Quantitative approach for the early detection of selection for virulence of Meloidogyne incognita on resistant tomato in plastic greenhouses

Resistant tomato cultivars are an important tool to control Meloidogyne spp., which cause the highest yield losses attributed to plant‐parasitic nematodes. However, the repeated cultivation of Mi resistant cultivars can select virulent populations. In the present study, the susceptible tomato cv. Durinta and the resistant cv. Monika were cultivated from March to July in a plastic greenhouse for 3 years to determine the maximum multiplication rate, maximum nematode density, equilibrium density, relative susceptibility and population growth rate of M. incognita; these were used as proxy indicators of virulence and yield losses. The values of population dynamics and growth rate on the resistant tomato increased year by year and were higher when it was repeatedly cultivated in the same plot compared to when it was alternated with the susceptible cultivar and the level of resistance decreased from very to moderately resistant. The relationship between the nematode density at transplanting (P_i) and the relative yield of tomato fitted to the Seinhorst damage model for susceptible, but not resistant, cultivars. The tolerance limit and the relative minimum yield were 2–4 J2 per 250 cm³ of soil and 0.44–0.48, respectively. The tomato yield did not differ between cultivars at low P_i, but it did at higher P_i values, at which the resistant yielded 50% more than the susceptible. This study demonstrates the utility of population dynamics parameters for the early detection of selection for virulence in Meloidogyne spp., and that three consecutive years were not sufficient to select for a completely virulent population.     

南方根结线虫中国分离群体种内变异分析

为调查我国不同地区和不同寄主上的根结线虫Meloidogyne spp.种类分布以及群体变异情况,基于酯酶和苹果酸脱氢酶同工酶图谱及SCAR分子标记技术对2017—2019年从6省19种植物根部组织分离到的40个根结线虫群体进行鉴定,针对南方根结线虫M. incognita群体分别通过寄主鉴别法进行生理小种鉴别,利用携带Mi抗性基因的番茄进行毒力测试,对2龄幼虫的口针长度和体长进行测量,并对核糖体ITS和线粒体Nad5基因序列进行比较分析。结果显示:根结线虫分离群体经鉴定包括38个南方根结线虫群体和2个象耳豆根结线虫M. erterolobii群体;38个南方根结线虫群体中有35个群体被鉴别为1号生理小种,其余3个群体被鉴别为2号生理小种;发现1个南方根结线虫群体CN19可在携带Mi抗性基因的番茄上侵染繁殖,为毒性群体,其余群体无法进行侵染和繁殖,为无毒群体。南方根结线虫群体2龄幼虫的口针长度和体长均差异较大,而不同寄主来源分离群体的ITS和Nad5基因序列也存在一定变异。基于ITS和Nad5基因序列构建的系统发育树将所有根结线虫群体归为南方根结线虫和象耳豆根结线虫组成的2个独立分支,...     

Diversity of Meloidogyne incognita populations from cotton and aggressiveness to Gossypium spp. accessions

The root-knot nematode (RKN) Meloidogyne incognita is the main nematode causing losses to the cotton crop in Brazil. In order to implement control strategies within integrated management, an accurate identification of the nematode populations prevailing in the cotton production areas is necessary. This study aimed to assess the genetic variability and aggressiveness of RKN populations from cotton production areas in Bahia state, Brazil. All populations were characterized biochemically and molecularly and identified as M. incognita. RAPD and AFLP markers detected 44% of polymorphic fragments among the 13 populations of this species. The 10 M. incognita populations collected in Bahia presented 33.7% of diversity when compared to each other, and 25% when the population from Barreiras (the most polymorphic) was excluded. This polymorphism increased when populations from other Brazilian states were included. The aggressiveness and virulence among populations from Bahia towards different cotton accessions (susceptible/resistant) was also studied. None of the populations showed virulence against the moderately resistant (Clevewilt 6, Wild Mexican Jack Jones and LA 887) and highly resistant (CIR1348 and M-315 RNR) cultivars. Two M. incognita populations from Barreiras were the most aggressive, reaching reproduction factors of 539 and 218, respectively, in the susceptible cultivar FiberMax 966. The most aggressive population (8) was also the most genetically divergent in phylogenetic analyses. These results demonstrate that diversity of M. incognita populations from cotton farms in Bahia is not related to virulence against resistant accessions, which suggests that cultivars containing one or two resistance genes with good agronomic characteristics could be used in infested commercial areas in Bahia state, Brazil.     

Occurrence of virulent root-knot nematode populations on tomatoes bearing the <Emphasis Type="Italic">Mi</Emphasis> gene in protected vegetable-growing areas of Turkey

Root-knot nematodes (Meloidogyne spp.; RKN) are one of the most important pathogens of vegetables in Turkey. Assessing the existing virulent RKN populations is of importance for pathogen mapping in the west Mediterranean region of Turkey. Therefore, 95 populations of RKN were collected from different protected vegetable-growing locations in the region. Pure cultures were obtained and identified by means of species-specific primers. Virulence of the populations against the Mi-1 gene conferring resistance to Meloidogyne incognita, M. javanica and M. arenaria was determined according to their egg masses and gall rating on resistant and susceptible tomato varieties. Results showed that seven populations of M. incognita and six populations of M. javanica were able to overcome the resistance controlled by the Mi-1 gene. The frequency of virulent populations of M. incognita and M. javanica collected from different protected-grown vegetables was 11.7% and 21.4%, respectively. To our knowledge, this is the first report of populations of RKN virulent to the Mi-1 gene in Turkey.     

Genetic variability and virulence of Meloidogyne incognita populations from Brazil to resistant cotton genotypes

The root-knot nematode Meloidogyne incognita is widely distributed and a major pathogen of cotton (Gossypium spp.) worldwide. The objectives of this study were to assess the genetic variability and aggressiveness of Brazilian populations of M. incognita in cotton. Five populations of M. incognita and one isolate of M. enterolobii (outgroup) were used in the molecular analysis. Our results showed that only 2.7 % of the RAPD and AFLP fragments were polymorphic. Despite the existence of two races (races 3 and 4) and two esterase phenotypes (I1 and I2), a low genetic variability among populations was observed, which might be due to the mitotic parthenogenetic mode of reproduction of this pathogen. The aggressiveness/virulence among populations towards different cotton genotypes was also studied. None of the populations was virulent to the resistant cotton genotypes M-315 RNR, TX-25, CIR1343, Wild Mexican Jack Jones and CIR1348 (reproduction factor <1). Two populations of M. incognita from the states of Mato Grosso do Sul and Parana (Umuarama) (races 4 and 3, respectively) were highly aggressive to the susceptible control FM966 and virulent to the accessions LA-887 and Clevewilt-6 that showed moderate resistance to other populations tested.     

Selection forMeloidogyne incognita virulence against resistance genes from tomato and pepper and specificity of the virulence/resistance determinants

Experiments were designed to analyze the relationships between the root-knot nematodeMeloidogyne incognita and resistant tomato and pepper genotypes. From a natural avirulent isolate, near-isogenic nematode lineages were selected with virulence either against the tomatoMi resistance gene or the pepperMe3 resistance gene. Despite the drastic selection pressure used, nematodes appeared unable to overcome the pepperMe1 gene, therefore suggesting some differences in the resistance conferred byMe1 andMe3 in this species. Nematodes virulent onMi-resistant tomatoes were not able to reproduce onMe1-resistant nor onMe3-resistant peppers, and nematodes virulent onMe3-resistant peppers were not able to reproduce onMi-resistant tomatoes nor onMe1-resistant peppers. These results clearly demonstrate the specificity ofM. incognita virulence against resistance genes from both tomato and pepper, and indirectly suggest that gene-for-gene relationships could occur between these two solanaceous crops and the nematode.     

The reproductive potential of the root-knot nematode <Emphasis Type="Italic">Meloidogyne incognita</Emphasis> is affected by selection for virulence against major resistance genes from tomato and pepper

The emergence of virulent root-knot nematode populations, able to overcome the resistance conferred by some of the resistance genes (R-genes) in Solanaceous crops, i.e., Mi(s) in tomato, Me(s) in pepper, may constitute a severe limitation to their use in the field. Research has been conducted to evaluate the durability of these R-genes, by comparing the reproduction of several laboratory-selected and wild virulent Meloidogyne incognita isolates, on both susceptible and resistant tomatoes and peppers. We first show that the Me1 R-gene in pepper behaves as a robust R-gene controlling avirulent and virulent Me3, Me7 or Mi-1 isolates. Although the reproductive potential of the virulent isolates was highly variable on susceptible and resistant plants, we also confirm that virulence is highly specific to a determined R-gene on which selection has occurred. Another significant experimental result is the observation that a reproductive fitness cost is associated with nematode virulence against Mi-1 in tomato and Me3 and Me7 in pepper. The adaptative significance of trade-offs between selected characters and fitness-related traits, suggests that, although the resistance can be broken, it may be preserved in some conditions if the virulent nematodes are counter-selected in susceptible plants. All these results have important consequences for the management of plant resistance in the field.     

Host suitability of Solanum torvum cultivars to Meloidogyne incognita and M. javanica and population dynamics

Several experiments were carried out to assess the performance of commercial Solanum torvum cultivars against the root knot nematodes Meloidogyne incognita and M. javanica in Spain. The response of S. torvum rootstock cultivars Brutus, Espina, Salutamu and Torpedo against M. incognita and Mi-1.2 (a)virulent M. javanica isolates was determined in pot experiments, and of ‘Brutus’ to an N-virulent isolate of M. incognita, compared with that of the eggplant S. melongena ‘Cristal’. The relationship between the initial and final population densities of M. javanica on ungrafted and grafted ‘Cristal’ onto the S. torvum ‘Brutus’ was assessed, together with the effect on dry shoot biomass. Finally, the population growth rate and the resistance level of the four S. torvum cultivars against M. incognita was assessed under plastic greenhouse conditions in two cropping seasons. All S. torvum rootstocks responded as resistant to the M. incognita isolates and from highly resistant to susceptible against M. javanica isolates. The maximum multiplication rates of M. javanica on the ungrafted or grafted eggplant were 270 and 49, respectively, and the equilibrium densities were 1318 and 2056 eggs and J2 per 100 cm³ soil, respectively. The tolerance of the ungrafted eggplant was 10.9 J2 per 100 cm³ soil, and the minimum relative dry shoot biomass was 0.76. The population growth rate of M. incognita on eggplant cv. Cristal differed from that of the S. torvum cultivars in both cropping seasons. These results suggest that S. torvum is a valuable rootstock for managing the two Meloidogyne species irrespective of the (a)virulence status.     

Occurrence of Resistance-breaking Populations of Root-knot Nematodes on Tomato in Greece

Nine populations of Meloidogyne spp. from Greece have been identified as M. javanica or M. incognita using either isozyme phenotypes or the sequence characterized amplified region-polymerase chain reaction (SCAR-PCR) technique. Virulence against the Mi resistance gene was assayed by pot experiments in controlled conditions and revealed the ability of five populations of M. javanica and one population of M. incognita to reproduce on tomato cultivars containing that gene. A resistance-breaking population of M. incognita is reported for the first time in the country; the M. javanica populations constitute new records for the Greek mainland.     

Effectiveness and profitability of the <Emphasis Type="Italic">Mi</Emphasis>-resistant tomatoes to control root-knot nematodes

Experiments were conducted to determine the effectiveness and profitability of the Mi-resistance gene in tomato in suppressing populations of Meloidogyne javanica in a plastic-house with a natural infestation of the nematode. Experiments were also conducted to test for virulence and durability of the resistance. Monika (Mi-gene resistant) and Durinta (susceptible) tomato cultivars were cropped for three consecutive seasons in non-fumigated or in soil fumigated with methyl bromide at 75 g m^–2 and at a cost of 2.44 euros m^–2. Nematode densities were determined at the beginning and end of each crop. Yield was assessed in eight plants per plot weekly for 6 weeks. The P_f/P_i values were 0.28 and 21.6 after three crops of resistant or susceptible cultivars, respectively. Growth of resistant as opposed to susceptible tomato cultivars in non-fumigated soil increased profits by 30,000 euros ha^–1. The resistant Monika in non-fumigated soil yielded similarly (P > 0.05) to the susceptible Durinta in methyl bromide fumigated soil but the resistant tomato provided a benefit of 8800 euros ha^–1 over the susceptible one because of the cost of fumigation. Selection for virulence did not occur, although the nematode population subjected to the resistant cultivar for three consecutive seasons produced four times more eggs than the population on the susceptible one. Such a difference was also shown when the resistant cultivar was subjected to high continuous inoculum pressure for 14 weeks. The Mi-resistance gene can be an effective and economic alternative to methyl bromide in plastic-houses infested with root-knot nematodes, but should be used in an integrated management context to preserve its durability and prevent the selection of virulent populations due to variability in isolate reproduction and environmental conditions.     

Genetic variability of Meloidogyne paranaensis populations and their aggressiveness to susceptible coffee genotypes

Meloidogyne paranaensis is one of the most destructive root‐knot nematode (RKN) species parasitizing coffee in Brazil and in the Americas generally. The objectives of this study were to assess the genetic variability, aggressiveness and virulence of seven different M. paranaensis populations on susceptible and resistant Coffea spp. All seven RKN populations were identified by biochemical and molecular methods. Coffee seedlings were inoculated in the greenhouse, and the nematode reproduction factor was used to infer their reproduction on coffee genotypes. Phylogenetic studies showed a low genetic variability in M. paranaensis populations, regardless of the existence of three esterase phenotypes (Est P1, P2 and P2a), except for the population Est P2a from Guatemala, which is genetically different from other M. paranaensis populations from Brazil. The Est P2a and Est P2 (Herculândia, SP, Brazil) populations were the most aggressive on two susceptible C. arabica cultivars under greenhouse conditions. None of the M. paranaensis populations were virulent on resistant coffee genotypes, confirming their resistance to the seven M. paranaensis populations tested. The resistant coffee cultivars, namely Clone 14 INCAPER, Catuaí Vermelho × Amphillo MR2161 (E1 16‐5 III), Apoatã IAC 2258, Timor Hybrid UFV 408‐01 (E1 6‐6 II) and IPR 100, exhibited segregation for resistance in the ratio of 0%, 2.4%, 12%, 26% and 29%, respectively. These are promising results, because they validate resistance against several M. paranaensis populations in different Coffea spp. genetic resources, which can be used in breeding programmes or as rootstocks, such as Apoatã IAC 2258 and Clone 14 INCAPER.     

PIN1 auxin efflux carrier absence in Meloidogyne incognita-induced root-knots of tomato plants

The nematode species Meloidogyne incognita infects a large variety of cultivated crops and is one of nature’s most notorious pests. One cultivated plant which is prone to M. incognita infestation is the tomato. Knowing that in A. thaliana the PIN auxin efflux transporters distribution is being altered upon early invasion by M. incognita, the PIN1 allocation patterns in the giant cells of tomato plants after 15 and 25 days of infection were investigated. PIN1 was absent from the giant cells’ membrane in both assessment timings examined, indicating the maintenance of a local auxin maxima, which was also supported by IAA immunodetection. PIN1 distribution pattern could be attributed on either the nematodes nutritional needs throughout its life cycle or due to the tomato plants differential responses upon M. incognita infection.

     

Stimulation of respiratory pathways in tomato roots infested by Meloidogyne incognita

Respiration of tomato roots susceptible and resistant to Meloidogyne incognita was measured during infestation. No significant changes in respiratory rate occurred in susceptible tomato roots, during infestation by M. incognita. In resistant tomato roots, a pronounced increase of both cyanide-sensitive and cyanide-resistant oxidases, was observed during nematode attack. The time-course of the respiration during 12 days, after nematode inoculation, showed that resistant tomato roots responded with a rapid increase in cyanide-sensitive and cyanide-resistant respiration as invading nematodes progressed; no changes were observed in the susceptible tomato roots.Change in the rate of oxygen uptake paralleled an increase in nematode density in resistant tomato roots; oxygen uptake rose linearly to an infestation level of 50 juveniles for each seedling, above which value it declined. The physiological significance of the alternative respiratory pathway is discussed.     

Screening Tomato Cultivars for Resistance to Meloidogyne Species

Abstract

Screening tests on 162 tomato cultivars and hybrids conducted at the Division of Horticulture, Indian Agricultural Research Institute, New Delhi during 1967-8 indicated a high degree of resistance to root-knot nematodes in Nematex, VFN-8, 65N215-1, 65N255-1 and S1-120, under field and laboratory conditions. The degree of resistance varied with the species of Meloidogyne. Nematex was immune to M. javanica, M. incognita and M. arenaria. VFN-8, 65N215-1 and 65N255-1 were immune to M. incognita and M. arenaria and resistant to M. javanica. S1-120, a commercial cultivar, showed a high degree of tolerance to M. javanica, M. incognita and M. arenaria. The degree of resistance varied with nematode population density.     

Yield Responses in Relation to the Chemical Control of Root-knot Nematodes in Southern Italy

Nematicide trials have been conducted in Southern Italy to determine the yield response of tobacco, tomato and egg-plant. The nematode species involved were mainly Meloidogyne incognita (Kofoid & White) Chitwood and M. arenaria (Neal) Chitwood. Many nematicides have an impact also on other soil pathogens or even on insects, so that the yied increase observed in the field has to be considered as a combined effect. On tobacco, most of the nematicides gave positive results, but it appeared that there might be important differences according to the variety considered. In certain tomato fields, which had not been specially attacked by Meloidogyne, the positive effects seemed to be also due to the insecticidal or fungicidal effect of the chemicals. No statistically significant yield increases were, however, obtained on treated egg-plant fields.     

Genetics of virulence in Ascochyta rabiei

In order to critically test the hypothesis that virulence variation in the Ascochyta rabiei/chickpea pathosystem is a discrete character under simple genetic control, a genetic cross was made between a highly virulent isolate of A. rabiei from Syria and a less virulent isolate from the USA. Two independent virulence assays conducted by inoculating susceptible and resistant chickpea cultivars under controlled conditions with 77 independent progeny isolates from this cross revealed a continuous distribution of disease phenotypes. Bimodality, as would be predicted for the segregation of virulence under simple genetic control, was not supported by statistical tests of the progeny phenotype distribution. anova revealed highly significant pathogen‐genotype × host‐genotype interactions demonstrating the segregation of genes controlling specialization on the two cultivars tested. These interactions could be localized to two isolates that changed virulence rank on the cultivars. It was concluded that variation in virulence to these two cultivars is under quantitative genetic control. If this conclusion applies to other cultivars, it can be speculated that the discrete categories of virulence variation identified in previous studies were probably the result of incomplete sampling of host resistance or pathogen virulence variation and/or of selection for increased virulence in contemporary A. rabiei populations.     

ROS and NO production in compatible and incompatible tomato-<Emphasis Type="Italic">Meloidogyne incognita</Emphasis> interactions

Nitric oxide (NO) has been shown to be an essential regulatory molecule in plant response to pathogen infection in synergy with reactive oxygen species (ROS). At the present, nothing is known about the role of NO in disease resistance to nematode infection. We used a resistant tomato cultivar with different sensitivity to avirulent and virulent populations of the root-knot nematode Meloidogyne incognita to investigate the key components involved in oxidative and nitrosative metabolism. We analyzed the superoxide radical production, hydrogen peroxide content, and nitric oxide synthase (NOS)-like and nitrate reductase activities, as potential sources of NO. A rapid NO accumulation and ROS production were found at 12 h after infection in compatible and incompatible tomato-nematode interactions, whereas the amount of NO and ROS gave different results 24 and 48 h after infection amongst compatible and incompatible interactions. NOS-like arginine-dependent enzyme rather than nitrate reductase was the main source of NO production, and NOS-like activity increased substantially in the incompatible interaction. We can envisage a functional overlap of both NO and ROS in tomato defence response to nematode invasion, NO and H₂O₂ cooperating in triggering hypersensitive cell death. Therefore, NO and ROS are key molecules which may help to orchestrate events following nematode challenge, and which may influence the host cellular metabolism.     

Toxicity of fungal endophyte secondary metabolites to plant parasitic nematodes and soil-borne plant pathogenic fungi

Fungi isolated from the cortical tissue of surface sterilized tomato roots collected from field plots produced secondary metabolites in nutrition broth that were highly toxic toMeloidogyne incognita. Especially strains ofFusarium oxysporum were highly active with 13 of 15 strains producing culture filtrates toxic to nematodes. The mechanism of action of the toxic metabolites produced by the non-pathogenicF. oxysporum strain 162 with proven biological control ofM. incognita in pot experiments was investigated. These metabolites reducedM. incognita mobility within 10 min of exposure. After 60 min, 98% of juveniles were inactivated. Juveniles were initially inactivated within a few minutes of exposure, but with exposure of 5 h 50% of the juveniles were dead and 24 h exposure resulted in 100% mortality. In a bioassay with lettuce seedlings metabolite concentrations > 100 mg/l reduced the number ofM. incognita juveniles on the roots comparing to the water control. TheF. oxysporum toxins were highly effective towards sedentary parasites and less effective towards migratory endoparasites. Nonparasitic nematodes were not influenced at all. Metabolites of strain 162 also reduced significantly the growth ofPhytophthora cactorum, Pythium ultimum andRhizoctonia solani in vitro. Secondary metabolites of endophytic fungi on plant-parasitic nematodes and soil-borne fungi should be considered for control of plant parasitic nematodes and plant pathogenic fungi. The results also show the need for proper selection of target nematodes inin vitro bioassays.     

Thermal requirements for the embryonic development and life cycle of Meloidogyne hispanica

The life cycle of a Portuguese Meloidogyne hispanica isolate on susceptible cv. Easypeel and resistant (Mi‐1.2 gene) cv. Rossol tomato plants was studied in growth chambers at constant temperatures (10–35°C). The development within the egg and hatching were compared to those of a Portuguese M. arenaria isolate. The base temperature was 10·11 and 8·31°C with 179·5 and 235·3 thermal units for M. hispanica and M. arenaria, respectively, suggesting better potential adaptation to low temperatures by M. arenaria than M. hispanica. No egg development occurred at 10 or 35°C. An increase in invasion of tomato roots by M. hispanica second‐stage juveniles (J2s) was correlated with an increase in temperature on both tomato cultivars. Tomato cv. Rossol limited M. hispanica development at 20, 25 and 30°C, but not at 35°C, indicating that these high temperatures blocked the resistance mechanism provided by the Mi‐1.2 gene. At 15°C, J2s penetrated tomato cv. Rossol roots, but failed to develop and establish feeding sites. On tomato cv. Easypeel, nematode development and reproduction occurred at 20, 25 and 30°C, but at 20°C the life cycle was 1·5 and 2·0 times longer than at 25 and 30°C, respectively. No egg production was observed at 15°C. The results of this study showed that M. hispanica is most suited to soil temperatures around 25°C. Predicted climate change might favour the spread of this nematode species into southern Europe and northwards. The thermal requirements for M. hispanica development are analysed and compared with those of M. arenaria, M. hapla, M. incognita and M. javanica.     

Response of tomato rootstocks carrying the <Emphasis Type="Italic">Mi</Emphasis>-resistance gene to populations of <Emphasis Type="Italic">Meloidogyne arenaria,M. incognita and M. javanica</Emphasis>

The response of four Mi-resistance gene tomato rootstocks to seven populations of Meloidogyne was determined in pot tests conducted in a glasshouse. Rootstocks PG76 (Solanum lycopersicum × Solanum sp.) and Brigeor (S. lycopersicum × S. habrochaites) and resistant cv. Monika (S. lycopersicum) were assessed against one population of M. arenaria, three of M. incognita, and three of M. javanica. Rootstocks Beaufort and Maxifort were assessed against one population of M. arenaria, two of M. incognita and two of M. javanica. Rootstock PG76 was highly resistant (reproduction index <10%) to all the populations, whereas rootstock Brigeor and cv. Monika were highly to moderate resistant. Rootstocks Beaufort and Maxifort showed reduced resistance or inability to suppress nematode reproduction, and their responses varied according to the population tested. Beaufort and Maxifort were susceptible to the two populations of M. javanica as Maxifort was to one of M. incognita. The reproduction index of the nematode was higher (P < 0.05) on Maxifort than Beaufort for all root-knot nematode populations.