Blight in Four Potato Cultivars: Effect of Leaf Position

Components of early blight resistance were quantified in leaves of different ages in four potato cultivars. The components of resistance: incubation period (IP), lesion number (LN), early blight severity, lesion expansion rate (LER), latent period (LP) and spore production by lesion area (SPLA), were evaluated separately in the lower, middle and upper leaves of four potato cultivars. Plants of cultivar Aracy (resistant), Delta (moderately resistant), Desirée (susceptible) and Bintje (susceptible) were inoculated with an Alternaria solani isolate at the beginning of the flowering stage. Disease severity varied in different plant parts. In all cultivars, regardless of resistance, the smallest values of LN, and severity were recorded on the upper leaves, suggesting that young tissues are less susceptible. In cultivar Aracy, the IP was long, with small values of LN and LER and consequently, low values of early blight severity in all leaf positions were recorded. Although IP was long in cultivar Aracy, no differences between the moderately resistant cultivar Delta and the susceptible cultivars Bintje and Desirée could be detected for this component. The IP was only influenced by leaf position in cultivar Aracy. Clear differences in resistance levels among cultivars could be detected regarding LN, severity and LER. However, neither LP nor SPLA were associated with resistance level of cultivars or with leaf position. Analyses according to plant part suggest that evaluations on leaves of the middle third part are most suitable for screening for early blight resistance in potato.     

Biochemical responses associated with resistance to bacterial stalk rot caused by Dickeya zeae in maize

Bacterial stalk rot (BSR) of maize caused by Dickeya zeae is an important disease in northwest region of India. In the current study, eighty maize lines were evaluated for resistance against this disease. Of these, 20 were moderately resistant, 25 were moderately susceptible and the rest were highly susceptible to BSR. Six lines from each set were randomly selected. Activities of three antioxidant enzymes, viz. phenylalanine ammonia lyase (PAL), peroxidase (POX) and polyphenol oxidase (PPO) were analysed from these three sets of maize lines representing different levels of resistance. A trend of elevated activity of PAL, POX and PPO was observed in all the three sets of maize lines. The results showed significantly more activity of these three enzymes in moderately resistant than highly susceptible maize lines. The activity of PAL and PPO peaked after 48 hr and of POX after 72 hr of challenge inoculation by D. zeae in all the maize lines. The activity of these enzymes further correlated negatively with disease development. Our results show that PAL, POX and PPO play an important role in contributing towards resistance in maize against BSR.     

Biological Relationship of Meloidogyne hapla Populations to Alfalfa Cultivars

Greenhouse and growth chamber studies were established to determine if there are pathological and physiological differences among Meloidogyne hapla populations from California (CA), Nevada (NV), Utah (UT), and Wyoming (WY) on alfalfa cultivars classified as resistant or susceptible to root-knot nematodes. In the greenhouse, plant survival was not consistent with resistance classifications. While all highly resistant Nevada Synthetic germplasm (Nev Syn XX) plants survived inoculation with all nematode populations, two cultivars classified as moderately resistant (''Chief'' and ''Kingstar'') survived (P ≤ 0.05) inoculation with M. hapla populations better than did ''Lobo'' cultivar, which is classified as resistant. Plant growth of Nev Syn XX was suppressed by only the CA population, whereas growth of the other alfalfa cultivars classified as M. hapla resistant or moderately resistant was suppressed by all nematode populations. Excluding Nev Syn XX, all alfalfa cultivars were severely galled and susceptible to all nematode populations. Except for Nev Syn XX, reproduction did not differ among the nematode populations on alfalfa cultivars. Nev Syn XX was not as favorable a host to CA as were the other cultivars; but, it was a good host (reproductive factor [Rf] = 37). Temperature affected plant resistance; the UT and WY populations were more pathogenic at 15-25 C, and CA was more pathogenic at 30 C. Nev Syn XX was susceptible to all nematode populations, except for CA, at only 30 C, and all other alfalfa cultivars were susceptible to all nematode populations at all temperatures.     

Feeding‐induced changes in defence enzymes and PR proteins and their implications in host resistance to Nilaparvata lugens

Six rice genotypes showing susceptible and resistant reactions to brown planthopper (BPH), Nilaparvata lugens were studied for feeding‐induced changes in defence enzymes and pathogenesis‐related (PR) proteins. The high resistant genotypes PTB 33, ADT 45 and ASD 7 and moderately resistant genotypes CO 43 and KAU 1661 recorded the greater expression of defence enzymes peroxidase, polyphenol oxidase, phenylalanine ammonia lyase, total phenol and β‐1,3 glucanase in response to N. lugens feeding at 1 day after infestation (DAI) compared with susceptible genotype TN1. The greater activity of chitinase was observed in resistant cultivars at 3 DAI and the activity was sustained for more than 1 week compared with susceptible TN1. In conclusion, the current study revealed that these defence enzymes and PR proteins might attribute to the resistance mechanisms in rice plants against BPH infestation.     

Changes in Peroxidase Activity during Potato Ring Rot Infection

The effects of the ring rot causal agent Clavibacter michiganensis subsp. sepedonicus (a virulent strain 5369) on the peroxidase activity of various tissues of potato plants grown under axenic conditions were studied. Root infection enhanced peroxidase activity in all plant tissues (roots, leaves, and stems). In the resistant cv. Lugovskoi, peroxidase activity was much higher than in the susceptible cv. Luk'yanovskii. Co-culturing of the suspension cells of these potato cultivars with the bacterial pathogen also activated peroxidase in the cells of the resistant cultivar; in the cells of the susceptible cultivar, peroxidase activation was less pronounced. Treating suspension cell with exopolysaccharides secreted by the pathogen enhanced the activity of extra- and intracellular peroxidases, and the degree of this enhancement differed in the two potato cultivars.     

The Role of Peroxidase and Polyphenol Oxidase Isozymes in Wheat Resistance to Alternaria triticina

Polyphenol oxidase activity was higher in resistant wheat cultivar ACC-8226 than in susceptible cultivar MP-845 in control sets and after inoculation of Alternaria triticina. However, similar polyphenol oxidase isozyme pattern was found in control and inoculated sets of both the cultivars, but the band intensity was higher after inoculation. Three and four peroxidase isozymes were found in ACC-8226 and MP-845, respectively. An extra peroxidase isozyme band was observed in both the cultivars after inoculation. The results suggest an active role of peroxidase and polyphenol oxidase in defence mechanism of wheat plants.     

The effect of leaf exudates on the spore germination of phylloplane mycoflora of chilli (Capsicum annuum L.) cultivars

Summary Ninetten aminoacids, twelve sugars, eleven organic acids and ten phenols were detected in the leaf exudates of three cultivars of chilli. The number of aminoacids, sugars, organic acids and phenols increased as the plants grew older. More aminoacids and sugars were detected in the exudate from the susceptible cultivar (Malwa). More organic acids and phenols were detected from the resistant cultivar (Simla). The leaf exudate of the resistant cultivar (Simla) inhibited spore germination of the pathogen (Alternaria solani) while that of susceptible (Malwa) stimulated spore germination. The cultivar ‘Patna’ which is moderately resistant, occupied an intermediate position. Spore germination of the isolated fungi was enhanced in leaf exudate of susceptible cultivar (Malwa), while leaf exudates of the moderately resistant (Patna) and resistant (Simla) inhibited spore germination of the majority of fungi isolated. Most of the antagonistic fungi were not isolated from the susceptible cultivar and the percentage spore germination of these fungi was less in leaf exudate of the susceptible cultivar, while leaf exudates of resistant cultivars enhanced the percentage spore germination of antagonistic fungi,viz Aspergillus flavus, A. fumigatus, A. versicolor, Penicillium citrinum, P. restrictum andTrichoderma viride.     

Comparative proteomic analysis of methyl jasmonate‐induced defense responses in different rice cultivars

Jasmonate is an important endogenous chemical signal that plays a role in modulation of plant defense responses. To understand its mechanisms in regulation of rice resistance against the fungal pathogen Magnaporthe oryzae, comparative phenotype and proteomic analyses were undertaken using two near‐isogenic cultivars with different levels of disease resistance. Methyl‐jasmonate (MeJA) treatment significantly enhanced the resistance against M. oryzae in both cultivars but the treated resistant cultivar maintained a higher level of resistance than the same treated susceptible cultivars. Proteomic analysis revealed 26 and 16 MeJA‐modulated proteins in resistant and susceptible cultivars, respectively, and both cultivars shared a common set of 13 proteins. Cumulatively, a total of 29 unique MeJA‐influenced proteins were identified with many of them known to be associated with plant defense response and ROS accumulation. Consistent with the findings of proteomic analysis, MeJA treatment increased ROS accumulation in both cultivars with the resistant cultivar showing higher levels of ROS production and cell membrane damage than the susceptible cultivar. Taken together, our data add a new insight into the mechanisms of overall MeJA‐induced rice defense response and provide a molecular basis of using MeJA to enhance fungal disease resistance in resistant and susceptible rice cultivars.