Meloidogyne Research Articles

Identification of a Promising Novel Genetic Source for Rice Root-Knot Nematode Resistance through Markers Associated with Trait-Specific Quantitative Trait Loci.

Direct-seeded rice (DSR) is gaining popularity among farmers due to its environmentally safe and resource-efficient production system. However, managing the rice root-knot nematode (RRKN), Meloidogyne graminicola, remains a major challenge in DSR cultivation. Developing genetic resistance is a pragmatic and effective approach compared to using hazardous pesticides. Pusa Basmati 1121 (PB1121) is the most popular Basmati rice variety, but it is highly susceptible to RRKN. In contrast, Phule Radha (PR) has shown highly resistant reaction to RRKN, as reported in our earlier study. We generated an F2:3 population from the cross of PB1121/PR and evaluated it for RRKN resistance-related traits under artificial inoculation conditions. The distribution pattern of traits in the F2:3 population indicated that resistance may be governed by a few major-effect genes and many minor-effect genes. The molecular markers reported to be associated with QTLs governing RRKN resistance traits were used to test in the current population. Although the simple linear regression identified significant associations between the markers and RRKN resistance-associated traits, these associations were spurious as the LOD score was below the threshold limit. This indicates that PR possesses novel genomic regions for resistance to RRKN as it does not possess any of the earlier reported QTLs.

Study on the Function of SlWRKY80 in Tomato Defense against Meloidogyne incognita.

WRKY transcription factors (TFs) can participate in plant biological stress responses and play important roles. SlWRKY80 was found to be differentially expressed in the Mi-1- and Mi-3-resistant tomato lines by RNA-seq and may serve as a key node for disease resistance regulation. This study used RNAi to determine whether SlWRKY80 silencing could influence the sensitivity of 'M82' (mi-1/mi-1)-susceptible lines to M. incognita. Further overexpression of this gene revealed a significant increase in tomato disease resistance, ranging from highly susceptible to susceptible, combined with the identification of growth (plant height, stem diameter, and leaf area) and physiological (soluble sugars and proteins; root activity) indicators, clarifying the role of SlWRKY80 as a positive regulatory factor in tomato defense against M. incognita. Based on this phenomenon, a preliminary exploration of its metabolic signals revealed that SlWRKY80 stimulates different degrees of signaling, such as salicylic acid (SA), jasmonic acid (JA), and ethylene (ETH), and may synergistically regulate reactive oxygen species (ROS) accumulation and scavenging enzyme activity, hindering the formation of feeding sites and ultimately leading to the reduction of root gall growth. To our knowledge, SlWRKY80 has an extremely high utilization value for improving tomato resistance to root-knot nematodes and breeding.

Screening, identification of root-knot nematode resistance sources using multivariate analysis and validation of molecular markers linked to Me genes in chilli (Capsicum annuum L.)

Screening, identification of root-knot nematode resistance sources using multivariate analysis and validation of molecular markers linked to Me genes in chilli (Capsicum annuum L.)

Perennial cultivation of green pepper provides pest suppression, ecosystem development and high crop yield

Perennial cultivation (PC) provides economic benefits to farmers and environmental benefits to society such as soil protection. However, little is known about the effects of PC on soil pest occurrence. This 4-year study used overwintered green pepper (GP) and root-knot nematodes (RKN) as a model plant–soil pest system. We compared the levels of RKN, its antagonists, and other organisms in strip-shaped PC microplots to those of annual cultivation (AC) and no-tillage AC (NT) microplots. To promote RKN occurrence, we inoculated the ends of all microplots with RKN in the spring of year 3 of study. During autumn in years 3 and 4, we determined abundance of RKN and antagonistic animals, DNA operational taxonomical units (OTUs) counts of antagonistic microbes. The results showed that the rate of RKN occurrence was lower in PC and NT than in AC due to minimal soil movement, rather than to suppression by antagonists, although some antagonists such as collembolan and nematode-trapping fungi increased in PC. Moreover, plant biomass was higher in PC than in NT and AC during fruiting period due to persistence of the plant body over years.We also found signs of ecosystem development in PC, which is an essential background for pest suppression: the maturity index (a measure of disturbance) of the nematode community, fungal diversity, frequency of plant and animal symbionts/parasites were greater in PC than in NT or AC. These findings suggested that ecosystem development had commenced within the 4-year study period. We conclude that the long-lasting plant body and undisturbed soil environment are specific to PC, and could provide agronomical and ecological benefits.

Field study on the effect of different planting periods on root-knot nematodes, Meloidogyne spp. infecting tomato

Field study on the effect of different planting periods on root-knot nematodes, Meloidogyne spp. infecting tomato

Bio-control efficacy of selected indigenous nematophagous fungi against Meloidogyne enterolobii in vitro and on dry bean (Phaseolus vulgaris L.).

Dry bean (Phaseolus vulgaris L.) is an important commercialized field crop in South Africa for aiding in food security as a cheap protein source. However, it is highly susceptible to root-knot nematodes (RKN), Meloidogyne species. Use of indigenous nematophagous fungi as bio-control agents (BCA) of Meloidogyne nematodes is a promising research focus area. This is because indigenous fungal species are naturally part of the ecosystem and therefore compatible with other biological processes unlike most synthetic chemicals. The objective of the study was to identify indigenous nematophagous fungal BCA and establish their potential efficacy in reducing M. enterolobii population densities on dry bean with and without incorporation of compost. Screened indigenous fungal species included Aspergillus terreus, Talaromyces minioluteus, T. sayulitensis, Trichoderma ghanense, and T. viride. There were observed significant parasitism differences (P ≤ 0.05) among the BCA, with T. ghanense showing the highest egg parasitism (86%), followed by T. minioluteus (72%) and T. sayulitensis (70%). On the other hand, the highest J2 parasitism was observed on T. minioluteus (95%), followed by A. terreus and T. viride (63%). A similar trend was observed under in vivo conditions, with higher efficacy with compost incorporation. This provides a highly encouraging alternative and ecologically complementary Meloidogyne management in dry bean production.

Protective role of native root-associated bacterial consortium against root-knot nematode infection in susceptible plants

Root-knot nematodes (RKNs) are a global menace to agricultural crop production. The role of root-associated microbes (RAMs) in plant protection against RKN infection remains unclear. Here we observe that cucumber (highly susceptible to Meloidogyne incognita) exhibits a consistently lower susceptibility to M. incognita in the presence of native RAMs in three distinct soils. Nematode infection alters the assembly of bacterial RAMs along the life cycle of M. incognita. Particularly, the loss of bacterial diversity of RAMs exacerbates plant susceptibility to M. incognita. A diverse range of native bacterial strains isolated from M. incognita-infected roots has nematode-antagonistic activity. Increasing the number of native bacterial strains causes decreasing nematode infection, which is lowest when six or more bacterial strains are present. Multiple simplified synthetic communities consisting of six bacterial strains show pronounced inhibitory effects on M. incognita infection in plants. These inhibitory effects are underpinned via multiple mechanisms including direct inhibition of infection, secretion of anti-nematode substances, and regulation of plant defense responses. This study highlights the role of native bacterial RAMs in plant resistance against RKNs and provides a useful insight into the development of a sustainable way to protect susceptible plants.

Host status of cover crops for the management of Meloidogyne izalcoensis in coffee orchards infested with root-knot nematodes

Summary The root-knot nematode Meloidogyne izalcoensis was described parasitising coffee trees and causing damage in the Izalco volcano region, Sonsonate, El Salvador. In Brazil, it was detected in coffee roots in the municipality of Indianópolis, Minas Gerais state (MG). Intercropping cover crops is a traditional practice in Brazilian coffee orchards, and the use of plants that are non-hosts or antagonistic to the nematodes is a very promising strategy for coffee in infested areas. Due to the scarce information on the host status of M. izalcoensis, the aim of this study was to evaluate 24 plants of different species of cover crops and select the non-hosts or poor hosts. Two assays were conducted at different times, and all plants were inoculated with 10 000 eggs. After 90 days, the following parameters were determined: fresh root weight, reproduction factor (RF) and final reaction. Plants with RF < 1.0 were classified as non-hosts (NH), those with RF > 1.0 were classified as hosts and using statistical analyses as: good host (GH), intermediate host (IH) and poor host (PH). Most of the botanical species tested were classified as NH or PH to M. izalcoensis: cotton, rice, white oat, black oat, ryegrass, corn, three species of grass, two cultivars of millet, two wheat cultivars, Crotalaria breviflora, C. ochroleuca, C. spectabilis, jack bean, gray mucuna bean and two species of rice grass. Only tomato and bean were classified as GH, and C. juncea and soybean were classified as IH. The plants identified as non-hosts or poor hosts can be recommended for intercropping or for crop rotation in Brazilian coffee orchards in regions infested by M. izalcoensis.

Efficient control of root-knot nematodes by expressing Bt nematicidal proteins in root leucoplasts

Root-knot nematodes (RKNs) are plant pests that infect the roots of host plants. Bacillus thuringiensis (Bt) nematicidal proteins exhibited toxicity to nematodes. However, the application of nematicidal proteins for plant protection is hampered by the lack of effective delivery systems in transgenic plants. In this study, we discovered the accumulation of leucoplasts (root plastids) in galls and RKN-induced giant cells. RKN infection causes the degradation of leucoplasts into small vesicle-like structures, which are responsible for delivering proteins to RKNs, as observed through confocal microscopy and immunoelectron microscopy. We showed that different-sized proteins from leucoplasts could be taken up by Meloidogyne incognita female. To further explore the potential applications of leucoplasts, we introduced the Bt crystal protein Cry5Ba2 into tobacco and tomato leucoplasts by fusing it with a transit peptide. The transgenic plants showed significant resistance to RKNs. Intriguingly, RKN females preferentially took up Cry5Ba2 protein when delivered through plastids rather than the cytosol. The decrease in progeny was positively correlated with the delivery efficiency of the nematicidal protein. In conclusion, this study offers new insights into the feeding behavior of RKNs and their ability to ingest leucoplast proteins, and demonstrates that root leucoplasts can be used for delivering nematicidal proteins, thereby offering a promising approach for nematode control.

Optimization andutilization ofemerging waste (fly ash) for growth performance of chickpea (Cicer arietinum L.) plant and mitigation of root-knot nematode (Meloidogyne incognita) stress.

The sustainable management of large amounts of fly ash (FA) is a concern for researchers, and we aim to determine the FA application in plant development and nematicidal activity in the current study. A pot study is therefore performed to assess the effects of adding different, FA-concentrations to soil (w/w) on the infection of chickpea plants with the root-knot nematode Meloidogyne incognita. Sequence characteristic amplified region (SCAR) and internal transcribed spacer (ITS) region-based-markers were used to molecularly confirm M. incognita. With better plant growth and chickpea yield performance, FA enhanced the nutritious components of the soil. When compared with untreated, uninoculated control (UUC) plants, the inoculation of M. incognita dramatically reduced chickpea plant growth, yield biomass, and metabolism. The findings showed that the potential of FA to lessen the root-knot nematode illness in respect of galls, egg-masses, and reproductive attributes may be used to explain the mitigating effect of FA. Fascinatingly, compared with the untreated, inoculated control (UIC) plants, the FA treatment, primarily at 20%, considerably (p ≤ 0.05) boosted plant growth, yield biomass, and pigment content. Additionally, when the amounts of FA rose, the activity of antioxidants like superoxide dismutase-SOD, catalase-CAT, and peroxidase-POX as well as osmo-protectants like proline gradually increased. Therefore, our findings imply that 20% FA can be successfully applied as a potential strategy to increase biomass yield and plant growth while simultaneously reducing M. incognita infection in chickpea plants.

Effect of Temperature on the Embryogenesis of Three Geographically Distinct Populations of Meloidogyne incognita Is Driven by Intrinsic Thermal Acclimation Reaction.

Research interest in the mechanisms enabling plant-parasitic nematodes to adjust their physiological performance and cope with changing temperatures has intensified in light of global warming. Here, we show that geographically distinct populations of the root-knot nematode Meloidogyne incognita, which is prevalent in the three main pepper-growing regions in Israel-Carmel Valley (Carmel), Jordan Valley (JV), and Arava Rift (Arava)-possess persistent differences in their thermal acclimation capacity, which affect pre- and postembryonic development. The optimal temperature for embryonic growth completion was 25°C for the Carmel population; 25 and 30°C for the JV population; and 30°C for the Arava population. Cumulative hatching percentages showed variations among populations; relative to hatching at 25°C, the Carmel population experienced hatching reduction at the higher studied temperatures 30 and 33°C, while the JV and Arava populations exhibited an increase in hatching at 30 and 33°C, respectively. Juvenile survival indicates that at the lowest temperature (20°C), the Carmel population gained the highest survival rates throughout the experimental duration, while at the same duration at 33°C, the Arava population gained the highest survival rate. Infective juveniles of the Carmel population demonstrated increased penetration of tomato roots at 25°C compared to the JV and Arava populations. Inversely, at 33°C, increased penetration was observed for the Arava compared to the Carmel and JV populations. Altogether, the Arava population's performance at 33°C might incur distinct fitness costs, resulting in consistent attenuation compared to the Carmel population at 25°C. Precisely defining a population's thermal acclimation response might provide essential information for models that predict the impact of future climate change on these populations.

Optimizing sustainable control of Meloidogyne javanica in tomato plants through gamma radiation-induced mutants of Trichoderma harzianum and Bacillus velezensis

This study investigates the efficacy of Trichoderma spp. and Bacillus spp., as well as their gamma radiation-induced mutants, as potential biological control agents against Meloidogyne javanica (Mj) in tomato plants. The research encompasses in vitro assays, greenhouse trials, and molecular identification methodologies to comprehensively evaluate the biocontrol potential of these agents. In vitro assessments reveal significant nematicidal activity, with Bacillus spp. demonstrating notable effectiveness in inhibiting nematode egg hatching (16–45%) and inducing second-stage juvenile (J2) mortality (30–46%). Greenhouse trials further confirm the efficacy of mutant isolates, particularly when combined with chitosan, in reducing nematode-induced damage to tomato plants. The combination of mutant isolates with chitosan reduces the reproduction factor (RF) of root-knot nematodes by 94%. By optimizing soil infection conditions with nematodes and modifying the application of the effective compound, the RF of nematodes decreases by 65–76%. Molecular identification identifies B. velezensis and T. harzianum as promising candidates, exhibiting significant nematicidal activity. Overall, the study underscores the potential of combined biocontrol approaches for nematode management in agricultural settings. However, further research is essential to evaluate practical applications and long-term efficacy. These findings contribute to the development of sustainable alternatives to chemical nematicides, with potential implications for agricultural practices and crop protection strategies.

Building accelerated plant breeding pipelines: Screening to evaluate lima bean resistance to root-knot nematode in diverse inbred lines and segregating breeding populations.

Lima beans (Phaseolus lunatus) are a cornerstone crop of Delaware's processing vegetable industry. Root-knot nematodes (RKN; Meloidogyne spp.) cause galling of root systems which severely reduces yield. Durable host resistance is an effective management strategy for RKN, but availability of resistant lima bean cultivars is limited. To overcome these challenges, breeding pipelines must simultaneously advance pre-commercial lines and identify new resistance sources with potential for incorporation into the breeding program. Inoculated field trials were conducted in 2021-2022 to evaluate three M. incognita resistant, pre-commercial experimental lines for resistance traits and yield potential in comparison to commercial standards, 'Cypress' and 'C-elite Select'. DE1306635 had the highest yield and reduced galling and reproduction compared to 'Cypress', and is a candidate for commercial release. To continue the breeding pipeline, 256 lima bean inbred accessions from around the world were assessed from 2022-2023 in greenhouse and field screenings to identify novel sources of resistance in the lima bean gene pool. This method allows for evaluation and/or advancement of three generations per year. The full panel was initially evaluated for root galling and 60 accessions were selected for additional field and greenhouse screening: 25 large- and 25 small-seeded with the lowest gall ratings, 5 high-gall controls, and 5 commercial standards. Seven accessions with reduced M. incognita galling and reproduction were identified, including two known resistant lines and five newly identified genotypes. The resistance carried by these genotypes will be further characterized to assess their potential use in lima bean RKN resistance breeding.

Metabolomics and histopathological analysis of two tomato cultivars after co-infection with soil-borne pathogens (Southern root-knot nematode and Fusarium wilt fungus)

Southern root-knot nematode (Meloidogyne incognita) and Fusarium wilt fungus (Fusarium oxysporum) are one of the most predominant pathogens responsible for substantial agricultural yield reduction of tomato. The current study planned to assess the effects of M. incognita (Mi) and F. oxysporum (Fo) and their co-infection on two tomato cultivars, Zhongza 09 (ZZ09) and Gailing Maofen 802 (GLM802). The present study examined the effects of co-infection on leaf morphology, chlorophyll content, leaf area, and histopathology. The present study used metabolomics to evaluate plant-pathogen interactions. The outcomes of the current study revealed that chlorophyll content and leaf area decreased more in GLM802 during co-infection. In co-infection (Fo + Mi), the chlorophyll content reduction in ZZ09 was 11%, while in GLM802 the reduction reached up to 31% as compared to control. Moreover, the reduction in leaf are in ZZ09 was 31%, however, in the GLM802 reduction was observed 54% as compared to control plants. Similarly, GLM802 stems exhibited larger brown patches on their vascular bundles than ZZ09 stems. The rate of browning of GLM802 stems was 247% more than ZZ09, during co-infection. Moreover, GLM802 roots exhibited a higher abundance of hyphae and larger galls than ZZ09 roots. In metabolic studies, glutathione, succinic acid, and 2-isopropylmalic acid decreased, whereas spermine and fumaric acid increased in GLM802 co-infected stems. It indicates that GLM802 is weakly resistant; therefore, F. oxysporum and other pathogens readily damage tissue. In the co-infected stem of ZZ09, L-asparagine and shikimic acid increased, but pipecolic acid, L-saccharine, and 2-isopropylmalic acid declined. L-asparagine was crucial in preserving the stability of nitrogen metabolism, chlorophyll synthesis, and leaf growth in ZZ09. Shikimic acid's substantial accumulation could explain the limited extent of browning observed in the vascular bundles of ZZ09. Thus, the present study provides insight into M. incognita and F. oxysporum co-infection in two tomato cultivars, which may aid breeding efforts to generate commercially viable resistant cultivars. However, further research on the relationship between M. incognita and F. oxysporum in different host plants is required in the future.

THE POTENTIAL OF TEQUIL, A PLANT EXTRACT BASED ON QUILLAJA SAPONARIA, IN SUSTAINABLE AGRICULTURE

More attention is being given to researches of eco-sustainable solutions to be implemented in agriculture, particularly in the presence of plant parasitic nematodes, whose damages have significant global importance. Unsustainable practices, such as fumigation and monoculture, have led to depletion of biodiversity, allowing harmful trophic groups (bacteria, fungi, and nematodes) to prevail, and their negative synergistic action further exacerbating yield losses. Considering the notable and progressive restriction of usable molecules, nematode control strategies must necessarily consider the appropriate integration of methods. The main objective of control is to limit pathogen populations below the damage threshold, rather than aiming at their eradication as mistakenly pursued in the past. This goal can be achieved with the help of solutions that promote plant development through biostimulation of root systems, which are often affected by various organisms in the biosphere. Tequil, through its well-documented synergy of saponins, tannins, and polyphenols, produces invigorating, biostimulating, and strengthening effects on the root system. Indeed, in vitro tests and field trials have highlighted Tequil’s appreciable activity in containing trophic groups, particularly root-knot nematodes of the Meloidogyne genus. Therefore, its use could indirectly contribute to nematode control and limit yield losses. Key Words: Quillaja saponaria, nematodes, bio stimulation, biocontrol, abiotic stresses resistance.

Distinct changes in tomato-associated multi-kingdom microbiomes during Meloidogyne incognita parasitism

BackgroundThe interplay between root-knot nematode (RKN) parasitism and the complex web of host-associated microbiota has been recognized as pivotal for effective management of the pest. However, studies assessing this relationship have focussed on the bacterial and fungal communities, neglecting the unicellular eukaryotic members. Here, we employed amplicon sequencing analysis of the bacterial 16S rRNA, fungal ITS and eukaryotic 18S rRNA genes, and comprehensively examined how the microbiome composition, diversity and networking developed with time in the rhizospheres and roots of RKN-inoculated and non-inoculated tomato plants.ResultsAs expected, infection with the RKN Meloidogyne incognita decreased plant growth. At individual timepoints, we found distinct bacterial, fungal and eukaryote community structures in the RKN-inoculated and non-inoculated rhizospheres and roots, and RKN inoculation affected several taxa in the root-associated microbiome differentially. Correlation analysis revealed several bacterial and fungal and few protist taxa that correlated negatively or positively with M. incognita. Moreover, network analysis using bacterial, fungal and eukaryotic data revealed more dynamic networks with higher robustness to disturbances in the RKN-inoculated than in the non-inoculated rhizospheres/roots. Hub taxa displayed a noticeable successional pattern that coincided with different phases of M. incognita parasitism. We found that fungal hubs had strong negative correlations with bacteria and eukaryotes, while positive correlations characterized hub members within individual kingdoms.ConclusionOur results reveal dynamic tomato-associated microbiomes that develop along different trajectories in plants suffering M. incognita infestation and non-infested plants. Overall, the results identify stronger associations between RKN and bacterial and fungal taxa than between eukaryotic taxa and RKN, suggesting that fungal and bacterial communities could play a larger role in the regulation of RKN. The study identifies several putative RKN-antagonistic bacterial and fungal taxa and confirms the antagonistic potential previously identified in other taxa.

Seasonal Fluctuations in Plant-Parasitic Nematode Vertical Distributions and Their Interactions with Edaphic Factors in Vegetable Fields of South Georgia, U.S.A.

Plant-parasitic nematodes (PPNs) can be found deep in the soil profile, compounding nematode management decisions and detection. This study aimed to understand how seasonal fluctuations in edaphic factors are associated with the vertical distribution of PPNs in south Georgia's vegetable cropping systems. Five-core composite soil samples were taken monthly (March 2020 to February 2022) at three random locations in six vegetable fields. Fields represented three cropping systems (vegetable plasticulture, bare ground cucumber, and plastic bed watermelon rotation) and two regions (north and south) sampled from five 15-cm strata (0 to 15, 15 to 30, 30 to 45, 45 to 60, and 60 to 75 cm). Only soil temperatures and precipitation had seasonal fluctuations; the other edaphic factors were vertically stratified. Latitude and stratum had the strongest associations with nematode composition, showing a clear separation between the north and south regions. Variations in soil texture, porosity, moisture, and PPN vertical distribution suggested that the illuvial zone in the north region was the main factor in the differences observed between regions and strata. Seasonal fluctuations in vertical distribution were found among Meloidogyne incognita and Nanidorus spp. Higher abundances of deep-dwelling PPNs were limited to the south region in the winter for deep-dwelling M. incognita and summer and winter for Nanidorus spp., probably due to the illuvial zone barrier in the north region. Because most root-knot nematodes dwell at ≤30 cm during the summer, fumigant/nematicide applications are recommended during this season to minimize the risk of deep-dwelling escapees.

Nematicidal Activity of Volatiles against the Rice Root-Knot Nematode and Environmental Safety in Comparison to Traditional Nematicides.

The rice root-knot nematode (RRKN), Meloidogyne graminicola Golden and Birchfield 1965, is a dangerous crop pest that affects rice production on a global scale. The largest rice-producing countries struggle with the impacts of RRKN infestation, namely, underdeveloped plants and a reduction in rice grain that can reach up to 70% of crop yield. In addition, the shift to strategies of sustainable pest management is leading to a withdrawal of some of the most effective pesticides, given the dangers they pose to the environment and human health. Volatile metabolites produced by plants can offer safer alternatives. The present study characterized the nematicidal activity of volatile phytochemicals against the RRKN and compared the most active with commercial nematicides concerning their safety to the environment and human health. Rice plants were used to grow large numbers of RRKNs for direct-contact bioassays. Mortality induced by the volatiles was followed for four days on RRKN second-stage juveniles. Of the 18 volatiles tested, carvacrol, eugenol, geraniol, and methyl salicylate showed the highest mortalities (100%) and were compared to traditional nematicides using (eco)toxicological parameters reported on freely available databases. While methyl salicylate had a faster activity, carvacrol had more lasting effects. When compared to synthetic nematicides, these volatile phytochemicals were reported to have higher thresholds of toxicity and beneficial ecotoxicological parameters. Ultimately, finding safer alternatives to traditional pesticides can lower the use of damaging chemicals in farming and leverage the transition to agricultural practices with a lower impact on biodiversity.

Local and systemic transcriptome and spliceome reprogramming induced by the root-knot nematode Meloidogyne incognita in tomato.

Root-knot nematodes (Meloidogyne spp.) are widely spread root parasites that infect thousands of vascular plant species. These highly polyphagous nematodes engage in sophisticated interactions with host plants that results in the formation of knot-like structures known as galls whose ontogeny remains largely unknown. Here, we determined transcriptome changes and alternative splicing variants induced by Megalaima incognita in galls and neighboring root cells at two distinct infective stages. M. incognita induced substantial transcriptome changes in tomato roots both locally in galls and systemically in neighboring cells. A considerable parallel regulation of gene expression in galls and neighboring cells were detected, indicative of effective intercellular communications exemplified by suppression of basal defense responses particularly during the early stage of infection. The transcriptome analysis also revealed that M. incognita exerts a tight control over the cell cycle process as a whole that results in an increase of ploidy levels in the feeding sites and accelerated mitotic activity of the gall cells. Alternative splicing analysis indicated that M. incognita significantly modulates pre-mRNA splicing as a total of 9064 differentially spliced events from 2898 genes were identified where intron retention and exon skipping events were largely suppressed. Furthermore, a number of differentially spliced events were functionally validated using transgenic hairy root system and found to impact gall formation and nematode egg mass production. Together, our data provide unprecedented insights into the transcriptome and spliceome reprogramming induced by M. incognita in tomato with respect to gall ontogeny and nematode parasitism.

Utilizing endophytic plant growth-promoting bacteria and the nematophagous fungus Purpureocillium lilacinum as biocontrol agents against the root-knot nematode (Meloidogyne incognita) on tomato plants

AbstractThe present investigation was designed to assess how administering biocontrol agents (BCAs) made from the nematophagous endophytic fungus MR20 (Purpureocillium lilacinum) or a mixture of endophytic plant growth-promoting bacteria MR12 (Pseudomonas fluorescens), and MR25 (Serratia marcescens), could enhance tomato growth and resistance to Meloidogyne incognita under in vivo conditions. The three strains’ cell-free culture filtrates showed a strong nematocidal impact (P < 0.05) on M. incognita infective second-stage juveniles (J2s). The highest mortality rates by the three BCAs were at concentrations of 80%, followed by 60%, 40%, and 20%. The in vitro hatching of free eggs was found to be considerably (P < 0.05) reduced as the concentrations of the endophytic bacteria MR12, and MR25 were increased in the order 1 × 105, 1 × 106, 1 × 107, 0.5 × 108, to 1 × 108 colony forming units (CFU)/mL, and the maximum nematicidal activity in killing M. incognita free eggs occurred at 1 × 108 CFU/mL. A statistically significant (P < 0.05) reduction in the percentage of M. incognita-free eggs retrieved was seen in vitro when various concentrations of the endophytic fungus MR20 were applied compared to the negative control (distilled water). The endophytic fungus MR20 had the highest nematicidal activity against M. incognita free eggs at a concentration of 3 × 106 CFU/mL. The application of P. lilacinum or a combination of P. fluorescens and S. marcescens to tomato plants in the presence of M. incognita under greenhouse conditions resulted in a significant increase (P < 0.05) in root and shoot fresh weight, number of leaves, weight of leaves, and stem diameter when compared to the positive control treatment that contained only M. incognita. Treatment with P. lilacinum was more effective (P < 0.05) than P. fluorescens and S. marcescens in reducing egg masses per root, J2s per 100 g of soil, egg masses per 100 g of soil, J2s + eggs of M. incognita per 100 g of soil, reproduction factor, and reduction percentage after 60 days under greenhouse conditions. Nevertheless, the reduction of M. incognita gall size categorization (> 4 mm, 2–4 mm, and < 2 mm) was more effectively achieved by treatment with P. fluorescens and S. marcescens than by treatment with P. lilacinum. Under in vivo conditions, the tested P. lilacinum or a mixture of P. fluorescens and S. marcescens effectively controlled nematode population densities below the economic threshold.