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The study assessed Trichoderma spp. as a biocontrol agent for managing wilt-nematode complex in tomato crops, aiming to mitigate yield losses. Fusarium sp. and Meloidogyne sp. were isolated from infected plant samples and confirmed molecularly and morphologically as Fusarium oxysporum f.sp. lycopersici and Meloidogyne incognita. Four Trichoderma spp. procured from the Department of Plant Pathology, Tamil Nadu Agricultural University, India were identified molecularly as Trichoderma asperellum (Tv1), Trichoderma asperelloides (Tasd1), Trichoderma harzianum (Th1), and Trichoderma koningiopsis (Tk1) utilizing ITS and TEF1 primer pairs. Among them, Tv1 effectively inhibited the mycelial growth of pathogen isolates. Furthermore, crude metabolite of Tv1 exhibited similar effects. The mortality rate of M. incognita J2s ranged from 90.48% to 100% after 24-72 h of incubation and inhibition percentage of egg hatching reached 90.20%. The shoot length, root length, fresh weight, and dry weight of the tomato plants treated with Tv1 conidia were increased. In a pot experiment, Tv1 treatment reduced disease incidence by 64.57%, comparable to carbendazim treatment (82.32%). Tv1-treated plants had fewer root galls, egg masses, and J2s per 100 g of roots than the inoculated-untreated control plants. The root-knot index (RKI) was significantly lower in plants treated with carbafuran (1.98 ± 0.047) and Tv1 (3.06 ± 0.086) than in control (4.47 ± 0.109). The bio-control efficiency of Tv1 against M. incognita was 21.04%, and the nematodes' reproductive factor (RF) declined to 0.53 in Tv1 treatment group. Based on the findings above, it was established that Tv1 effectively controlled nematode populations and wilt disease when applied in soil., (© 2024 Wiley‐VCH GmbH.) more...

Background: Root-knot nematode is one of the most significant diseases of vegetable crops in the world. Biological control with microbial antagonists has been emerged as a promising and eco-friendly treatment to control pathogens. The aim of this study was to screen and identify novel biocontrol agents against root-knot nematode, Meloidogyne incognita., Results: A total of 890 fungal isolates were obtained from rhizosphere soil of different crops and screened by nematicidal activity assays. Snef1910 strain showed high virulence against second stage juveniles (J2s) of M. incognita and identified as Trichoderma citrinoviride by morphology analysis and biomolecular assay. Furthermore, T. citrinoviride Snef1910 significantly inhibited egg hatching with the hatching inhibition percentages of 90.27, 77.50, and 67.06% at 48, 72, and 96 h after the treatment, respectively. The results of pot experiment showed that the metabolites of T. citrinoviride Snef1910 significantly decreased the number of root galls, J2s, and nematode egg masses and J2s population density in soil and significantly promoted the growth of tomato plants. In the field experiment, the biocontrol application showed that the control efficacy of T. citrinoviride Snef1910 against root-knot nematode was more than 50%. Meanwhile, T. citrinoviride Snef1910 increased the tomato plant biomass., Conclusions: T. citrinoviride strain Snef1910 could be used as a potential biological control agent against root-knot nematode, M. incognita. more...

Plant root chemistry is altered by the parasitism of plant-parasitic nematodes (PPN). Here, we investigated the influence of the infective stage juveniles (J2) of Meloidogyne javanica in inducing tomato ( Solanum lycopersicum ) root volatiles and chemotactic effect on conspecifics. In olfactometer assays, J2 avoided the roots of 2-day infected plants but preferred 7-day-infected tomato compared to healthy plants. Chemical analysis showed a 2-7-fold increase in the amounts of monoterpenes emitted from tomato roots infected with M. javanica relative to healthy roots. In further bioassays, the monoterpenes β-pinene, (+)-(2)-carene, α-phellandrene, and β-phellandrene differentially attracted (51-87%) J2 relative to control. Concurrent reduction and increase in the levels of methyl salicylate and ( Z )-methyl dihydrojasmonate, respectively, in the root volatiles reduced J2 responses. These results demonstrate that the host plant can alter its root volatile composition to inhibit PPN attack. The observed plant-produced inhibition of J2 warrants further investigation as a potential management tool for growers. more...

To date, there has been great demand for ecofriendly nematicides with beneficial properties to the nematode hosting plants. Great efforts are made towards the chemical characterization of botanical extracts exhibiting nematicidal activity against Meloidogyne spp. , but only a small percentage of these data are actually used by the chemical industry in order to develop new formulates. On the other hand, the ready to use farmer produced water extracts based on edible plants could be a sustainable and economic solution for low income countries. Herein, we evaluate the nematicidal potential of Stevia rebaudiana grown in Greece against Meloidogyne incognita and Meloidogyne javanica , two most notorious phytoparasitic nematode species causing great losses in tomato cultivation worldwide. In an effort to recycle the plant's remnants, after leaves selection for commercial use, we use both leaves and wooden stems to test for activity. In vitro tests demonstrate significant paralysis activity of both plant parts' water extracts against the second-stage juvenile (J2) of the parasites; while, in vivo bioassays demonstrated the substantial efficacy of leaves' powder (95% at 1 g kg -1 ) followed by stems. Interestingly, the incorporation of up to 50 g powder/kg of soil is not phytotoxic, which demonstrates the ability to elevate the applied concentration of the nematicidal stevia powder under high inoculum level. Last but not least, the chemical composition analyses using cutting edge analytical methodologies, demonstrated amongst components molecules of already proven nematicidal activity, was exemplified by several flavonoids and essential oil components. Interestingly, and to our knowledge, for the flavonoids, morin and robinin, the anthocyanidin, keracyanin, and a napthalen-2-ol derivative is their first report in Stevia species. more...

Meloidogyne incognita is an economically important plant parasitic nematode. Here we demonstrate substantial variation in the invasiveness of four M. incognita populations relative to tomato. Infective (J2) stage transcriptomes reveal significant variation in the expression of protein-coding and non-coding RNAs between populations. We identify 33 gene expression markers that correlate with invasiveness, and which map to genes with predicted roles in host finding and invasion, including neuropeptides, ion channels, G Protein-Coupled Receptors, cell wall-degrading enzymes and microRNAs. These data demonstrate a surprising diversity in microRNA complements between populations, and identify gene expression markers for invasiveness of M. incognita, to our knowledge for the first time., (Copyright © 2019 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.) more...

A study of the processes of lipid peroxidation and the activity of the peroxidase enzyme, as well as photosynthetic pigments in susceptible tomato plants treated with salicylic acid (SA), during infection with the root-knot nematode Meloidogyneincognita. It was shown that, in the roots of SA-treated plants, the activity of lipid peroxidation is higher compared to the untreated plants, especially in the case of nematode invasion. A significant increase in the activity of lipid peroxidation in SA-treated invasive plants compared with untreated was noted during the transition of larvae to the sedentary stage and the beginning of the formation of feeding areas-giant cells (3-5 days after invasion). This, apparently, contributes to the inhibition of the development of the parasite and the reduction of plant infection and also indicates the involvement of oxidative processes in the mechanism of the induced resistance of plants to root-knot nematodes. In the SA-treated plants, the qualitative and quantitative composition of photosynthetic pigments, disturbed by invasion, was restored and corresponded to the control level. more...

The efficacy of Verticillium leptobactrum isolate (HR1) was evaluated in the control of root-knot nematode and Fusarium wilt fungus under laboratory and greenhouse conditions. Five concentrations of V. leptobactrum (HR1) isolate were tested for their nematicidal and fungicidal activities against Meloidogyne javanica and Fusarium oxysporum f.sp. lycopersici in vitro. Laboratory trials showed that mycelium growth inhibition of Fusarium wilt fungus was correlated to the increase of the concentration of culture filtrate. All dilutions showed efficiency in reducing the growth of Fusarium oxysporum f.sp. lycopersici. The greatest nematicidal activity was observed at 50, 75, and 100% filtrate dilutions. The egg hatching percentage reached 42%, and the juvenile's corrected mortality registered 90% for the above treatments. In greenhouse experiment, the biocontrol agent fungus enhanced significantly tomato growth components (height and weight of plant and root). The multiplication rate of root-knot nematode and the Fusarium wilt disease incidence declined significantly with soil application of V. leptobactrum as with chemical treatments. The isolate HR1 was efficient to control wilt disease complex caused by M. javanica and Fusarium oxysporum f.sp. lycopersici. more...

In the course of screening for microbes with nematicidal activity, we found that Enterobacter asburiae HK169 displayed promising nematicidal activity against the root-knot nematode Meloidogyne incognita , along with plant growth-promoting properties. Soil drenching of a culture of HK169 reduced gall formation by 66% while also increasing root and shoot weights by 251% and 160%, respectively, compared with an untreated control. The cell-free culture filtrate of the HK169 culture killed all juveniles of M. incognita within 48 h. In addition, the nematicidal activity of the culture filtrate was dramatically reduced by a protease inhibitor, suggesting that proteolytic enzymes contribute to the nematicidal activity of HK169. In order to obtain genomic information about the HK169 isolate related to its nematicidal and plant growth-promoting activities, we sequenced and analyzed the whole genome of the HK169 isolate, and the resulting information provided evidence that the HK169 isolate has nematicidal and plant growth-promoting activities. Taken together, these observations enable the future application of E. asburiae HK169 as a biocontrol agent for nematode control and promote our understanding of the beneficial interactions between E. asburiae HK169 and plants. more...

The increased incidence of the crown gall disease caused by Agrobacterium tumefaciens has long been associated with activities of root-knot nematodes (Meloidogyne spp.). Pot experiments on tomato were designed to assess plant vitality, nematode reproduction, and crown gall incidence in combined infection with Agrobacterium and Meloidogyne spp. on tomato roots. Results suggest that tomato plants infected with pathogenic A. tumefaciens 2 days before the nematodes show enhanced plant defense against M. ethiopica resulting in lower egg and gall counts on roots 45 and 90 days postinoculation (dpi); no significantly enhanced defense was observed when the plant was inoculated with bacteria and nematodes at the same time. Split-root experiments also showed that the observed interaction was systemic. Reverse-transcription quantitative polymerase chain reaction analysis that targeted several genes under plant hormonal control suggests that the suppression was mediated via systemic acquired resistance by the pathogenesis-related protein 1 and that M. ethiopica did not enhance the defense reaction of tomato against Agrobacterium spp. Nematodes completely inhibited tumor growth in a 45-day experiment if inoculated onto the roots before the pathogenic bacteria. We conclude that the observed antagonism in the tested pathosystem was the result of initially strong plant defense that was later suppressed by the invading pathogen and pest. more...

Meloidogyne enterolobii is one of the most important plant-parasitic nematodes that can overcome the Mi-1 resistance gene and damage many economically important crops. Translationally controlled tumour protein (TCTP) is a multifunctional protein that exists in various eukaryotes and plays an important role in parasitism. In this study, a novel M. enterolobii TCTP effector, named MeTCTP, was identified and functionally characterized. MeTCTP was specifically expressed within the dorsal gland and was up-regulated during M. enterolobii parasitism. Transient expression of MeTCTP in protoplasts from tomato roots showed that MeTCTP was localized in the cytoplasm of the host cells. Transgenic Arabidopsis thaliana plants overexpressing MeTCTP were more susceptible to M. enterolobii infection than wild-type plants in a dose-dependent manner. By contrast, in planta RNA interference (RNAi) targeting MeTCTP suppressed the expression of MeTCTP in infecting nematodes and attenuated their parasitism. Furthermore, MeTCTP could suppress programmed cell death triggered by the pro-apoptotic protein BAX. These results demonstrate that MeTCTP is a novel plant-parasitic nematode effector that promotes parasitism, probably by suppressing programmed cell death in host plants., (© 2016 BSPP and John Wiley & Sons Ltd.) more...

Small peptides play important roles in intercellular signaling. Inflorescence deficient in abscission (ida) is an Arabidopsis mutant that does not abscise (shed) its flower petals. The IDA gene encodes a small, secreted peptide that putatively binds to two redundant receptor-like kinases (HAESA and HAESA-like2) that initiate a signal transduction pathway. We identified IDA-like (IDL) genes in the genomic sequence for Meloidogyne incognita and Meloidogyne hapla. No orthologous sequences were found in any other genus of nematodes. Transcript for both M. incognita and M. hapla IDLs were found in total RNA isolated from infected root systems of tomato, Solanum lycopersicum. Five and three prime RACE of RNA from M. incognita infected tomato roots revealed a sequence of 392 nt that includes a poly (A) tail of 39 nt. The open reading frame encodes a 47 aa protein with a putative 25 aa N-terminal signal peptide. Expression of MiIDL1 is very low in eggs and pre-parasitic J2 and rapidly increases in the first four days post inoculation (dpi) and then declines at approximately 14 dpi. A proposed role for the root-knot nematode IDL is discussed., (Published by Elsevier Inc.) more...

Parasitism genes encode effector proteins that are secreted through the stylet of root-knot nematodes to dramatically modify selected plant cells into giant-cells for feeding. The Mi8D05 parasitism gene previously identified was confirmed to encode a novel protein of 382 amino acids that had only one database homolog identified on contig 2374 within the Meloidogyne hapla genome. Mi8D05 expression peaked in M. incognita parasitic second-stage juveniles within host roots and its encoded protein was limited to the subventral esophageal gland cells that produce proteins secreted from the stylet. Constitutive expression of Mi8D05 in transformed Arabidopsis thaliana plants induced accelerated shoot growth and early flowering but had no visible effects on root growth. Independent lines of transgenic Arabidopsis that expressed a double-stranded RNA complementary to Mi8D05 in host-derived RNA interference (RNAi) tests had up to 90% reduction in infection by M. incognita compared with wild-type control plants, suggesting that Mi8D05 plays a critical role in parasitism by the root-knot nematode. Yeast two-hybrid experiments confirmed the specific interaction of the Mi8D05 protein with plant aquaporin tonoplast intrinsic protein 2 (TIP2) and provided evidence that the Mi8D05 effector may help regulate solute and water transport within giant-cells to promote the parasitic interaction. more...

Arbuscular mycorrhizal (AM) fungi and plant growth-promoting rhizobacteria (PGPR) have potential for the biocontrol of soil-borne diseases. The objectives of this study were to quantify the interactions between AM fungi [Glomus versiforme (Karsten) Berch and Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe] and PGPR [Bacillus polymyxa (Prazmowski) Mace and Bacillus sp.] during colonization of roots and rhizosphere of tomato (Lycopersicon esculentum Mill) plants (cultivar Jinguan), and to determine their combined effects on the root-knot nematode, Meloidogyne incognita, and on tomato growth. Three greenhouse experiments were conducted. PGPR increased colonization of roots by AM fungi, and AM fungi increased numbers of PGPR in the rhizosphere. Dual inoculations of AM fungi plus PGPR provided greater control of M. incognita and greater promotion of plant growth than single inoculations, and the best combination was G. mosseae plus Bacillus sp. The results indicate that specific AM fungi and PGPR can stimulate each other and that specific combinations of AM fungi and PGPR can interact to suppress M. incognita and disease development., (© Springer-Verlag 2012) more...

The root-knot nematode, Meloidogyne incognita, is among the most damaging agricultural pests, particularly to tomato. The mutualistic endophytes Fusarium oxysporum strain Fo162 (Fo162) and Rhizobium etli strain G12 (G12) have been shown to systemically induce resistance toward M. incognita. By using triple-split-root tomato plants, spatially separated but simultaneous inoculation of both endophytes did not lead to additive reductions in M. incognita infection. More importantly, spatially separated inoculation of Fo162 and G12 led to a reduction in Fo162 root colonization of 35 and 39% when G12 was inoculated on a separate root section of the same plant in two independent experiments. In an additional split-root experiment, spatial separation of Fo162 and G12 resulted in a reduction of Fo162 root colonization of approximately 50% over the water controls in two independent experiments. The results suggested that the suppressive activity of G12 on Fo162 and M. incognita is possibly related to the induction of specific plant defense mechanisms. Thus, although Fo162 and G12 have the ability to systemically repress M. incognita infection in tomato, they can be considered incompatible biocontrol agents when both organisms are present simultaneously on the same root system. more...

The root-knot nematode Meloidogyne incognita poses a worldwide threat to agriculture, with an increasing demand for alternative control options since most common nematicides are being withdrawn due to environmental concerns. The biocontrol potential of arbuscular mycorrhizal fungi (AMF) against plant-parasitic nematodes has been demonstrated, but the modes of action remain to be unraveled. In this study, M. incognita penetration of second-stage juveniles at 4, 8 and 12 days after inoculation was compared in tomato roots (Solanum lycopersicum cv. Marmande) pre-colonized or not by the AMF Glomus mosseae. Further life stage development of the juveniles was also observed in both control and mycorrhizal roots at 12 days, 3 weeks and 4 weeks after inoculation by means of acid fuchsin staining. Penetration was significantly lower in mycorrhizal roots, with a reduction up to 32%. Significantly lower numbers of third- and fourth-stage juveniles and females accumulated in mycorrhizal roots, at a slower rate than in control roots. The results show for the first time that G. mosseae continuously suppresses root-knot nematodes throughout their entire early infection phase of root penetration and subsequent life stage development. more...

Aim: To screen and evaluate the biocontrol potential of Paenibacillus strains against disease complex caused by Meloidogyne incognita and Fusarium oxysporum f. sp. lycopersici interactions., Methods and Results: Paenibacillus strains were collected from rotten ginseng roots. The strains were tested under in vitro and pots for their inhibitory activities, and biocontrol potential against disease complex caused by M. incognita and F. oxysporum f. sp. lycopersici on tomato. In in vitro experiments, among 40 tested strains of Paenibacillus spp., 11 strains showed antifungal and nematicidal activities against F. oxysporum f. sp. lycopersici and M. incognita, respectively. Paenibacillus polymyxa GBR-462; GBR-508 and P. lentimorbus GBR-158 showed the strongest antifungal and nematicidal activities. These three strains used in pot experiment reduced the symptom development of the disease complex (wilting and plant death), and increased plant growth. The control effects were estimated to be 90-98%, and also reduced root gall formation by 64-88% compared to the untreated control., Conclusion: The protective properties of selected Paenibacillus strains make them as potential tool to reduce deleterious impact of disease complex plants., Significance and Impact of the Study: The study highlights biocontrol potential of Paenibacillus strains in management of disease complex caused by nematode-fungus interaction. more...

The root-knot nematode Meloidogyne javanica induces giant cells and feeds from them during its development and reproduction. To study the cellular processes underlying the formation of giant cells, laser microdissection was used to isolate the contents of early-stage giant cells 4 and 7 days post-infection (dpi) from tomato, and cDNA libraries from both stages were generated with 87 [250 expressed sequence tag (EST) clones] and 54 (309 EST clones) individual transcripts identified, respectively. These transcripts have roles in metabolism, stress response, protein synthesis, cell division and morphogenesis, transport, signal transduction, protein modification and fate, and regulation of cellular processes. The expression of 25 selected transcripts was studied further by real-time quantitative reverse transcriptase-polymerase chain reaction. Among them, 13 showed continuous up-regulation in giant cells from 4 to 7 dpi. The expression of two transcripts was higher than in controls at 4 dpi and remained at the same level at 7 dpi; a further five transcripts were highly expressed only at 7 dpi. The Phi-1 protein gene, a cell cycle-related homologue in tobacco, was expressed 8.5 times more strongly in giant cells than in control cells at 4 dpi, but was reduced to 6.7 times at 7 dpi. Using in situ hybridization, the expression of the Phi-1 gene was preferentially localized in the cytoplasm of giant cells at 4 dpi, together with a pectinesterase U1 precursor gene. The identification of highly expressed transcripts in developing giant cells adds to the knowledge of the plant genes responsive to nematode infection, and may provide candidate genes for nematode control strategies. more...

Bacillus firmus, commercial WP formulation (BioNem) was evaluated against the root-knot nematode Meloidogyne incognita in a laboratory, greenhouse and under field conditions on tomato plants. In the laboratory tests, an aqueous suspension of BioNem at 0.5%, 1%, 1.5% and 2% concentration reduced egg hatching from 98% to 100%, 24-days after treatment. Treatment of second-stage juveniles with 2.5% and 3% concentration of BioNem, caused 100% inhibition of mobility, 24 h after treatment. In the green house trials, BioNem applied at 8 g/pot (1200 cc soil) planted with a tomato seedlings reduced gall formation by 91%, final nematode populations by 76% and the number of eggs by 45%. Consequently, plant height and biomass was increased by 71% and 50%, respectively, compared to the untreated control, 50-days after treatment application. Application of BioNem at 16 g/pot was phytotoxic to plants. In the field trails, BioNem applied at 200 and 400 kg ha(-1) was effective in reducing the number of galls (75-84%), and increased shoot height (29-31%) and weight (20-24%) over the untreated control, 45-days after treatment. Our results indicate that B. firmus is a promising microorganism for the biological control of M. incognita in tomato pots. more...

This investigation was undertaken to determine the effect of culture filtrates of different isolates of Fusarium oxysporum f. sp. lycopersici on mortality of Meloidogyne incognita juveniles and egg hatching. It was observed that different concentrations including standard extract (S.E), 1:10 and 1:100 dilutions of all fungal filtrates inhibited egg hatch when compared with control. Minimum mortality and maximum hatching was observed in BRT (showing least mortality) isolate of F. oxysporum, while maximum mortality and minimum egg hatching was recorded in BGT (showing maximum mortality) isolate. Larval mortality was decreased with a decrease in concentration and the least mortality was observed in 1:100 when compared with SE and 1:10. The potentiality of both the isolates (BRT and BGT) against root-knot nematode M. incognita was confirmed by the pathogenicity test on tomato. These observations confirmed that F. oxysporumisolates possesses variability in pathogenicity ranging from pathogenic to bio-control agent. The plants inoculated with BRT isolate failed to show wilt symptoms while plants inoculated with BGT isolate showed wilt indices. more...

Nonchemical methods and strategies for nematode management including cultural methods and engineered measures have been recommended as an alternative to methyl bromide (a major soil fumigant), due to its role in the depletion of the ozone layer. Hence, an international agreement has recently been reached calling for its reduced consumption and complete phasing out. This present research evaluates the potential of Ecologic, a biological, marine shell meal chitin material, as a soil amendment management agent for root knot nematode, Meloidogyne incognita, control, and its effect on the growth of Floradel tomato plant, Lycopersicon esculentum. To accomplish this goal, studies were conducted during which, experimental pots were set up in greenhouse environments using sterilized soil inoculated with 5,000 root-knot eggs per 1500 g soil. There were 4 treatments and 5 replications. Treatments were: No chitin; 50 g chitin; 100 g chitin; and 200 g chitin. A two-week wait period following Ecologic amendment preceded Floradel tomato planting to allow breakdown of the chitin material into the soil. Fresh and dry weights of shoot and root materials were taken as growth end-points. A statistically significant difference (p < or = 0.05) was obtained with regard to the growth rate of L. esculentum at 100 g chitin treatment compared to the control with no chitin. Mean fresh weights of Floradel tomato were 78.0 +/- 22.3 g, 81.0 +/- 20.3 g, 109.0 +/- 25.4 g and 102.0 +/- 33.3 g at 0, 50, 100 and 200 g chitin, respectively. The analysis of root knot nematode concentrations indicated a substantial effect on reproduction rate associated with chitin amendment. Study results showed a significant decrease in both root knot nematode eggs and juveniles (J2) at 100g and 200 g Ecologic chitin levels, however, an increase in nematode concentrations was recorded at the 50 g Ecologic chitin level (p < or = 0.05). The mean amounts of J2 population, as expressed per 1500 cm3 soil, were 49,933 +/- 38,819, 86,050 +/- 25248, 103 +/- 133 and 103 +/- 133 for 0, 50, 100 and 200 g chitin, respectively. Similarly, the mean numbers of root knot nematode eggs (per 1500 cm3 of soil) were 40,759 +/- 36,712, 66,048 +/- 39,730, 9,904 +/- 16,591 and 9,257 +/- 17,204. Root gall rating was also significantly lower (p < or = 0.05) at the 100 g and 200 g chitin levels compared to the control. Percent gall ratings were 3.3 +/- 1.0%, 3.2 +/- 1.0%, 1.0 +/- 0.5%, and 1.0% +/- 0.6% for amendment levels of 0, 50, 100, and 200 g chitin, respectively. more...

Management of root knot nematode disease infecting tomato, by the use of fungal bioagents Acremonium strictum and Trichoderma harzianum isolated from egg masses of M. incognita infecting tomato has been carried out. The rhizosphere and rhizoplane of root knot nematode infested tomato revealed consistent association of Acremonium strictum. In the present study A. strictum and other fungal bioagents viz. Aspergillus niger, Paecilomyces lilacinus, Rhizoctonia solani and Trichoderma harzianum isolated earlier from egg-masses of M. incognita, identified and maintained have been investigated through in-vitro and in-vivo trials for their potentiality against M. incognita. Out of the above, isolated mycoflora A. niger was identified to be toxic against M. incognita while A. strictum and T. harzianum was found to possess both egg parasitic or opportunistic and toxic properties. A field trial with all the above fungal bioagents both alone and together showed significant promising performance by the dual treatment of A. strictum and T. harzianum in improving the health of the tomato plant with a remarkable reduction in M. incognita population. more...

Fresh crop and garden residues were applied both under laboratory conditions and in commercial greenhouse in order to asses their effect on soil nematodes populations and soil fertility. In the laboratory experiments, dosages of 5 to 20 g of cabbage residues, chicken manure, cabbage residues+chicken manure, grass+chicken manure, as well as leaves and stems of orange tree, pine tree, oleander, olive tree, palm tree and boxwood were mixed with 500 g soil having root-knot nematodes (Meloidogyne incognita) and soil moisture was adjusted at field capacity. A control treatment without residues was also included. The mixtures were kept into plastic bags, with four replications, and the bags were incubated for four weeks at 30 degrees C, when nematological and soil fertility analyses were carried out. In general, all these materials significantly (P < 0.05) reduced M. incognita populations and increased saprophagous nematodes, with slight effects on soil fertility except for the K increase with residues application. Tomato plants susceptible to M. incognita were planted in pots with 300 cm3 of the treated soils and kept for five weeks in a growth chamber (24 +/- 1 degrees C, 14 hours light), when root galling indices were evaluated. Most materials applied reduced root galling indices as regards to the control. In the greenhouse experiment, cabbage residues, cabbage residues+chicken manure, grass+chicken manure and grass+cabbage residues were applied to the soil and covered with a polyethylene sheet for 5 weeks. A cabbage residues:chicken manure treatment and a control (not-amended) treatment, without polyethylene, were also included. At the end of the experiment, the nematological analysis showed that all materials successfully controlled M. incognita populations, reaching 86-100% mortality with organic amendments vs. 6% for the control. After the greenhouse biodesinfestation experiment, a tomato crop was grown for one month, when root galling indices were determined. All materials significantly reduced this value from 4.75 in the control to 1.0-2.25 with the organic amendments, except for the cabbage residues+chicken manure treatment without polyethylene (index = 4.0). Our results show that fresh crop and garden residues successfully reduced M. incognita populations and root galling indices when applied with polyethylene covers, having good potential to be considered in integrated management programs. more...

Experiments were carried out to study the effect of two fungal bioagents along with mustard oil cake and furadan against root knot nematode Meloidogyne incognita infecting tomato under greenhouse condition. Bioagents viz., Paecilomyces lilacinus and Trichoderma viride alone or in combination with mustard cake and furadan promoted plant growth, reduced number of galls/plant, egg masses/root system and eggs/egg mass. The fungal bioagents along with mustard cake and nematicide showed least nematodes reproduction factor as compared to untreated infested soil. more...

Trichoderma harzianum and Trichoderma viride were tested for their capacity to reduce the incidence of the root-knot nematode Meloidogyne incognita on tomato. In vitro studies demonstrated that all tested isolates were effective in causing nematode mortality compared with the control. Trichoderma slightly reduced nematode damage to tomato in vivo. Treatment of the soil with the biocontrol agents before transplanting, improved control over treatment directly at transplanting. The Trichoderma isolates could not be re-isolated from the endorhiza, but were successfully re-isolated from the rhizosphere 45 days after fungal inoculation. Only slight increases in plant growth could be measured. The mutualistic endophyte F. oxysporum 162, used as positive control, was more effective in root-knot nematode biocontrol than the Trichoderma isolates. more...

In pot trial, dried ground weed leaves of Cynodon dactylon, Datura stramonium, Eichhomia crassipes, Emex spinosus, Ricinus communis and Sisymbrium irio were mixed with soil at the rate of 1, 3, 5 and 10 g/kg soil and compared their nematicidal potential with carbofuran as a standard against the root-knot nematode, M. incognita infecting tomato. In addition, their effects on growth rate of tomato plants were also investigated. The results showed that M. incognita populations in the soil and root galling were significantly suppressed when the dried leaves of the tested weeds at all rates were allowed to decompose in the soil. All amendments exhibited varying degree of reduction compared to control. The highest reduction was noticeable with the plants grown in Sisymbrium irio amended soil followed by Datura stramonium and Emex spinosus. In addition, employing high rate of the tested weeds gave higher activity in suppressing the nematode both in the soil and in tomato roots than using low rate. The data also indicated that all amendments at low rates significantly increased growth indices of tomato over control treatment, except Cynodon dactylon and Emex spinosus which decreased it, particularly in the shoot system. On the other hand, their high rates showed phytotoxic effects. These weed species may offer considerable promise as soil amendments for control of root-knot nematode, M. incognita. more...

Pasteuria penetrans will build-up faster where there is a high initial nematode density and can suppress root-knot nematode populations in the roots of tomato plants. The effect of different initial densities of nematode (Meloidogyne javanica) (150, 750, 1500, 3000) and P. penetrans infected females (F1, F3) densities (F0=control and AC=absolute control without nematode or P. penetrans inoculum) on the build-up of Pasteuria population was investigated over four crop cycles. Two major points of interest were highlighted. First, that within a confined soil volume, densities of P. penetrans can increase >100 times within 2 or 3 crop cycles. Second, from a relatively small amount of spore inoculum, infection of the host is very high. There were more infected females in the higher P. penetrans doses. The root growth data confirms the greater number of females in the controls particularly at the higher inoculum densities in the third and fourth crops. P. penetrans generally caused the fresh root weights to be higher than those in the control. P. penetrans has shown greater reduction of egg masses per plant at most densities. The effects of different initial densities of M. javanica and P. penetrans on the development of the pest and parasite populations were monitored. And no attempt was made to return the P. penetrans spores to the pots after each crop so the build-up in actual numbers of infected females and spores under natural conditions may be underestimated. more...

The microbial and nematode populations associated with two plants (tomato and cabbage) inoculated with the nematophagous fungus, Pochonia chlamydosporia var. chlamydosporia or root knot nematode (Meloidogyne incognita), or both, were compared with those in unplanted controls. The dominant factor affecting culturable microbial populations was found to be the presence or absence of tomato plants. Generally microbial colony counts were lowest in unplanted soil, small increases were associated with cabbage and significantly greater numbers with tomato plants. Differences in microbial diversity (estimated from community profiles of carbon substrate utlisation, using Biolog) were observed between planted and unplanted soils, however, there were few differences between soils with either of the two plants. The presence of P. chlamydosporia was associated with a reduction in the numbers of plant parasitic nematodes (51%-78%) including the migratory ectoparasites, whereas free-living nematodes, culturable bacteria and bacterial populations assessed by Biolog were unaffected by the application of fungus. more...

On the first of seeds germination, Meloidogyne javanica induced knots on the stem eventually by to penetrate into the coleoptiles. In this experiment, used soil was loamy-sandy type, pH=7.2, including 1.5% composts. Tomato seedlings of cultivar Rutgers were used as host. This experiment was conducted in Completely Randomized Design with 3 treatments and 10 replicates as fallow: (1) control, (2) 10 J2s/cm3 of soil, (3) 20 J2s/cm3 of soil. In each pot (1000 cm3), 50 seeds were planted. Inoculation was done, a day after to plant seeds, in 2 cm depth. Pre-emergence and post-emergence damping-off, number of stem-knots, height of knots on the stem in 3 treatments was more than 2 treatments. In some of the pots of 2 treatments were not observed stem-knots. As long as four weeks after planting, there was not significant difference in fresh weight and height of the seedlings in treatments. Numbers of adult females in 3 treatments were more than 2. In the stem-knots, collenchymas tissue, endodermis and stem cambium were found deformed J2s and J3s, and adult females. more...

An integrated approach with the obligate bacterial parasite, Pasteuria penetrans and nematicides was assessed for the management of the root-knot nematode, Meloidogyne incognita infestation in tomato and grapevine. Seedlings of tomato cv. Co3 were transplanted into pots filled with sterilized soil and inoculated with nematodes (5000 juveniles/pot). The root powder of P. penetrans at 10 mg/pot was applied alone and in combination with carbofuran at 6 mg/pot. Application of P. penetrans along with carbofuran recorded lowest nematode infestation (107 nematodes/200 g soil) compared to control (325 nematodes/200 g soil). The rate of parasitization was 83.1% in the carbofuran and P. penetrans combination treatment as against 61.0% in the P. penetrans treatment only. The plant growth was also higher in the combination treatment compared to all other treatments. A field trial was carried out to assess the efficacy of P. penetrans and nematicides viz., carbofuran and phorate in the management of root-knot nematode, M. incognita infestation of grapevine cv. Muscat Hamburg. A nematode and P. penetrans infested grapevine field was selected and treatments either with carbofuran or phorate at 1 g a.i/vine was given. The observations were recorded at monthly interval. The results showed that the soil nematode population was reduced in nematicide treated plots. Suppression of nematodes was higher under phorate (117 nematodes/200 g soil) than under carbofuran (126.7 nematodes/200 g soil) treatment. The number of juveniles parasitized was also influenced by nematicides and spore load carried/juvenile with phorate being superior and the increase being 17.0 and 29.0% respectively over the control. The results of these experiment confirmed the compatibility of P. penetrans with nematicides and its biological control potential against the root-knot nematode. more...

The alternative ecological methods require investigation of many organo-biological means for plant protection against dangerous root parasites such as root-knot nematode Meloidogyne arenaria and some ectoparasites (Xiphinema index). The Bulgarian organic product - dry extract of green alga Chlorella vulgaris ("The Golden Apple"-Plamen Barakov) is the latest product, which in comparative aspect gives the best results. Series of laboratory and pot experiments are carried out with tomato (cv. Bele and cv. Ideal) and grape seedlings (cv. Cabernet Sauvignon). Different dosages of Chlorella from 0.5 g to 2 g per plant/pot are investigated. The first results show that even low dosages had double effect - on the one hand they suppress the parasite development and on the other hand they strongly stimulate plant growing. The very important conclusion is that Chlorella vulgaris ignores the negative influence of M. arenaria and X. index. These results give us opportunity for future model and field investigations of Chlorella vulgaris with the aim of its practical application. more...

Aims: The aim was to determine the influence of some Aspergillus species on the production of nematicidal agent(s) in vitro and biocontrol of Meloidogyne javanica in tomato by Pseudomonas fluorescens strains CHA0 and CHA0/pME3424., Methods and Results: Six species of Aspergillus, isolated from the rhizosphere of certain crops, produced a variety of secondary metabolites in vitro. Culture filtrate (CF) obtained from Ps. fluorescens strain CHA0 and its2,4-diacetylphloroglucinol overproducing mutant CHA0/pME3424 grown in King's B liquid medium caused significant mortality of M. javanica juveniles in vitro. Bacterial growth medium amended with CF of A. niger enhanced nematicidal and beta-galactosidase activities of fluorescent pseudomonads while A. quadrilineatus repressed such activities. Methanol or ethyl acetate extracts of the CF of A. niger markedly optimized bacterial efficacy to cause nematode deaths while hexane extract of the fungus had no influence on the nematicidal activity of the bacterial strains. A. niger applied alone or in conjunction with the bacterial inoculants inhibited root-knot nematode galling in tomato. On the other hand, A. quadrilineatus used alone or together with CHA0 did not inhibit nematode galling but when used in combination with strain CHA0/pME3424 did reduce galling intensity., Conclusions: Aspergillus niger enhances the production of nematicidal compounds by Ps. fluorescensin vitro and improves biocontrol potential of the bacterial inoculants in tomato while A. quadrilineatus reduces bacterial performance to suppress root-knot nematodes., Significance and Impact of the Study: Rhizosphere harbours a variety of micro-organisms including bacteria, fungi and viruses. Aspergillus species are ubiquitous in most agricultural soils and generally produce a variety of secondary metabolites. Such metabolites synthesized by Aspergillus species may influence the production of nematicidal agents and subsequent biocontrol performance of the bacterial inoculants against plant-parasitic nematodes. This fact needs to be taken into consideration when using biocontrol strains in an agriculture system. more...

Aims: To determine the influence of soil-borne fungus Trichoderma harzianum on the biocontrol performance of Pseudomonas fluorescens strain CHA0 and its 2,4-diacetylphloroglucinol (DAPG) overproducing derivative CHA0/pME3424 against Meloidogyne javanica., Methods and Results: Amendment of the culture filtrate (CF) or methanol extract of the CF of a T. harzianum strain Th6 to P. fluorescens growth medium enhanced the production of nematicidal compound(s) by bacterial inoculants in vitro. In addition, bacteria overwhelmingly expressed phl'-'lacZ reporter gene when the medium was amended with CF of T. harzianum. Pseudomonas fluorescens and T. harzianum applied together in unsterilized sandy loam soil caused greater reduction in nematode population densities in tomato roots., Conclusions: Trichoderma harzianum improves root-knot nematode biocontrol by the antagonistic rhizobacterium P. fluorescens both in vitro and under glasshouse conditions., Significance and Impact of the Study: The synergistic effect of T. harzianum on the production of nematicidal compound(s) critical in biocontrol may improve the efficacy of biocontrol bacteria against plant-parasitic nematodes. Considering the inconsistent performance of the biocontrol agents under field conditions, application of a mixture of compatible T. harzianum and P. fluorescens would more closely mimic the natural situation and might broaden the spectrum of biocontrol activity with enhanced efficacy and reliability of control. more...

The root galling index and the densities of eggs in roots and juveniles in soil of the root-knot nematode Meloidogyne javanica (Treub) Chitwood on tomato, and the effect of these on crop yield were assessed in greenhouse experiments applying various treatments at two different sites in Crete, Greece. Tomato crops were grown for four cycles by rotating nematode-resistant (first and third spring crops) with susceptible (second and fourth autumn crops) cultivars and receiving the following treatments: (a) untreated control; (b) methyl bromide application before the first and third crops; (c) application of the fungus Pochonia chlamydosporia (Goddard) Zare, Gams & Evans before planting the first, third and fourth crops with a supplementary application three weeks after the beginning of the fourth crop; (d) application of oxamyl in both sites and fenamiphos in site 1 only at the second and fourth crops; (e) combination of treatments (c) and (d). The fungus density in soil was monitored three weeks after application and at the end of each crop, when roots were lifted. Pochonia chlamydosporia had a variable establishment and did not control the nematode. Its pathogenicity on eggs was not demonstrated, as in all cases galls were big, with all egg masses inside and protected from infection. The methyl bromide treatment significantly reduced root galling and egg production compared to other treatments in all crop cycles and the yield of the fourth crop was significantly greater. Nematicides reduced nematode densities compared with untreated controls and the fungus treatment, but they were less effective than methyl bromide and resulted in increased yield in one site only. more...

Aims: To assess the effects of various carbon and mineral sources on the nematicidal potential of biocontrol inoculants of Pseudomonas aeruginosa IE-6S+ and Ps. fluorescens CHA0 under laboratory and glasshouse conditions., Methods and Results: Culture filtrates of strains IE-6S+ and CHA0, cultured in nutrient yeast extract broth, caused substantial mortality of the juveniles of Meloidogyne javanica. The nematicidal activities of the culture filtrates were altered after amendment with various carbon and mineral sources. Soil amendment with zinc alone or in combination with glycerol improved the biocontrol efficacy against root-knot nematode, promoted tomato plant growth and enhanced bacterial rhizosphere and endophytic colonization., Conclusions: Appropriate quantities of glycerol and zinc alone or in combination enhance the nematicidal activity of Ps. aeruginosa and Ps. fluorescens. Glucose reduces the activity of these bacteria against nematodes., Significance and Impact of the Study: Minerals and carbon sources are appealing because they are easy and economical to provide during liquid fermentation of inoculants or as fertilizer amendments to improve the biocontrol activity of indigenous and introduced bacteria. more...

TerraPy, Magic Wet and Chitosan are soil and plant revitalizers based on natural renewable raw materials. These products stimulate microbial activity in the soil and promote plant growth. Their importance to practical agriculture can be seen in their ability to improve soil health, especially where intensive cultivation has shifted the biological balance in the soil ecosystem to high numbers of plant pathogens. The objective of this study was to investigate the plant beneficial capacities of TerraPy, Magic Wet and Chitosan and to evaluate their effect on bacterial and nematode communities in soils. Tomato seedlings (Lycopersicum esculentum cv. Hellfrucht Frühstamm) were planted into pots containing a sand/soil mixture (1:1, v/v) and were treated with TerraPy, Magic Wet and Chitosan at 200 kg/ha. At 0, 1, 3, 7 and 14 days after inoculation the following soil parameters were evaluated: soil pH, bacterial and fungal population density (cfu/g soil), total number of saprophytic and plant-parasitic nematodes. At the final sampling date tomato shoot and root fresh weight as well as Meloidogyne infestation was recorded. Plant growth was lowest and nematode infestation was highest in the control. Soil bacterial population densities increased within 24 hours after treatment between 4-fold (Magic Wet) and 19-fold (Chitosan). Bacterial richness and diversity were not significantly altered. Dominant bacterial genera were Acinetobacter (41%) and Pseudomonas (22%) for TerraPy, Pseudomonas (30%) and Acinetobacter (13%) for Magic Wet, Acinetobacter (8.9%) and Pseuodomonas (81%) for Chitosan and Bacillus (42%) and Pseudomonas (32%) for the control. Increased microbial activity also was associated with higher numbers of saprophytic nematodes. The results demonstrated the positive effects of natural products in stimulating soil microbial activity and thereby the antagonistic potential in soils leading to a reduction in nematode infestation and improved plant growth. more...

In previous work, the four endophytic bacteria Pantoea agglomerans MK-29, Cedeca davisae MK-30, Enterobacter spp. MK-42 and Pseudomonas putida MT-19 were shown to reduce Meloidogyne incognita on tomato when applied as a seed treatment and/or soil drench. The objective of this work was to study these bacteria for their potential to induce systemic resistance against root knot nematodes on tomato. To guarantee spatial separation between inducing agent and pathogen a split-root system was chosen and inoculated with the bacteria as a drench application on one side of the root system and 6 days later with 2000 juveniles of Meloidogyne incognita on the other side of the split-root system. The experiment was maintained in the greenhouse and repeated once. The penetration rate of juveniles as well as the total number of root-knot galls and egg masses was recorded. Treatment with all four bacteria significantly reduced juvenile penetration and the number of root-knot galls when compared with the non-treated control. Induced systemic resistance is considered a possible control mechanism of endophytic bacteria against root-knot nematodes. more...

Various lines of evidence show that local changes in the auxin concentration are involved in the initiation and directional expansion of syncytia induced by cyst nematodes. Analysis of nematode infections on auxin-insensitive tomato and Arabidopsis mutants revealed various phenotypes ranging from complete inhibition of syncytium development to a decrease in hypertrophy and lateral root formation at the infection site. Specific activation of an auxin-responsive promoter confirmed the role of auxin and pointed at a local accumulation of auxin in developing syncytia Disturbance of auxin gradients by inhibiting polar auxin transport with N-(1-naphthyl)phtalamic acid (NPA) resulted in abnormal feeding cells, which were characterized by extreme galling, massive disordered cell divisions in the cortex, and absence of radial expansion of the syncytium initial toward the vascular bundle. The role of auxin gradients in guiding feeding cell morphogenesis and the cross-talk between auxin and ethylene resulting in a local activation of cell wall degrading enzymes are discussed. more...

Efficient selection of fungi for biological control of nematodes requires a series of screening assays. Assessment of genetic diversity in the candidate species maximizes the variety of the isolates tested and permits the assignment of a particular genotype with high nematophagous potential using a rapid novel assay. Molecular analyses also facilitate separation between isolates, allowing the identification of proprietary strains and trace biocontrol strains in the environment. The resistance of propagules to UV radiation is an important factor in the survival of a biocontrol agent. We have analyzed 15 strains of the nematophagous fungus Paecilomyces lilacinus using these principles. Arbitrarily primed DNA and allozyme assays were applied to place the isolates into genetic clusters, and demonstrated that some genetically related P. lilacinus strains exhibit widespread geographic distributions. When exposed to UV radiation, some weakly nematophagous strains were generally more susceptible than effective isolates. A microtitre tray-based assay used to screen the pathogenic activity of each isolate to Meloidogyne javanica egg masses revealed that the nematophagous ability varied between 37%-100%. However, there was no clear relationship between nematophagous ability and genetic clusters. Molecular characterizations revealed sufficient diversity to allow tracking of strains released into the environment. more...