Plant parasitic nematodes associated with ornamental plant Pseuderanthemum atropurpureum in Aligarh district of western Uttar Pradesh

A survey was conducted to determine the nematodes associated with the soil, substratum and roots of the ornamental plant, Pseuderanthemum atropurpureum, grown in the Aligarh district of western Uttar Pradesh, India. Ten genera of plant parasitic nematodes, viz. Meloidogyne spp., Helicotylenchus sp., Hemicriconemoides sp., Hoplolaimus sp., Xiphinema sp., Pratylenchus sp., Trichodorus sp., Tylenchorhynchus sp., Aphelenchoides sp. and Rotylenchus sp., were isolated from 345 soil samples collected from 15 different localities. Our study indicates that among the 10 genera of the plant parasitic nematodes, Meloidogyne spp. was widely spread in all the examined localities of Aligarh district, except at Khair. It was observed that the population density and diversity of nematodes in all the soil samples was not uniform. Meloidogyne spp. showed the highest absolute frequency and relative frequency followed by Helicotylenchus sp., Hemicriconemoides sp., Xiphinema sp., Hoplolaimus sp., Pratylenchus sp., Trichodorus sp., Tylenchorhynchus sp., Aphelenchoides sp. and Rotylenchus sp. Since no records are available in the literature related to the association of root-knot nematode in P. atropurpureum, it is worthy to note that this study may be considered as a new hope towards further study of the interaction between the root-knot nematode and ornamental plant P. atropurpureum if any.     

Comparative study on the host susceptibility of different banana cultivars to plant parasitic nematodes in Malappuram district of Kerala,India

Six cultivars of banana grown widely in Malappuram district of Kerala were evaluated for the diversity of plant parasitic nematodes in their respective rhizosphere. The genotypes evaluated include Nendran, Poovan (Mysore), Njali poovan, Mysore poovan, Chenkadali and Ponnan. The study aims on a comparative analysis of rhizosphere of different banana varieties for the occurrence of plant parasitic nematodes, diversity, frequency of occurrence and to determine the most vulnerable type of banana variety. Cobb’s decanting and sieving method was used to isolate nematodes from soil. Thirty three samples randomly collected from different blocks of Malappuram district are subjected to analysis and a ten species of nematodes were reported. Among the six varieties analysed both Nendran and Mysore Poovan variety seems to be more susceptible to phytoparasitic nematodes. With respect to eight blocks of Malappuram district, Helicotylenchus sp. shows highest prominence value followed by Radopholus similis, reniform nematode and Meloidogyne incognita.     

Effect of tillage,subsidiary crops and fertilisation on plant‐parasitic nematodes in a range of agro‐environmental conditions within Europe

The overall goal in nematode management is to develop sustainable systems where nematode populations are kept under the economic damage threshold. Conservation tillage and subsidiary crops, applied as cover crops and living mulches, generally improve soil health by increasing soil organic matter content and stimulating soil microbial activity. However, more permanent crop and weed cover associated with subsidiary crops and noninversion tillage, respectively, may benefit plant‐parasitic nematodes with broad host spectra such as Meloidogyne and Pratylenchus. These genera are major constraints to many field crops throughout Europe and there is a need to identify effective and reliable management options that can be applied to avoid excessive infestations. The dynamics of the indigenous fauna of plant‐parasitic nematodes were studied in eight coordinated multi‐environment field experiments (MEEs) under four agro‐environmental conditions in Europe (Continental, Nemoral, Atlantic North and Mediterranean North). The MEEs consisted of a 2‐year sequence of wheat combined with a living mulch or subsequent cover crops and second main crops maize, potatoes or tomatoes depending on site. Additionally, the effects of inversion tillage using the plough were compared with various forms of conservation tillage (no‐tillage, shallow and deep noninversion tillage). Overall, Helicotylenchus, Paratylenchus, Pratylenchus and Tylenchorhynchus were the most frequent genera across sites while Meloidogyne occurred only in Germany at very low densities. During the wheat–maize sequences in Switzerland, the populations of Pratylenchus increased from 63 to 146 nematodes per 100 mL soil and Helicotylenchus from 233 to 632 nematodes per 100 mL soil. The effects of tillage on plant‐parasitic nematodes were generally minor, although no tillage in Italy supported higher densities of Pratylenchus (184 nematodes per 100 mL soil) than inversion tillage (59 nematodes per 100 mL soil). Furthermore, Pratylenchus densities were 160 nematodes per 100 mL soil when leguminous subsidiary crops were grown, 122 nematodes per 100 mL soil in the green fallow and 84 nematodes per 100 mL soil after growing black oat (Avena strigosa) or oilseed radish (Raphanus sativus). The differences were greatest in Italy, in a sandy soil with low organic matter. Application of compost or nitrogen fertiliser had no consistent effects on plant‐parasitic nematodes. We conclude that crop rotations including specific subsidiary crops are prominent factors affecting the indigenous nematode community, while tillage and fertiliser are of lower importance.     

Differential Colonization Patterns of Bananas (Musa spp.) by Physiological Race 1 and Race 4 Isolates of Fusarium oxysporum f.sp. cubense

Fusarium oxysporum f.sp. cubense (Foc) is the causative agent of Fusarium wilt of bananas (Musa spp.). To clarify the colonization patterns of Foc in bananas, two green fluorescent protein‐tagged isolates, NT320 (race 1) and B2‐gfp (race 4), were used to follow infection of the banana varieties Pisang Awak and Brazil. Penetration and colonization of both isolates in roots of these two banana varieties were observed within 6 days, but sporulation in xylem vessels was not observed until day 30 postinoculation. Interestingly, B2‐gfp penetrated into xylem vessels of Pisang Awak banana roots more quickly than NT320, implying that the race 4 isolate is more virulent than the race 1 isolate. This result was further confirmed by comparing the disease severity of plants inoculated with NT320 with that of plants inoculated with B2‐gfp. Quantitative real‐time PCR revealed that some pathogenicity‐associated genes, including Fga1, Fhk1, Fow2 and Ste12, were upregulated by B2‐gfp during exposure to Brazil bananas, while they were either downregulated by NT320 or not significantly changed. These data might partly explain why the race 4 isolate was more virulent than the race 1 isolate.     

Combined application of native Trichoderma isolates possessing multiple functions for the control of Fusarium wilt disease in banana cv. Grand Naine

Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is considered as a lethal disease of bananas worldwide. To manage the disease effectively, 20 rhizospheric and 43 endophytic Trichoderma isolates obtained from 12 different Foc resistant banana accessions were evaluated against Foc in vitro and in vivo. In vitro screening among Trichoderma isolates for their multiple functions (mycelial and spore germination inhibition, hydrogen cyanide, chitinolytic enzymes, non-volatile and volatile metabolites production) in suppressing Foc and promoting plant growth (IAA production and phosphate solubilisation) indicated that the multiple biocontrol actions were significantly higher in 6 isolates of rhizospheric Trichoderma and 10 isolates of endophytic Trichoderma compared to other isolates. The greenhouse evaluation of individual application of these rhizospheric and endophytic Trichoderma isolates against Fusarium wilt pathogen in cv. Grand Naine (AAA) indicated significant suppression of Fusarium wilt disease and increased plant growth characters as compared to Foc pathogen inoculated plants. However, none of these individual Trichoderma isolates recorded complete suppression of Fusarium wilt disease. Therefore, the greenhouse evaluation involving combination of rhizospheric Trichoderma sp. NRCB3 + endophytic Trichoderma asperellum Prr2 recorded 100% reduction of Fusarium wilt disease and increased plant growth parameters up to 250% when compared to individual isolates application and Foc alone-inoculated plants. Further, the field evaluation of this combination of Trichoderma isolates applied for three times: (1) at 15 days before planting, (2) second month after planting and (3) fourth month after planting resulting in significant reduction of Fusarium wilt disease and also increase in bunch weight as compared to untreated control plants. Therefore, these Trichoderma isolates may be used in combination for the effective suppression of Fusarium wilt disease in banana.     

生物肥与甲壳素和恶霉灵配施对香蕉枯萎病的防治效果

通过盆栽试验研究了生物肥与甲壳素和恶霉灵配施防治香蕉枯萎病效果,试验结果表明,生物肥与恶霉灵配施(H+F)处理香蕉枯萎病病情指数最高,生物肥与甲壳素配施(C+F)处理病情指数最低。单独生物肥处理防病效果为32.8%,生物肥与甲壳素配施处理为42.5%,而生物肥与恶霉灵配施加重了香蕉枯萎病病情。Biolog Eco微平板研究发现,AWCD(平均每孔颜色变化率)和Shannon等4个多样性指数变化趋势与防病效果相反:防病效果好的处理,土壤细菌功能多样性指数反而低,经检测发现病原真菌(Fusarium oxysporum f. sp. cubense)可利用Biolog Eco微平板上碳源底物并发生颜色变化,干扰测定结果。T-RFLP分析土壤细菌DNA多样性,对照(灭菌生物肥)土壤中TRFs末端限制性片段最少,生物肥与甲壳素配施处理最多。与网上数据库比较,生物肥与甲壳素配施增加了土壤中芽胞杆菌种类,与恶霉灵配施降低了芽胞杆菌种类。分析发现,T-RFLP和Biolog的主成份分析载荷图具有较高一致性。因此,生物肥与生物农药甲壳素配施,从生态角度控制土传病害,优势互补,提高了土壤细菌多样性,改善了土壤细菌群落结构,有利于提高防病效果。     

Combined application of botanical formulations and biocontrol agents for the management of Fusarium oxysporum f. sp. cubense (Foc) causing Fusarium wilt in banana

Plant products along with biocontrol agents were tested against Fusarium wilt of banana caused by Fusarium oxysporum f. sp. cubense (Foc). Of the 22 plant species tested, the leaf extract of Datura metel (10%) showed complete inhibition of the mycelial growth of Foc. Two botanical fungicides, Wanis 20 EC and Damet 50 EC along with selected PGPR strains with known biocontrol activity, Pseudomonas fluorescens 1, Pf1 and Bacillus subtilis, TRC 54 were tested individually and in combination for the management of Fusarium wilt under greenhouse and field conditions. Combined application of botanical formulation and biocontrol agents (Wanis 20 EC + Pf1 + TRC 54) reduced the wilt incidence significantly under greenhouse (64%) and field conditions (75%). Reduction in disease incidence was positively correlated with the induction of defense-related enzymes peroxidase (PO) and polyphenol oxidase (PPO). Three antifungal compounds (two glycosides and one ester) in D. metel were separated and identified using TLC, RP-HPLC (Reverse Phase-High Pressure Liquid Chromatography) and mass spectrometry. In this study it is clear that combined application of botanical formulations and biocontrol agents can be very effective in the management of Fusarium wilt of banana.     

Identification and validation of reference genes for RT‐qPCR analysis in banana (Musa spp.) under Fusarium wilt resistance induction conditions

Banana (Musa spp.) is severely damaged by Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc). Biocontrol by inducing systemic resistance has been considered as one of the most important strategies to improve plant health. Very few studies have investigated appropriate reference gene selection for RT‐qPCR (quantitative real‐time polymerase chain reaction) analysis suitable for conditions of systemic activated resistance. In this study, we assessed over a time‐course the expression of seven candidate reference genes (EF1, TUB, ACT1, ACT2, L2, RPS2 and RAN) for Cavendish cultivar Brazilian (Musa spp. AAA) and dwarf banana cultivar Guangfen No. 1 (Musa spp. ABB) that were inoculated by Bacillus subtilis strain TR21 and Foc. We choose these plants because they are commonly planted in Southern China. Expression stability of the candidate genes was evaluated using various software packages (GeNorm, NormFinder and BestKeeper). L2 and TUB genes displayed maximum stability in Guangfen No. 1. In Brazilian, ACT1 and TUB were the most stable genes. To further validate the suitability of the reference genes identified in this study, the expression of pathogenesis‐related 1 (PR1) gene under TR21 and Foc strains Foc004/Foc009 treatments was also studied. Identified reference genes in this work that are most suitable for normalizing gene expression data in banana under Fusarium wilt resistance induction conditions will contribute to the understanding of disease resistance mechanisms induced by biocontrol strains in banana.     

Interaction of Population Levels of Fusarium oxysporum f. sp. vasinfectum and Meloidogyne incognita on Cotton

In autoclaved greenhouse soil without Fusarium oxysporum f. sp. vasinfectum, Meloidogyne incognita did not cause leaf or vascular discoloration of 59-day-old cotton plants. Plants had root galls with as few as 50 Meloidogyne larvae per plant. Root galling was directly proportional to the initial nematode population level. Fusarium wilt symptoms occurred without nematodes with 77,000 fungus propagules or more per gram of soil. As few as 50 Meloidogyne larvae accompanying 650 fungus propagules caused Fusarium wilt. With few exceptions, leaf symptoms appeared sooner as numbers of either or both organisms increased. In soils infested with both organisms, the extent of fungal invasion and colonization was well correlated with the extent of nematode galling and other indications of the Fusarium wilt syndrome.     

Seasonal dynamics of phytonematodes associated with date palm cvs. Barhi and Hyani as affected by soil temperature and moisture

Seasonal fluctuation of the reniform nematode, Rotylenchulus reniformis in soil and roots and the spiral nematode, Helicotylenchus sp. in soil and roots of date palm (Phoenix dactylifera L.) cvs. Barhi and Hyani over one year revealed that these nematodes were negatively correlated with the prevailing soil temperature (°C) and were positively correlated with percentage soil moisture at different sampling months and seasons. Correlation coefficients(r) were calculated to ascertain these relationships.     

Induction of defence-related enzymes in susceptible variety of banana: role of Fusarium-derived elicitors

Elicitor prepared from the Fusarium oxysporum f. sp cubense (Foc) isolated from infected banana rhizosphere induced the accumulation of resistance-associated enzymes in leaves of susceptible and resistant variety of banana. Roots of Grand Naine (susceptible) and robusta (resistant) variety were inoculated with 1 g/l Foc elicitors. Distinct difference in peroxidase, polyphenol oxidase, β-1,3-glucanase, chitinase and phenolics was observed in control plants of resistant and susceptible varieties. Induced defence-related enzymes in susceptible variety were increased tothe level of untreated resistant variety. This depicted that Fusarium-derived elicitor effectively induced defence in susceptible variety to the apparent level of untreated resistant variety.     

Development of a Real-Time Fluorescence Loop-Mediated Isothermal Amplification Assay for Rapid and Quantitative Detection of Fusarium oxysporum f. sp. cubense Tropical Race 4 In Soil

Fusarium oxysporum f. sp. cubense (Foc), the causal agent of Fusarium wilt (Panama disease), is one of the most devastating diseases of banana (Musa spp.). The Foc tropical race 4 (TR4) is currently known as a major concern in global banana production. No effective resistance is known in Musa to Foc, and no effective measures for controlling Foc once banana plants have been infected in place. Early and accurate detection of Foc TR4 is essential to protect banana industry and guide banana planting. A real-time fluorescence loop-mediated isothermal amplification assay (RealAmp) was developed for the rapid and quantitative detection of Foc TR4 in soil. The detection limit of the RealAmp assay was approximately 0.4 pg/µl plasmid DNA when mixed with extracted soil DNA or 10³ spores/g of artificial infested soil, and no cross-reaction with other relative pathogens were observed. The RealAmp assay for quantifying genomic DNA of TR4 was confirmed by testing both artificially and naturally infested samples. Quantification of the soil-borne pathogen DNA of Foc TR4 in naturally infested samples was no significant difference compared to classic real-time PCR (P>0.05). Additionally, RealAmp assay was visual with an improved closed-tube visual detection system by adding SYBR Green I fluorescent dye to the inside of the lid prior to amplification, which avoided the inhibitory effects of the stain on DNA amplification and makes the assay more convenient in the field and could thus become a simple, rapid and effective technique that has potential as an alternative tool for the detection and monitoring of Foc TR4 in field, which would be a routine DNA-based testing service for the soil-borne pathogen in South China.     

Apoptosis-related genes confer resistance to Fusarium wilt in transgenic 'Lady Finger' bananas

Fusarium wilt, caused by Fusarium oxysporum f. sp. cubense (Foc), is one of the most devastating diseases of banana (Musa spp.). Apart from resistant cultivars, there are no effective control measures for the disease. We investigated whether the transgenic expression of apoptosis-inhibition-related genes in banana could be used to confer disease resistance. Embryogenic cell suspensions of the banana cultivar, 'Lady Finger', were stably transformed with animal genes that negatively regulate apoptosis, namely Bcl-xL, Ced-9 and Bcl-2 3' UTR, and independently transformed plant lines were regenerated for testing. Following a 12-week exposure to Foc race 1 in small-plant glasshouse bioassays, seven transgenic lines (2 × Bcl-xL, 3 × Ced-9 and 2 × Bcl-2 3' UTR) showed significantly less internal and external disease symptoms than the wild-type susceptible 'Lady Finger' banana plants used as positive controls. Of these, one Bcl-2 3' UTR line showed resistance that was equivalent to that of wild-type Cavendish bananas that were included as resistant negative controls. Further, the resistance of this line continued for 23-week postinoculation at which time the experiment was terminated. Using TUNEL assays, Foc race 1 was shown to induce apoptosis-like features in the roots of wild-type 'Lady Finger' plants consistent with a necrotrophic phase in the life cycle of this pathogen. This was further supported by the observed reduction in these effects in the roots of the resistant Bcl-2 3' UTR-transgenic line. This is the first report on the generation of transgenic banana plants with resistance to Fusarium wilt.     

Development of an in vitro hydroponic system for studying the interaction between banana plantlet and Fusarium oxysporum f. sp. cubense

Plant Cell, Tissue and Organ Culture (PCTOC) - Fusarium wilt of banana, caused by Fusarium oxysporum f. sp. cubense (Foc), is one of the most destructive diseases of banana. Several experimental...     

米修链霉菌TF78对香蕉枯萎病的田间防效及根际土壤微生物的影响

【背景】香蕉枯萎病是香蕉生产上的毁灭性病害,生物防治是遏制该病害发生的有效手段。在前期的研究中,从健康香蕉根际土壤中分离获得一株对香蕉枯萎病具有良好盆栽防治效果的生防菌——米修链霉菌(Streptomyces misionensis) TF78,但其对香蕉枯萎病的田间生防潜力和对土壤微生物环境的影响尚不清楚。【目的】评价米修链霉菌TF78对香蕉枯萎病的田间防治效果,明确其对香蕉根际土壤微生物群落的影响。【方法】选取两块发病香蕉园,测定该生防菌株对香蕉枯萎病的防治效果,并利用扩增子测序技术分析施用菌剂组和空白对照组共12份香蕉根际土壤的微生物多样性和丰度。【结果】米修链霉菌TF78对两块香蕉园的田间防效分别达55.30%和45.32%。该生防菌株处理组的物种稀释曲线坡度大于空白对照组,并显著富集了优势种群梳霉门(Kickxellomycota),消减了绿弯菌门(Chloroflexi)、酸杆菌门(Acidobacteria)和苔藓杆菌(Bryobacter)的丰度,对土壤中优势种群变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)、放线菌门(Actinobacteria)、芽单胞菌门(Gemmatimonadetes)及木霉属(Trichoderma)、鞘氨醇单胞菌属(Sphingomonas)、寡养单胞菌属(Stenotrophomonas)和芽孢杆菌属(Bacillus)的相对丰度影响不显著。【结论】米修链霉菌TF78塑造了不利于香蕉枯萎病菌Fusarium oxysporum f.sp.cubense存活的土壤环境,有效降低了田间香蕉枯萎病的发生,同时对土壤中大部分具有重要生态功能和抑菌功能的优势微生物种群影响不显著。该研究结果为米修链霉菌TF78的进一步开发应用奠定了基础。