Induced resistance by the mutualistic endophyte, Fusarium oxysporum strain 162, toward Meloidogyne incognita on tomato

The non-pathogenic endophytic fungus, Fusarium oxysporum strain 162, originally isolated from the endorhiza of tomato roots, reduces damage caused by Meloidogyne incognita, by inhibiting juvenile penetration of and development in the root. However, little is known about the mode of action of this endophyte fungus against the nematode. This study aimed at investigating how the endophyte affects nematode motility and survival and if induced resistance plays a role in the relationship. In a previous study, F. oxysporum strain 162 decreased nematode penetration of tomato up to 60%. In experiments using a split-root chamber to test for induced resistance, nematode penetration, number of galls, and number of egg masses were investigated 2 and 5 weeks after nematode inoculation. Split-root plants treated with F. oxysporum strain 162 showed 26-45% less nematode penetration, 21-36% less galls and a 22-26% reduction in the number of egg masses in the roots not directly inoculated with the fungus when compared to untreated control plants in repeated tests. In conclusion, inoculation of tomato plants with the non-pathogenic fungal endophyte F. oxysporum strain 162 resulted in a signficant reduction of nematode infection, which was in part due to induced resistance in the first 2-3 weeks after fungal inoculation.     

Induction of metabolite organic compounds by mutualistic endophytic fungi to reduce the greenhouse whitefly Trialeurodes vaporariorum (Westwood) infection on tomato

Background and aims

Six mutualistic endophytic fungi that are known to colonize the endorhiza have shown biological control properties against plant-parasitic nematodes. In this study we aim to investigate the potential of these endophytic fungi to reduce the phloem-feeding insect Trialeurodes vaporariorum (Westwood) on tomato.

Methods

To determine the host plant choice of T. vaporariorum, the total number of insects present on each plant was counted daily for 10 days, and then the second leaf below the shoot apex were examined for its chlorophyll content index (CCI).To separate and quantify the active compounds produced in the tomato leaves, a reversed phase high liquid chromatography (RP-HPLC) analysis was performed.

Principle results

A greenhouse choice test showed that Trichoderma atroviride strain MT-20, T. atroviride strain S-2 and Fusarium oxysporum strain 162 (Fo162) reduced the number of greenhouse whiteflies fifty percent when compared to the untreated control during ten days after insect release. The highest level of biocontrol activity was attained with Fo162. The strains MT20, S-2, and Fo162 all demonstrated acropedal induction of resistance to the insects. The isolate Fusarium sp. strain Bonn-7 enhanced plant growth. The negative effect on insect attraction to the leaves of the endophyte treated plants was not associated with leaf altered chlorophyll content. RP-HPLC analysis revealed that inoculation of the fungus Fo162 induced a change in the accumulation of specific organic compounds in the tomato leaves that could be the cause of insect repellence.

Conclusions

This study demonstrated the high potential of mutualintic endophytic fungi, in particular of Fo162, to induce resistance in tomato against the phloemfeeding T. vaporariorum.     

Monoxenic liquid culture with Escherichia coli of the free-living nematode Panagrolaimus sp. (strain NFS 24-5), a potential live food candidate for marine fish and shrimp larvae

The free-living, bacterial-feeding nematode Panagrolaimus sp. (strain NFS 24-5) has potential for use as live food for marine shrimp and fish larvae. Mass production in liquid culture is a prerequisite for its commercial exploitation. Panagrolaimus sp. was propagated in monoxenic liquid culture on Escherichia coli and parameters, like nematode density, population dynamics and biomass were recorded and compared with life history table data. A mean maximum nematode density of 174,278 mL^?1 and a maximum of 251,000 mL^?1 were recorded on day 17 after inoculation. Highest average biomass was 40 g L^?1 at day 13. The comparison with life history table data indicated that the hypothetical potential of liquid culture is much higher than documented during this investigation. Nematode development is delayed in liquid culture and egg production per female is more than five times lower than reported from life history trait analysis. The latter assessed a nematode generation time of 7.1 days, whereas the process time at maximum nematode density in liquid culture was 16 days indicating that a reduction of the process time can be achieved by further investigating the influence of nematode inoculum density on population development. The results challenge future research to reduce process time and variability and improve population dynamics also during scale-up of the liquid culture process.     

The Influence of Trichoderma harzianum on the Root-knot Fusarium Wilt Complex in Cotton

Wilt-susceptible cultivar ''Rowden'' cotton was inoculated wilh Meloidogyne incognita (N), Trichoderma harzianum (T), and Fusarium oxysporum f. sp. vasinfectum (F) alone and in all combinations in various time sequences. Plants inoculated with F alone or in combination with T did not develop wilt, Simultaneous inoculation of 7-day-old seedlings with all three organisms (NTF) produced earliest wilt. However, plants receiving nematodes at 7 days and Fusarium and Trichoderma at 2 or 4 weeks later (N-T-F, N-TF) developed the greatest wilt between 49-84 days after initial nematode inoculation. During the same period, Fusarium added 4 weeks after initial nematode inoculation (N-F) and Fusarium added 4 weeks after initial simultaneous inoculation of nematode and Trichoderma (NT-F) produced the least wilt. The addition of Fusarium inhibited nematode reproduction. Simultaneous inoculation with nematodes and Trichoderma (NT-) resulted in the greatest root gall development, whereas nematodes alone produced the greatest number of larvae. In comparison with noninoculated controls (CK), treatments involving all three organisms inhibited plant growth, plants inoculated with the nematode alone (N-) or with nematodes and Trichoderma (NT-) simultaneously had greatest root weight. Any treatment involving the nematode resulted in fewer bolls per plant and greater necrosis on roots than the noninoculated checks.     

Identification and evaluation of a potential biocontrol agent,Bacillus subtilis,against Fusarium sp. in apple seedlings

To find a potential biocontrol agent against Fusarium sp. in apple seedlings, an endophytic bacterium strain was isolated from apple tree tissues. The inhibitive efficiency of the isolated strain against the hyphal growth of Fusarium sp. and Rhizoctonia solani was tested. Strain Y-1 showed significant inhibitory effects against Fusarium oxysporum, F. moniliforme, F. proliferatum, F. solani and R. solani. Its antifungal activity against F. oxysporum was the highest, reaching up to 64.90 %. In vivo tests indicated that strain Y-1 effectively protects apple from F. oxysporum infections. The control effect reached 92.26 % when bacterial inoculation was performed 3 days prior to pathogen inoculation. Strain Y-1 could colonize the rhizosphere and tissues within 30 days. It was also able to induce systemic resistance in apple seedlings as shown by the activities of SOD and POD. Strain Y-1 significantly increased the root length, root wet and dry weights, and plant height of the apple seedlings compared with the control group. The homology analysis of the 16S rRNA sequence, together with morphological, physical, and biochemical analyses, revealed that strain Y-1 is Bacillus subtilis.     

Interactions between Zeldia Punctata (Cephalobidae) and bacteria in the presence or absence of maize plants

Bacterial-feeding nematodes constitute one of the primary grazers of soil bacteria. We investigated the effects of selective grazing of a representative nematode (Zeldia punctata, Cephalobidae) on nematode life history and population biology and on the soil microbial community. Firstly, we measured (i) the effect of five different bacterial strains on the nematode life cycle using petri dishes and (ii) the impact of bacterial inoculation on nematode population growth in a soil microcosm. Selection of the five bacterial strains was based on morphology, cell-wall characteristics and mucus production. Z. punctata development was strongly affected by the type of bacteria ingested, independent of experimental design. Bacterial cell-wall characteristics seemed to directly affect Z. punctata development since high nematode densities were only reached with gram-negative strains (Pseudomonas monteilii and Methylobacterium nodulans). In petri dishes, the filamentous organisms (Actinomyces sp.) and mucus-producing bacteria (Bradyrhizobium sp.) led to the least reproduction. Duration of the various nematode life phases (egg, juvenile, reproductive stage and non-reproductive stage) was significantly affected by the bacterial food source. Total life span varied from 12.5 days (Bradyrhizobium sp.) to 40 days (Pseudomonas monteilii). Secondly, we monitored the influence of Z. punctata on the indigenous soil microbial community in the presence or absence of a maize plantlet. Nematode inoculation led to an increase in bacterial activity (as measured by alkaline phosphatase activity) but did not significantly influence bacterial biomass. The genetic fingerprint (DGGE) of soil bacteria was more influenced by plant presence than by nematode inoculation. Nematode activity has important repercussions on N flux in the soil since inoculation of Z. punctata in the absence of plants resulted in a net increase of N mineralization (2 mg N per pot) while a decrease of mineral N (0.5 mg N per pot) was observed in the absence of the nematodes, due to bacterial immobilization. This study underscores the close relationship between selective bacterial grazing and nematode development. Nevertheless, the impact of nematode grazing on the overall soil microbial community seems to primarily affect microbial activity and relative dominance rather than microbial diversity.     

Real time PCR detection targeting nifA gene of plant growth promoting bacteria Azospirillum brasilense strain FP2 in maize roots

The plant growth promoting bacteria (PGPB) Azospirillum brasilense has been recommended for use in commercial inoculants in Brazil. Effective methods are necessary to monitor PGPB strains in the rhizosphere. Our purpose was to develop a real time PCR method for detection of A. brasilense strain FP2 in maize seedlings targeting nifA. Primer pairs were designed and their specificity was verified using DNA from 12 different bacterial species. Standard curves were prepared for DNA quantification using serial dilution of A. brasilense DNA extracts. PCR efficiencies and correlation coefficient presented values within the acceptable range for qPCR, mean PCR efficiency was 95 % and correlation coefficient was 0.98, indicating that nifA gene was suitable for the quantitative analysis of the target bacterial genome. Inoculated maize seedlings were grown in vitro or in pots, bacterial DNA copy number per gram of fresh root was quantified 1, 4, 7 and 10 days after inoculation. The developed primers targeting nifA will be useful for monitoring Azospirillum brasilense strain FP2 in crops.     

Developmental and behavioural effects of the endophytic Fusarium moniliforme Fe14 towards Meloidogyne graminicola in rice

The root‐knot nematode, Meloidogyne graminicola, is an important pest of rice in many rice production areas worldwide. The endophyte Fusarium moniliforme strain Fe14, isolated from a disinfected root of rice, has previously shown potential antagonistic activity against M. graminicola. This study shows the effects of Fe14 on M. graminicola behaviour, infection, development and reproduction. The endophyte Fe14 colonisation significantly reduced M. graminicola penetration into rice roots by 55% and increased the male to female ratio nine times. The endophyte also delayed juvenile development into female inside the rice root. These results suggest a suboptimal performance of the giant cell and a cumulative effect of the endophyte on the long‐term root‐knot nematode population development. In split‐root assays, the application of Fe14 at the inducer side significantly reduced nematode invasion at the responder side by 38% and 60% in two independent trials. This result suggests a systemic effect of the endophyte on rice plants. The root exudates from Fe14‐treated plants were either less attractive or had repellent effect on nematode movement. The results, when compared to what was described for other endophytic Fusarium against other nematode species, may indicate a basal response mechanism initiated in the plant by endophytic Fusarium spp. The present study may give leads for unravelling the molecular mechanisms responsible for the induced systemic defence responses in plants.     

Dynamics of Belonolaimus longicaudatus Parasitism on a Susceptible St. Augustinegrass Host

St. Augustinegrass (Stenotaphrum secundatum) cv FX-313 was used as a model laboratory host for monitoring population growth of the sting nematode, Belonolaimus longicaudatus, and for quantifying the effects of sting nematode parasitism on host performance in two samples of autoclaved native Margate fine sand with contrasting amounts of organic matter (OM = 7.9% and 3.8%). Following inoculation with 50 Belonolaimus longicaudatus per pot, nematodes peaked at a mean of 2,139 nematodes per pot 84 days after inoculation, remained stable through 168 days at 2,064 nematodes per pot, and declined at 210 days. The relative numbers of juveniles and adults demonstrated senescence after 84 days. Root dry weight of nematode-inoculated plants increased briefly to an apparent equilibrium 84 days after inoculation, whereas root weights of uninoculated controls continued to increase, exceeding those of inoculated plants from 84 to 210 days (P < 0.01). At 210 days, uninoculated plants had 227% the root dry weight of inoculated plants. Transpiration of FX-313 was reduced by nematodes (P < 0.0001) at 84 and 126 days after inoculation; reduction was first observed at 42 days and last observed 168 days after inoculation (P < 0.05). OM content affected all plant performance variables at multiple dates, and generally there were no inoculation x OM content interactions. OM content had no effect on nematode numbers per pot, although there was a slight (P < 0.05) increase in the number of nematodes per gram root dry weight in the low-OM soil compared with the high-OM soil.     

Dual inoculation of Fusarium oxysporum endophytes in banana: effect on plant colonization,growth and control of the root burrowing nematode and the banana weevil

The burrowing nematode (Radopholus similis (Cobb) Thorne) and the banana weevil (Cosmopolites sordidus Germar, Coleoptera: Curculionidae) are major pests of banana (Musa spp.) in the Lake Victoria basin region of Uganda. Among biological options to control the two pests is the use of non-pathogenic Fusarium oxysporum Schltdl.: Fries endophytes of banana. We investigated the ability of endophytic F. oxysporum isolates Emb2.4o and V5w2 to control the banana weevil and the burrowing nematode, alone and in combination. Plant colonization by the endophytes was determined by inoculating their chemical-resistant mutants separately and in combination, onto banana roots. Plant growth promotion was determined by measuring plant height, girth, number of live roots and fresh root weight at harvest, and control of the nematode and weevil was determined by challenging endophyte-inoculated plants with the pests 8 weeks after endophyte inoculation. Endophytic root colonization was highest in plants inoculated with both endophytes, compared with those inoculated with only one of the endophytes. Root colonization was better for isolate V5w2 than Emb2.4o. Dually inoculated plants showed a significant increase in height, girth, fresh root weight and number of functional roots following nematode challenge. Nematode numbers in roots were reduced 12 weeks after challenge of 8-week-old endophyte-inoculated plants. Significant reductions in weevil damage were observed in the rhizome periphery, inner and outer rhizomes, compared with endophyte non-inoculated controls. We conclude that dual inoculation of bananas with endophytic isolates Emb2.4o and V5w2 increases root colonization by the endophytes, reduces nematode numbers and weevil damage, and enhances plant growth in the presence of nematode infestation.     

Penetration and Postinfection Development of Meloidogyne incognita on Cotton as Affected by Glomus intraradices and Phosphorus

The influence of the vesicular-arbuscular mycorrhizal fungus Glomus intraradices (Gi) and superphosphate (P) on penetration, development, and reproduction of Meloidogyne incognita (Mi) was studied on the Mi-susceptible cotton cultivar Stoneville 213 in an environmental chamber at 28 C. Plants were inoculated with Mi eggs at planting or after 28 days and destructively sampled 7, 14, 21, and 28 days after nematode inoculation. Mi penetration after 7 days was similar in all treatments at either inoculation interval. At 28 days, however, nematode numbers were least in mycorrhizal root systems and greatest in root systems grown with supplemental P. The rate of development of second-stage juveniles to ovipositing females was unaffected by Gi or P when Mi was added at planting, but was delayed in mycorrhizal root systems when Mi was added 28 days after planting. Nematode reproduction was lower in mycorrhizal than in nonmycorrhizal root systems at both Mi inoculation intervals. Nematode reproduction was stimulated by P when Mi was added at planting, but was similar to reproduction in the low P nonmycorrhizal treatment when Mi was added 28 days after planting. Eggs per female were increased by P fertility when Mi was added at planting.     

Variability among Populations of Meloidogyne arenaria

Variability in reproduction and pathogenicity of 12 populations of Meloidogyne arenaria race 1 was evaluated on Florunner peanut, Centennial soybean, Rutgers tomato, G70, K326, and Mc944 tobacco, and Carolina Cayenne, Mississippi Nemaheart, and Santanka pepper. Differences among M. arenaria populations in rates of egg production 45 days after inoculation were observed for all cultivars except Santanka pepper. Differences among populations in dry top weights or fresh root weights were recorded on all cultivars. Numbers of nematode eggs produced on Florunner peanut varied from 3,419 to 11,593/g fresh root weight. On resistant tobacco cultivars (G70 and K326), one nematode population produced high numbers of eggs (12,042 and 6,499/g fresh root weight on G70 and K326, respectively), whereas the other populations produced low numbers of eggs (less than 500 eggs/g fresh root weight on both cultivars). Two variant M. arenaria race 1 populations were identified by factor analysis of reproductive rates on all nine cultivars. Differences m reproduction and pathogenicity observed among populations would affect the design of sustainable management systems for M. arenaria.     

Isolation and Characterization of a Rhizobacterial Antagonist of Root-Knot Nematodes

The rhizobacterial strain Jdm2 was isolated from the rhizosphere of the traditional Chinese medicinal herb Trichosanthes kirilowii in Jiangsu province, China, and was identified as Bacillus subtilis. Exposure of cell-free filtrate of the strain to the root-knot nematode Meloidogyne incognita under in vitro conditions caused substantial mortality of the second stage juvenile (J2) and significantly reduced egg hatchability. A greenhouse trial demonstrated that 56 days after treatment with Jdm2, the number of galls associated with M. incognita infection in the tomato (Solanum lycopersicum) roots was significantly reduced compared to controls, and the disease severity of infected plants was lower in treated plants (36%) compared to water control (75%). Consistently, in the field trial, the biocontrol efficacy of Jdm2 reached 69%, 51% and 48% after 30, 60 and 90 days post-transplantation, respectively. As indicated by PCR-DGGE analysis, inoculation with Jdm2 strain had an effect on the bacterial community of the tomato rhizosphere at the first stage, but was not able to imperil the bacterial community stability for long time. The novel bacterial strain Jdm2 enhances plant growth and inhibits nematode activity, and has the potential to be a safe and effective microbial pesticide.     

Glyceollin I in soybean-cyst nematode interactions : spatial and temporal distribution in roots of resistant and susceptible soybeans

Accumulation of the phytoalexin glyceollin I in roots of soybean (Glycine max [L.] Merr.) following inoculation with race 1 of Heterodera glycines Ichinohe, the soybean cyst nematode (SCN), was determined in a whole-root system by high performance liquid chromatography (HPLC) and in a cross-section system by a radioimmunoassay procedure. In the whole-root system, roots were harvested from controls and nematode-inoculated seedlings immediately after inoculation and at 2-day intervals for 8 days. The roots were extracted with ethanol, and the extracts were subjected to HPLC. Glyceollin I was not detected in roots of either resistant cultivar Centennial or susceptible cultivar Ransom immediately after inoculation with SCN but steadily accumulated in large quantity in roots of Centennial. Accumulation of glyceollin I in roots of Ransom following nematode inoculation was minimal. In the cross-section system, 3-day-old soybean seedlings were inoculated with juvenile nematodes, and root segments containing a single nematode were dissected from inoculated plants at 4-hour intervals under a dissecting microscope. The root segments were embedded in ice and cut into 16-micrometer sections with a cryostat microtome. The spatial and temporal distribution of glyceollin I was determined with a radioimmunoassay procedure specific for the phytoalexin. Glyceollin I was found to accumulate in tissues immediately adjacent to the head region of the nematode in Centennial but not in Ransom. Glyceollin I was detected 8 hours after nematode penetration, and the concentration increased steadily up to 0.3 micromole per milliliter in Centennial 24 hours after penetration.     

Effect of Soybean Tip Removal on Penetration and Development of Heterodera glycines

On a few occasions, soybeans with broken root tips were included in tests to evaluate resistance to Heterodera glycines. Although females developed on these plants, the numbers tended to be lower than on similarly treated intact roots. To test the possibility that removal of the root meristem affected nematode development, a culture system using pruned soybeans was devised that permitted access to the roots without disturbing the plants. Treatments included removal of 2 mm of root tip at various times ranging from 24 hours before to 10 days after inoculation, or roots left intact. In each experiment, all roots were inoculated at the same time with equal numbers of freshly hatched second-stage juveniles of Heterodera glycines. No differences in nematode development were detected in plants with root tips removed after inoculation compared to the control. When tips were removed at or before inoculation, fewer juveniles entered roots and relatively fewer nematodes developed. Penetration levels and development correlated with root tip removal such that progressively fewer nematodes entered roots and relatively greater numbers of nematodes remained undeveloped as the time interval between root tip removal and inoculation was increased.     

Real-Time PCR Quantification of the Plant Growth Promoting Bacteria Herbaspirillum seropedicae Strain SmR1 in Maize Roots

The plant growth promoting bacteria Herbaspirillum seropedicae SmR1 is an endophytic diazotroph found in several economically important crops. Considering that methods to monitor the plant–bacteria interaction are required, our objective was to develop a real-time PCR method for quantification of PGPB H. seropedicae in the rhizosphere of maize seedlings. Primer pairs were designed, and their specificity was verified using DNA from 12 different bacterial species. Ten standard curves of qPCR assay using HERBAS1 primers and tenfold serial dilutions of H. seropedicae SmR1 DNA were performed, and PCR efficiency of 91 % and correlation coefficient of 0.99 were obtained. H. seropedicae SmR1 limit of detection was 10¹ copies (corresponding to 60.3 fg of bacterial DNA). qPCR assay using HERBAS1 was used to detect and quantify H. seropedicae strain SmR1 in inoculated maize roots, cultivated in vitro and in pots, harvested 1, 4, 7, and 10 days after inoculation. The estimated bacterial DNA copy number per gram of root was in the range 10⁷–10⁹ for plants grown in vitro and it was around 10⁶ for plants grown in pots. Primer pair HERBAS1 was able to quantify H. seropedicae SmR1, and this assay can be useful for monitoring plant–bacteria interaction.     

Biology and Influence of Pterotylenchus cecidogenus on Desmodium ovalifolium

Pterotylenchus cecidogenus did not inhibit germination of Desmodium ovalifolium but did reduce survival of seedlings and root and stem growth. Stem gall formation was associated with nematode population densities which were positively correlated with plant age at time of inoculation. Nematode population densities increased 100 times in 52 days following inoculation.     

Interaction of Vesicular-arbuscular Mycorrhizal Fungi and Phosphorus with Meloidogyne incognita on Tomato

The influence of two vesicular-arbuscular mycorrhizal fungi and phosphorus (P) nutrition on penetration, development, and reproduction by Meloidogyne incognita on Walter tomato was studied in the greenhouse. Inoculation with either Gigaspora margarita or Glomus mosseae 2 wk prior to nematode inoculation did not alter infection by M. incognita compared with nonmycorrhizal plants, regardless of soil P level (either 3 μg [low P] or 30 μg [high P] available P/g soil). At a given soil P level, nematode penetration and reproduction did not differ in mycorrhizal and nonmycorrhizal plants. However, plants grown in high P soil had greater root weights, increased nematode penetration and egg production per plant, and decreased colonization by mycorrhizal fungi, compared with plants grown in low P soil. The number of eggs per female nematode on mycorrhizal and nonmycorrhizal plants was not influenced by P treatment. Tomato plants with split root systems grown in double-compartment containers which had either low P soil in both sides or high P in one side and low P in the other, were inoculated at transplanting with G. margarita and 2 wk later one-half of the split root system of each plant was inoculated with M. incognita larvae. Although the mycoorhizal fungus increased the inorganic P content of the root to a level comparable to that in plants grown in high P soil, nematode penetration and reproduction were not altered. In a third series of experiments, the rate of nematode development was not influenced by either the presence of G. margarita or high soil P, compared with control plants grown in low P soil. These data indicate that supplemental P (30 μ/g soil) alters root-knot nematode infection of tomato more than G. mosseae and G. margarita.     

Histopathology of Okra and Ridgeseed Spurge Infected with Meloidodera charis

Histological observations of okra Abelomoschus esculentus ''Clemson Spineless'' and ridgeseed spurge Euphorbia glyptosperma (a common weed) infected with Meloidodera charis Hopper, indicated that the juvenile nematode penetrated the roots intercellularly. Within 5 days after plant emergence the nematode positioned its body in the cortical tissue parallel to the vascular system. By 10 days after plant emergence the juvenile had extended its head into the vascular system and initiated giant cell formation, generally in protophloem tissue. Giant cells were one celled and usually multi-nucleate. Eggs were observed in the female body 30 days after plants emerged and juveniles were found within the female body by 40 days. Nematode development progressed equally in the root system of either host plant. Generally, throughout the nematode''s life cycle its entire body remained inside the cortical tissue of okra. In ridgeseed spurge, however, the posterior portion of the female erupted through the host epidermis as early as 15 days after plant emergence; only the head and neck remained embedded in the host. The nematode caused extensive tissue disruption in the cortical and vascular system of both plant species. Corn, Zea mays, was another host of the nematode.     

Interaction between Meloidogyne incognita and Rhizoctonia solani on green beans

The interaction between Meloidogyne incognita (race 2) and Rhizoctonia solani (AG 4) in a root rot disease complex of green beans (Phaseolus vulgaris) was examined in a greenhouse pot experiment. Three week-old seedlings (cv. Contender) were inoculated with the nematode and/or the fungus in different combinations and sequences. Two months after last nematode inoculation, the test was terminated and data were recorded. The synchronized inoculation by both pathogens (N + F) increased the index of Rhizoctonia root rot and the number of root galls; and suppressed plant growth, compared to controls. However, the severity of root rot and suppression of plant growth were greater and more evident when inoculation by the nematode preceded the fungus (N → F) by two weeks. Nematode reproduction (eggs/g root) was adversely affected by the presence of the fungus except by the synchronized inoculation. When inoculation by nematode preceded the fungus, plant growth was severely suppressed and roots were highly damaged and rotted leading to a decrease of root galls and eggs.