Effects of the Mi-1, N and Tabasco Genes on Infection and Reproduction of Meloidogyne mayaguensis on Tomato and Pepper Genotypes

Meloidogyne mayaguensis is a damaging root-knot nematode able to reproduce on root-knot nematode-resistant tomato and other economically important crops. In a growth chamber experiment conducted at 22 and 33°C, isolate 1 of M. mayaguensis reproduced at both temperatures on the Mi-1-carrying tomato lines BHN 543 and BHN 585, whereas M. incognita race 4 failed to reproduce at 22°C, but reproduced well at 33°C. These results were confirmed in another experiment at 26 ± 1.8°C, where minimal or no reproduction of M. incognita race 4 was observed on the Mi-1-carrying tomato genotypes BHN 543, BHN 585, BHN 586 and ‘Sanibel’, whereas heavy infection and reproduction of M. mayaguensis isolate 1 occurred on these four genotypes. Seven additional Florida M. mayaguensis isolates also reproduced on resistant ‘Sanibel’ tomato at 26 ± 1.8°C. Isolate 3 was the most virulent, with reproduction factor (Rf) equal to 8.4, and isolate 8 was the least virulent (Rf = 2.1). At 24°C, isolate 1 of M. mayaguensis also reproduced well (Rf ≥ 1) and induced numerous small galls and large egg masses on the roots of root-knot nematode-resistant bell pepper ‘Charleston Belle’ carrying the N gene and on three root-knot nematode-resistant sweet pepper lines (9913/2, SAIS 97.9001 and SAIS 97.9008) carrying the Tabasco gene. In contrast, M. incognita race 4 failed to reproduce or reproduced poorly on these resistant pepper genotypes. The ability of M. mayaguensis isolates to overcome the resistance of tomato and pepper genotypes carrying the Mi-1, N and Tabasco genes limits the use of resistant cultivars to manage this nematode species in infested tomato and pepper fields in Florida.     

Histology of the Interactions of Paecilomyces lilacinus with Meloidogyne incognita on Tomato

Excised tomato roots were examined histologically for interactions of the fungus Paecilomyces lilacinus and Meloidogyne incognita race 1. Root galling and giant-cell formation were absent in tomato roots inoculated with nematode eggs infected with P. lilacinus. Few to no galls and no giant-cell formation were found in roots dipped in a spore suspension of P. lilacinus and inoculated with M. incognita. Numerous large galls and giant cells were present in roots inoculated only with M. incognita. P. lilacinus colonized the surface of epidermal cells as well as the internal cells of epidermis and cortex. The possibility of biological protection of plant surfaces with P. lilacinus against root-knot nematodes is discussed.     

Sensitivity of Meloidogyne incognita and Rotylenchulus reniformis to Fluopyram

Fluopyram is a succinate dehydrogenase inhibitor (SDHI) fungicide that is being evaluated as a seed treatment and in-furrow spray at planting on row crops for management of fungal diseases and its effect on plant-parasitic nematodes. Currently, there are no data on nematode toxicity, nematode recovery, or effects on nematode infection for Meloidogyne incognita or Rotylenchulus reniformis after exposure to low concentrations of fluopyram. Nematode toxicity and recovery experiments were conducted in aqueous solutions of fluopyram, while root infection assays were conducted on tomato. Nematode paralysis was observed after 2 hr of exposure at 1.0 µg/ml fluopyram for both nematode species. Using an assay of nematode motility, 2-hr EC₅₀ values of 5.18 and 12.99 µg/ml fluopyram were calculated for M. incognita and R. reniformis, respectively. Nematode recovery in motility was greater than 50% for M. incognita and R. reniformis 24 hr after nematodes were rinsed and removed from a 1-hr treatment of 5.18 and 12.99 µg/ml fluopyram, respectively. Nematode infection of tomato roots was reduced and inversely proportional to 1-hr treatments with water solutions of fluopyram at low concentrations, which ranged from 1.3 to 5.2 µg/ml for M. incognita and 3.3 to 13.0 µg/ml for R. reniformis. Though fluopyram is nematistatic, low concentrations of the fungicide were effective at reducing the ability of both nematode species to infect tomato roots.     

Reproduction of Mi-Virulent Meloidogyne incognita Isolates on Lycopersicon spp.

Selection of detectable numbers of Mi-virulent root-knot nematodes has necessitated a greater understanding of nematode responses to new sources of resistance. During the course of this research, we compared the reproduction of four geographically distinct Mi-virulent root-knot nematode isolates on three resistant accessions of Lycopersicon peruvianum. Each accession carried a different resistant gene, Mi-3, Mi-7, or Mi-8. All nematode isolates were verified as Meloidogyne incognita using diagnostic markers in the mitochondrial genome of the nematode. Reproduction of Mi-virulent isolates W1, 133 and HM, measured as eggs per g of root, was greatest on the Mi-7 carrying accession and least on the Mi-8 carrying accession. In general, Mi-3 behaved similar to the Mi-8 carrying accession. Reproduction of the four nematode isolates was also compared on both Mi and non-Mi-carrying L. esculentum cultivars and a susceptible L. peruvianum accession. Resistance mediated by Mi in L. esculentum still impacted the Mi-virulent nematodes with fewer eggs per g of root on the resistant cultivar (P ≤ 0.05). Preliminary histological studies suggests that Mi-8 resistance is mediated by a hypersensitive response, similar to Mi.     

Pasteuria penetrans for Control of Meloidogyne incognita on Tomato and Cucumber,and M. arenaria on Snapdragon

Meloidogyne incognita and Meloidogyne arenaria are important parasitic nematodes of vegetable and ornamental crops. Microplot and greenhouse experiments were conducted to test commercial formulations of the biocontrol agent Pasteuria penetrans for control of M. incognita on tomato and cucumber and M. arenaria on snapdragon. Three methods of application for P. penetrans were assessed including seed, transplant, and post-plant treatments. Efficacy in controlling galling and reproduction of the two root-knot nematode species was evaluated. Seed treatment application was assessed only for M. incognita on cucumber. Pasteuria treatment rates of a granular transplant formulation ranged from 1.5 × 10⁵ endospores/cm³ to 3 × 10⁵ endospores/cm³ of transplant mix applied at seeding. Additional applications of 1.5 × 10⁵ endospores/cm³ of soil were applied as a liquid formulation to soil post-transplant for both greenhouse and microplot trials. In greenhouse cucumber trials, all Pasteuria treatments were equivalent to steamed soil for reducing M. incognita populations in roots and soil, and reducing nematode reproduction and galling. In cucumber microplot trials there were no differences among treatments for M. incognita populations in roots or soil, eggs/g root, or root condition ratings. Nematode reproduction on cucumber was low with Telone II and with the seed treatment plus post-plant application of Pasteuria, which had the lowest nematode reproduction. However, galling for all Pasteuria treatments was higher than galling with Telone II. Root-knot nematode control with Pasteuria in greenhouse and microplot trials varied on tomato and snapdragon. Positive results were achieved for control of M. incognita with the seed treatment application on cucumber.     

Relationships Between Tolerance and Resistance to Meloidogyne incognita in Cotton

The southern root-knot nematode, Meloidogyne incognita, is the most damaging pathogen of cotton in the United States, and both resistance and tolerance to M. incognita could be valuable management approaches. Our objectives were to evaluate advanced cotton breeding lines for resistance and tolerance to M. incognita and to determine if a relationship between resistance and tolerance exists. Reproduction of M. incognita was evaluated on 17 breeding lines, a susceptible control (Delta and Pine Land DP5415), and a resistant control (M-120) in two greenhouse trials with six replications in a randomized complete block design. Two-week-old seedlings were inoculated with 8,000 M. incognita eggs and assessed for egg production 8 weeks later. Reproduction on the resistant control was only 10% of that on the susceptible control. Eight breeding lines supported 45% to 57% less (P <= 0.05) nematode reproduction than the susceptible control, and none of them were as resistant as M-120. Yield was determined in 2001 and 2002 in fumigated (1,3-dichloropropene at 56 liters/ha) and nonfumigated plots in a strip-plot design with three replications in a field naturally infested with M. incognita. Yield suppression caused by nematode infection differed among genotypes (P ≤ 0.05 for genotype × fumigation interaction). Six genotypes in 2001 and nine in 2002 were tolerant to M. incognita based on no difference in yield between the fumigated and nonfumigated plots (P ≥ 0.10). However, only three genotypes had no significant yield suppression in both years, of which two also were resistant to M. incognita. Regression analysis indicated that yield suppression decreased linearly as nematode resistance increased.     

An In Vitro Test for Temperature Sensitivity and Resistance to Meloidogyne incognita in Tomato

An in vitro root explant tissue culture technique is described for determining susceptibility of tomato (Lycopersicon esculentum Mill.) breeding lines and cultivars to the root-knot nematode Meloidogyne incognita. Root explants were taken from 2-day-old seedlings cultured for 30 days at 28 C on Gamborg''s B-5 medium with or without nematode inoculum. The remaining portion of the root and stem from the excised root explants was transferred to soil in pots and grown to maturity in the greenhouse. In vitro root explants were evaluated for growth and occurrence of juveniles, adults, and egg masses. The regenerated plants were used to produce more seed, The proposed technique is simple, reliable, and adapted to routine screening of large numbers of F₁ and F₂ samples, and it utilizes less space than tests performed on intact plants in the greenhouse or growth chamber. Evidence is presented also on the breakdown of resistance to M. incognita under high temperature stress using this in vitro root explant technique.     

Effects of Temperature on Resistance in Phaseolus vulgaris Genotypes and on Development of Meloidogyne Species

Phaseolus vulgaris lines with heat-stable resistance to Meloidogyne spp. may be needed to manage root-knot nematodes in tropical regions. Resistance expression before and during the process of nematode penetration and development in resistant genotypes were studied at pre- and postinoculation temperatures of 24 °C and 24 °C, 24 °C and 28 °C, 28 °C and 24 °C, and 28 °C and 28 °C. Resistance was effective at all temperature regimes examined, with fewer nematodes in roots of a resistant line compared with a susceptible line. Preinoculation temperature did not modify resistance expression to later infections by root-knot nematodes. However, postinoculation temperatures affected development of Meloidogyne spp. in both the resistant and susceptible bean lines tested. The more rapid development of nematodes to adults at the higher postinoculation temperature of 28 °C in both bean lines suggests direct temperature effects on nematode development instead of on resistance expression of either of two gene systems. Also, resistance was stable at 30 °C and 32 °C.     

Evaluation of Dry Ice as a Potential Cryonematicide for Meloidogyne incognita in Soil

Solid CO₂ (dry ice) was added to pots containing soil that was infested either with eggs of the root-knot nematode, Meloidogyne incognita, or with tomato (Lycopersicon esculentum ''Rutgers'') root fragments that were infected with various stages of the nematode. Two hours after dry ice was added, thermocouples in the soil recorded temperatures ranging from -15 °C to -59 °C. One day after treatment with the dry ice, the temperature of the soil was allowed to equilibrate with that of the greenhouse, and susceptible tomato seedlings were planted in pots containing infested soil treated or untreated (controls) with dry ice. After 5 weeks, roots were removed from the pots and nematode eggs were extracted and counted. Plants grown in soil infested with eggs and receiving dry ice treatment had less than 1% of the eggs found in the controls; plants from soil infested with root fragments and receiving dry ice treatment had less than 4% of the eggs found in controls. Dry ice used to lower soil temperature may have potential as a cryonematicide.     

The Effects of Root-knot Nematode Infection and Mi-mediated Nematode Resistance in Tomato on Plant Fitness

The Mi-1.2 resistance gene in tomato (Solanum lycopersicum) confers resistance against several species of root-knot nematodes (Meloidogyne spp.). This study examined the impact of M. javanica on the reproductive fitness of near-isogenic tomato cultivars with and without Mi-1.2 under field and greenhouse conditions. Surprisingly, neither nematode inoculation or host plant resistance impacted the yield of mature fruits in field microplots (inoculum=8,000 eggs/plant), or fruit or seed production in a follow-up greenhouse bioassay conducted with a higher inoculum level (20,000 eggs/plant). However, under heavy nematode pressure (200,000 eggs/plant), greenhouse-grown plants carrying Mi-1.2 had more than ten-fold greater fruit production than susceptible plants and nearly forty-fold greater estimated lifetime seed production, confirming prior reports of the benefits of Mi-1.2. In all cases Mi-mediated resistance significantly reduced nematode reproduction. These results indicated that tomato can utilize tolerance mechanisms to compensate for moderate levels of nematode infection, but that the Mi-1.2 resistance gene confers a dramatic fitness benefit under heavy nematode pressure. No significant cost of resistance was detected in the absence of nematode infection.     

Reproduction of Meloidogyne incognita on Winter Cover Crops Used in Cotton Production

Substantial reproduction of Meloidogyne incognita on winter cover crops may lead to damaging populations in a subsequent cotton (Gossypium hirsutum) crop. The amount of population increase during the winter depends on soil temperature and the host status of the cover crop. Our objectives were to quantify M. incognita race 3 reproduction on rye (Secale cereale) and several leguminous cover crops and to determine if these cover crops increase population densities of M. incognita and subsequent damage to cotton. The cover crops tested were ‘Bigbee’ berseem clover (Trifolium alexandrinum), ‘Paradana’ balansa clover (T. balansae), ‘AU Sunrise’ and ‘Dixie’ crimson clover (T. incarnatum), ‘Cherokee’ red clover (T. pratense), common and ‘AU Early Cover’ hairy vetch (Vicia villosa), ‘Cahaba White’ vetch (V. sativa), and ‘Wrens Abruzzi’ rye. In the greenhouse tests, egg production was greatest on berseem clover, Dixie crimson clover, AU Early Cover hairy vetch, and common hairy vetch; intermediate on Balansa clover and AU Sunrise crimson clover; and least on rye, Cahaba White vetch, and Cherokee red clover. In both 2002 and 2003 field tests, enough heat units were accumulated between 1 January and 20 May for the nematode to complete two generations. Both AU Early Cover and common hairy vetch led to greater root galling than fallow in the subsequent cotton crop; they also supported high reproduction of M. incognita in the greenhouse. Rye and Cahaba White vetch did not increase root galling on cotton and were relatively poor hosts for M. incognita. Only those legumes that increased populations of M. incognita reduced cotton yield. In the southern US, M. incognita can complete one to two generations on a susceptible winter cover crop, so cover crops that support high nematode reproduction may lead to damage and yield losses in the following cotton crop. Planting rye or Meloidogyne-resistant legumes as winter cover crops will lower the risk of increased nematode populations compared to most vetches and clovers.     

Effect of Gamma-irradiation and Heat on Root-knot Nematode,Meloidogyne javanica

Effects of gamma-irradiation on the root-knot nematode Meloidogyne javanica were investigated. A dose of 7.5 kGy killed all second-stage juveniles (J2) within 1 day after treatment. Egg hatch was completely inhibited at 6.25 kGy. A bioassay on tomato measuring galling and egg production was used to determine the infectivity of irradiated J2 and J2 hatched from irradiated eggs. The J2 and eggs irradiated with a dose of 4.25 kGy did not induce galls or reproduce on tomato plants. When nematodes were exposed to combined irradiation and heat treatment, no synergistic effect on J2 or eggs was measured. Heat treatment at 49° C for 10 minutes or 20 minutes without irradiation immobilized J2 and prevented egg development. Irradiation rates needed to kill or incapacitate M. javanica were high and may be impractical as a quarantine measure.     

Detection and Investigation of Soil Biological Activity against Meloidogyne incognita

Greenhouse experiments with two susceptible hosts of Meloidogyne incognita, a dwarf tomato and wheat, led to the identification of a soil in which the root-knot nematode population was reduced 5- to 16-fold compared to identical but pasteurized soil two months after infestation with 280 M. incognita J2/100 cm³ soil. This suppressive soil was subjected to various temperature, fumigation and dilution treatments, planted with tomato, and infested with 1,000 eggs of M. incognita/100 cm³ soil. Eight weeks after nematode infestation, distinct differences in nematode population densities were observed among the soil treatments, suggesting the suppressiveness had a biological nature. A fungal rRNA gene analysis (OFRG) performed on M. incognita egg masses collected at the end of the greenhouse experiments identified 11 fungal phylotypes, several of which exhibited associations with one or more of the nematode population density measurements (egg masses, eggs or J2). The phylotype containing rRNA genes with high sequence identity to Pochonia chlamydosporia exhibited the strongest negative associations. The negative correlation between the densities of the P. chlamydosporia genes and the nematodes was corroborated by an analysis using a P. chlamydosporia-selective qPCR assay.     

南方根结线虫对不同砧木嫁接番茄苗活性氧清除系统的影响

采用盆栽人工接种方法,对番茄嫁接苗进行了抗性评价,研究了番茄嫁接苗叶片中抗氧化酶活性和活性氧代谢的动态变化。结果表明,接种南方根结线虫(J2)后,砧木嫁接苗表现为高抗,自根嫁接苗为高感。通过嫁接换根,与自根嫁接苗相比,砧木嫁接苗明显提高了接穗叶片的超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)活性,降低了超氧阴离子(O.2-)产生速率以及过氧化氢(H2O2)和丙二醛(MDA)含量。表明番茄植株体内的活性氧水平和抗氧化酶活性的高低与其抗根结线虫的能力密切相关,较低的活性氧水平和较高的抗氧化酶活性有利于减轻对膜系统的伤害,提高番茄植株的抗根结线虫能力。     

Effects of the Mi-1 and the N root-knot nematode-resistance gene on infection and reproduction of Meloidogyne enterolobii on tomato and pepper cultivars

Meloidogyne enterolobii is widely considered to be an aggressive root-knot nematode species that is able to reproduce on root-knot nematode-resistant tomato and pepper cultivars. In greenhouse experiments, M. enterolobii isolates 1 and 2 from Switzerland were able to reproduce on tomato cultivars carrying the Mi-1 resistance gene as well as an N-carrying pepper cultivar. Reproduction factors (Rf) ranged between 12 and 109 depending on the plant cultivar, with M. enterolobii isolate 2 being more virulent when compared to isolate 1. In contrast, M. arenaria completely failed to reproduce on these resistant tomato and pepper cultivars. Although some variability in virulence and effectiveness of root-knot nematode-resistance genes was detected, none of the plant cultivars showed Rf values less than 1 or less than 10% of the reproduction observed on the susceptible cv. 'Moneymaker' (Rf = 23-44) used to characterize resistance. The ability of M. enterolobii to overcome the resistance of tomato and pepper carrying the Mi-1 and the N gene makes it difficult to manage this root-knot nematode species, particularly in organic farming systems where chemical control is not an option.     

Host Plant Resistance as an Alternative to Methyl Bromide for Managing Meloidogyne incognita in Pepper

Pre-plant soil fumigation with methyl bromide and host resistance were compared for managing the southern root-knot nematode (Meloidogyne incognita) in pepper. Three pepper cultivars (Carolina Cayenne, Keystone Resistant Giant, and California Wonder) that differed in resistance to M. incognita were grown in field plots that had been fumigated with methyl bromide (98% CH₃Br : 2% CCl₃NO₂ [w/w]) before planting or left untreated. Carolina Cayenne is a well-adapted cayenne-type pepper that is highly resistant to M. incognita. The bell-type peppers Keystone Resistant Giant and California Wonder are intermediate to susceptible and susceptible, respectively. None of the cultivars exhibited root galling in the methyl bromide fumigated plots and nematode reproduction was minimal (<250 eggs/g fresh root), indicating that the fumigation treatment was highly effective in controlling M. incognita. Root galling of Carolina Cayenne and nematode reproduction were minimal, and fruit yields were not reduced in the untreated plots. The root-galling reaction for Keystone Resistant Giant was intermediate (gall index = 2.9, on a scale of 1 to 5), and nematode reproduction was moderately high. However, yields of Keystone Resistant Giant were not reduced in untreated plots. Root galling was severe (gall index = 4.3) on susceptible California Wonder, nematode reproduction was high, and fruit yields were reduced (P ≤ 0.05) in untreated plots. The resistance exhibited by Carolina Cayenne and Keystone Resistant Giant provides an alternative to methyl bromide for reducing yield losses by southern root-knot nematodes in pepper. The high level of resistance of Carolina Cayenne also suppresses population densities of M. incognita.     

Interaction of Fusarium oxysporum f. sp. ciceri and Meloidogyne javanica on Cicer arietinum

Interaction of Meloidogyne javanica and Fusarium oxysporum f. sp. ciceri was studied on Fusarium wilt-susceptible (JG 62 and K 850) and resistant (JG 74 and Avrodhi) chickpea cultivars. In greenhouse experiments, inoculation of M. javanica juveniles prior to F. oxysporum f. sp. ciceri caused greater wilt incidence in susceptible cultivars and induced vascular discoloration in roots of resistant cultivars. Nematode reproduction was greatest (P = 0.05) at 25 °C. Number of galls and percentage of root area galled increased when the temperature was increased from 15 °C to 25 °C. Wilt incidence was greater at 20 °C than at 25 °C. Chlorosis of leaves and vascular discoloration of plants did not occur at 15 °C. The nematode enhanced the wilt incidence in wilt-susceptible cultivars only at 25 °C. Interaction between the two pathogens on shoot and root weights was significant only at 20 °C, and F. o. ciceri suppressed the nematode density at this temperature. Wilt incidence was greater in clayey (48% clay) than in loamy sand (85% sand) soils. The nematode caused greater plant damage on loamy sand than on clayey soil. Fusarium wilt resistance in Avrodhi and JG 74 was stable in the presence of M. javanica across temperatures and soil types.     

Morphological and Molecular Evaluation of a Meloidogyne hapla Population Damaging Coffee (Coffea arabica) in Maui,Hawaii

An unusual population of Meloidogyne hapla, earlier thought to be an undescribed species, was found causing large galls, without adventitious roots, and substantial damage to coffee in Maui, Hawaii. Only in Brazil had similar damage to coffee been reported by this species. Unlike M. exigua from South and Central America, this population reproduced well on coffee cv. Mokka and M. incognita-susceptible tomato but poorly on tomato with the Mi resistance gene. Characterization included SEM images, esterase isozymes, and five DNA sequences: i) the D3 segment of the large subunit (LSU-D3 or 28S) rDNA, ii) internal transcribed spacer (ITS-1) rDNA, iii) intergenic spacer (IGS) rDNA, iv) the mitochondrial interval from cytochrome oxidase (CO II) to 16S mtDNA, and v) the nuclear gene Hsp90. Sequences for ITS-1, IGS, and COII were similar to other M. hapla populations, but within species ITS-1 variability was not less than among species. One LSU-D3 haplotype was similar to a previously analyzed population with two minor haplotypes. Hsp90 exhibited some variation between Maryland and Hawaiian populations distinct from other species. Females were narrow with wide vulval slits, large interphasmidial distances, and more posterior excretory pores; 20% of perineal patterns had atypical perivulval lines. Males had a low b ratio (<12 µm). Juveniles had a short distance between stylet and dorsal gland orifice. Juvenile body length was short (<355 µm) and was different between summer and winter populations.     

Identification of Sources of Resistance to Four Species of Root-knot Nematodes in Tobacco

Resistance to the southern root-knot nematode, Meloidogyne incognita races 1 and 3, has been identified, incorporated, and deployed into commercial cultivars of tobacco, Nicotiana tabacum. Cultivars with resistance to other economically important root-knot nematode species attacking tobacco, M. arenaria, M. hapla, M. javanica, and other host-specific races of M. incognita, are not available in the United States. Twenty-eight tobacco genotypes of diverse origin and two standard cultivars, NC 2326 (susceptible) and Speight G 28 (resistant to M. incognita races 1 and 3), were screened for resistance to eight root-knot nematode populations of North Carolina origin. Based on root gall indices at 8 to 12 weeks after inoculation, all genotypes except NC 2326 and Okinawa were resistant to M. arenaria race 1, and races 1 and 3 of M. incognita. Except for slight root galling, genotypes resistant to M. arenaria race 1 responded similarly to races 1 and 3 of M. incognita. All genotypes except NC 2326, Okinawa, and Speight G 28 showed resistance to M. javanica. Okinawa, while supporting lower reproduction of M. javanica than NC 2326, was rated as moderately susceptible. Tobacco breeding lines 81-R-617A, 81-RL- 2K, SA 1213, SA 1214, SA 1223, and SA 1224 were resistant to M. arenaria race 2, and thus may be used as sources of resistance to this pathogen. No resistance to M. hapla and only moderate resistance to races 2 and 4 of M. incognita were found in any of the tobacco genotypes. Under natural field infestations of M. arenaria race 2, nematode development on resistant tobacco breeding lines 81-RL-2K, SA 1214, and SA 1215 was similar to a susceptible cultivar with some nematicide treatments; however, quantity and quality of yield were inferior compared to K 326 plus nematicides.