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INTRODUCTION 1. Rationale Eggplant is a fruit vegetable and is high nutrient and economical value, developed and expanded in Vietnam as well as Lam Dong province. Although, eggplant is planted and took care easily but, disadvantages of eggplant production are detected by many diseases such as green wilt, leaf spot, Verticillium wilt and plant-parasitic nematodes. Root-knot nematodes (Meloidogyne spp.) are common pathogens parasitised on most plants worldwide. Among the nematodes discovered in Vietnam, root-knot nematodes is the most serious parasitism and thriving pest in recent years. In Lam Dong, root-knot nematodes is one of the major pest parasitised on solanaceae, in generally and on eggplant, as a result, decreasing of quality and yield, increasing other diseases. There have been several researches about root-knot nematodes parasitised on eggplant, however, very little has been conducted about distribution, as well as biological and ecological characteristics of root-knot nematodes on Solanaceae and eggplant in Lam Dong to control them. For these reasons, we carried out the topic "Research on biological and ecological characteristics of Meloidogyne sp. parasitised on eggplant and methods control by integrated nematodes management in Lam Dong" to (i) determine root-knot parasitised nematodes species composition on eggplant, (ii) study biological and ecological characteristics, and (iii) provide effective solutions for control root-knot nematodes toward integrated nematodes management in eggplant production in Lam Dong as well as in Vietnam. 2. Research aims and objectives Objectives: the objectives of the research are: (i) to identify species composition root-knot nematodes parasitised on eggplant, (ii) to determine biological and ecological characteristics of M.
incognita, and (iii) to provide proposing methods to prevent them toward integrated nematodes management, contributing production of eggplant and solanaceace safety, effectively and sustainably. 3. Significance Theoretical significance: The thesis has supplemented new scientific data of species composition of root-knot nematodes, biological and ecological characteristics, rules of reproduction, development and parasiticide of M. incognita. On that basis, effective methods and solutions to control root-knot nematodes, protecting the environment and contributing to safe production of eggplants safety in Lam Dong was proposed. The thesis is a reference for agricultural students, researcher, teacher and sciences. Practical significance: Proposing methods to control root-knot nematodes effectively and safely, contributing to improved process of the eggplant production toward integrated nematodes management for the purpose of stable, effective and sustainable production in Lam Dong as well as in Vietnam, particularly, chemical pesticides are abused to control nematodes today. This thesis is also a document to help managers and farmers identifying symptoms of M. incognita on eggplant and decide effectively management solutions of root-knot nematodes. 4. Subjects and scope of the research 4.1. Research subjects The subject of study is root-knot nematodes (Meloidogyne sp.) parasitised on eggplant. 4.2. Research scope Thesis content: Identifying species composition of parasitic root-knot nematodes on eggplant; Studying biological characteristics of M. incognita detected on eggplant; Accessing some of ecological conditions (soil type, organic fertilizers, rainfall, soil moisture, temperature 1
and different eggplant varieties) affected parasitic characteristics and population of M. incognita detected on eggplant; Providing solutions to control M. incognita detected on eggplant according to integrated management (cultivation methods, biological methods, physical methods, chemical methods). Location: Identifying species composition, affecting ecological conditions on root-knot nematodes and open field experiments were conducted in three vegetable growing areas in Don Duong, Duc Trong and Da Lat in Lam Dong province. Greenhouse experiments were conducted in the Department of Agriculture and Forestry, Dalat University. In vitro experiments including idetification of root-knot nematodes, extractation vermiform root-knot nematodes from soil and root, lilfe cycle of M. incognita were carried out in Plant Protection Laboratory, the Department of Agriculture and Forestry, Dalat University and laboratories of Nematology Department, Institute for Agricultural and Fisheries Research (ILVO), Flanders, Belgium. 5. Novel contributions of the thesis This thesis makes contributions: (i) To provide some new data of species composition of root-knot nematodes (Meloidogyne spp.), biological and ecological characteristics of M. incognita detected on eggplant in Lam Dong province. (ii) To propose methods to control root-knot nematodes detected on eggplant in Lam Dong according to integrated nematodes management to decrease damage; to reduce using chemical pesticides; to complete the protocol of eggplant production following safety and organic cultivation in Lam Dong as well as in Vietnam. CHAPTER 1. LITERATURE REVIEW 1.1 Overview Eggplant (Solanum melongena) is a fruit vegetable with high economic value. In over the world, there were about 1.6 million hectares growing eggplant. In Vietnam, eggplant is grown in recent years and becomes popularly in many areas throughout the country. In Lam Dong, eggplant was planted reaching about 1,944 ha, with an average yield of 47.6 tons/ha in 2017. Growing eggplant can bring high income but unstable, because eggplant is a susceptible crop to many diseases, insects and nematodes, in which, Meloidogyne spp. are an important group of plant parasites reduced eggplant yield and quality. Root-knot nematodes are the most important and common pest worldwide, causing economic damages of agricultural crops in tropical and subtropical areas. Root-knot nematodes were reduced eggplant yield up to 95% (Di Vito, 1986). There are many species of root-knot nematodes detected on eggplant, in which two species M. incognita, M. javanica detected in South Asia, Nepal and India, three species M. incognita, M. arenaria and M. javanica detected in Egypt. Until now, very few surveys have been conducted and evaluated the role of root-knot nematodes on crops in Vietnam, mainly focused on some industrial perennial plants. Some survey reported the species composition of root-knot nematodes on vegetables in South of Vietnam and Lam Dong provine in the 1990s. However, there is no report studying root-knot nematodes on eggplant. On over the world, there have been a number of studies involves the different ecological conditions such as temperature, humidity, climatic factors affecting life cycle of M. incognita on eggplant as well as control methods. However, in Vietnam, there are no research results of
biological and ecological characteristics as well as control methods root-knot nematodes on eggplant. 1.2 Research situation 1.2.1 Worldwide 22.214.171.124 Research history, distribution and damage thresholds of root knot nematodes in agriculture Root-knot nematodes are obligate parasites which can be found in varieties of plants play an important role in agriculture. They caused serious economic losses in agriculture. The earliest report of observation of plant parasitic nematodes was in the mid-18th century (1743) when Needham observed pests on wheat by microscope. However, until the middle of the 19th century, root-knot nematodes had been identified morphological characterictics clearly. Typical symptoms of root-knot nematodes are many galls on the root system. M. incognita, M. javanica and M. arenaria are the major of root-knot nematodes, in which, M. incognita is the most important plant parasitic nematodes. Like other plant pathogens, root-knot nematodes cause estimated crop yield decline. According to Taylor and Sasser (1978), for infected areas, without control methods, crop yield may be drop to 24.5% - 85.0%. 126.96.36.199 Classification and identificationy root knot nematodes Root-knot nematodes (Meloidogyne) belongs to kingdom Animalia, phylum Nematoda Potts, 1932; class Chromadorea Inglis, 1983; order Rhabditida Chitwood 933; suborder Tylenchida Thorne, 1949; family Meloidogynidae Skarbilovich, 1959; genus Meloidogyne Goldi, 1987. Initially, technique identification of root-knot nematodes is mainly based on morphological characteristics, describes perineal pattern characteristics of females and the length of secondstage juveniles. However, at the end of the 20th century, root-knot nematodes is identified by isozyme analysis and molecular identification methods. 188.8.131.52 Biological and ecological characteristics of root-knot nematodes There are many stages growth and development of root-knot nematodes changing in different shapes which could be called sexual dimorphism. Females are the pear-shaped, less moving into the root. The first-stage juveniles (J1) are within the eggs. The second-stage juveniles are worm shape, move in the soil. After infecting roots, the second-stage juveniles (J2) develop into the third juveniles (J3) and the four juveniles (J4), then swollen to adult females or form males moving in the soil. Usually, root-knot nematodes development depends on species, host and temperature. In general, life cycle is 15 days to 70 days. Root-knot nematode are detected on different varieties of crop with the different ways. The density of root-knot nematodes in soil depends on soil type and ecological climate conditions. Species composition and nematode density in soil are related to 65% of rainfall and 58% of soil temperature. Soil texture and structure also affects nematodes density. Sandy are more favorable than clay. The density of nematodes in soil is related to organic materials added to soil. In general, growth, density and survival of root-knot nematodes depend on the host, ecological conditions such as temperature, soil moisture, organic content and soil texture. 184.108.40.206 Methods control root-knot nematodes Using control methods aims to limit harmful effects of nematodes on crop, stabilize yield and quality, bring high economic efficiency. There are 6 control methods of root-knot nematodes to protect crops including: crop rotation, sanitation, resistant varieties, biological methods, physical methods and nematicide. Integrated nematode management is studied and implemented to efficiently control nematodes and reduce using nematicide. 3
1.2.2 Viet Nam 220.127.116.11 Research history, classification, distribution, biological and ecological characteristics of root knot nematodes Root-knot nematodes have been called many Vietnamese names. In Vietnam, the first nematode study was published by Hungarian scientist named Andrassy in 1970 with more than 30 species of plant parasitic nematodes and free living nematodes. According to Nguyen Ngoc Chau and Nguyen Vu Thanh (2000), plant parasitic nematodes were divided into 30 varieties, 11 families, 4 order. In particularly, root knot nematodes parasitised in Vietnam including 10 species of M. arenaria, M. cynariensis, M. graminicola, M. incognita, M. javanica, M. exigua, M. cofeicola, M. enterolobii, M. hapla and M. daklakensis which belonged to genus Meloidgyne, family Heteroderidae, suborder Tylenchina and order Tylenchida. Studies of investigating and classification of root-knot nematodes in Vietnam were mainly based on morphological methods. In 2005, identifying nematodes by molecular methods was first applied, but there was no results of Meloidogyne identification. In 2012, M. graminicola and M. incognita were identified by molecular methods. In 2018, Trinh et al. were identified new species called M. daklakensis based on morphological characteristics, genes and mitochondrial ADN-based identification. The study of root-knot nematodes was first published basing on biological characteristics of M. arenaria parasitised on rice, M. incognita, relationship between M. incognita, Rotylenchulus reniformis and Tylenchorhynchus brassicae and damage thresholds parasitised on tomato and tobacco. However, there are not any reports about biological and ecological characteristics of root-knot nematodes parasitised on eggplant. 18.104.22.168 Methods control root-knot nematodes In Vietnam, the first report method to control of root-knot nematode was in 1981, followed by reports about control methods for root-knot nematodes on black pepper and vegetable grown in Ha Noi. Control methods of root-knot nematodes in Vietnam were also carried out basing on the most popular methods on crops such as rice, tobacco, coffee, black pepper, Chinese cabbage and salad. Control methods include crop rotation, resistant varieties, physical methods, biological methods, chemical methods and integrated nematodes management. However, there have been publishing methods to control root-knot nematode parasitised on eggplant in Vietnam as well as Lam Dong. CHAPTER 2. METHODOLOGY 2.1. Research time Research period was from 2014 to 2017 2.2 Materials, equipments and instruments Materials were eggplant variety of TN525 Green King; sandy clay (49% sand, 10% limon and 41% clay) and clay (32% sand, 1% limon, 67% clay) and river sandy (70% sand, 20 % limon and 10% clay); nylon bags and pots; organic amendments and Yara NPK fertilizer 15-15-15; chemicals for specimens; sampling tools, sieve and laboratory equipment. 2.3 Research content - Conducting to survey, collect samples, identify species composition, recognize symptoms, to determine damage threshold and to observe density change of root-knot nematodes (Meloidogyne spp.) in open field parasitised on eggplant in Lam Dong. - Studying biological and ecological characteristics of M. incognita detected on eggplant in Lam Dong. 4
- Studying methods to control root-knot nematodes (M. incognita) following integrated nematode management. - Building protocol of integrated nematodes management (M. incognita) parasitised on eggplant in Lam Dong. 2.4 Research methods 2.4.1. Survey and identification of root-knot nematodes parasitised on eggplant in Lam Dong 22.214.171.124 Survey Investigating farmers was carried out by interviewing directly with semi-structured questionnaires. 126.96.36.199 Investigation of the main nematodes parasitised on eggplant in the field Samples were collected from 85 households. On the field, an random area was chosen about 100m2. Surveying from 5 points according to diagonal rules was to determine main pest and diseases in open field based on typical symptoms on the stem, leaf, fruit of eggplant (Daunay, (2008); Srinivasan (2009). 188.8.131.52 Sampling Soil and root samples were taken according to the W-pattern from 85 eggplant fields in Lam Dong. Samples were stored in a thermostatic cabinet at a temperature of 15oC. 184.108.40.206 Extraction root-knot nematodes from soil and root samples Activity second-stage juveniles were extracted from soil and root by modification Baermann funel. The sample was incubated in the funnel for 48 hours at room temperature. Activity second-stage juveniles were counted by stereo-microcopes with 4X magnification. 220.127.116.11 Determining the ratio of root damage and root gall-index Root gall-index was assessed by the degree of infection from 1-10 according to Zeck (1971), Bridge and Page (1980). Determining the incidence of root knot nematodes 𝑁𝑢𝑚𝑏𝑒𝑟 𝑜𝑓 𝑟𝑒𝑝𝑒𝑎𝑡 𝑓𝑖𝑒𝑙𝑑𝑠 𝑠𝑢𝑟𝑣𝑒𝑦𝑒𝑑 Incidence of root knot nematodes(%) = . 100 𝑇𝑜𝑡𝑎𝑙 𝑓𝑖𝑒𝑙𝑑 𝑠𝑢𝑟𝑣𝑒𝑦𝑒𝑑 Determining the frequency of species occurrence 𝑁𝑢𝑚𝑏𝑒𝑟 𝑜𝑓 𝑡𝑖𝑚𝑒𝑠 𝑒𝑛𝑐𝑜𝑢𝑛𝑡𝑒𝑟𝑒𝑑 𝑠𝑢𝑟𝑣𝑒𝑦 Frequency of species occurrence(%) = . 100 𝑇𝑜𝑡𝑎𝑙 𝑜𝑓 𝑡𝑖𝑚𝑒𝑠 𝑠𝑢𝑟𝑣𝑒𝑦𝑒𝑑 Determining ratio of root infestation 𝑁𝑢𝑚𝑏𝑒𝑟 𝑜𝑓 𝑟𝑜𝑜𝑡 𝑖𝑛𝑓𝑒𝑐𝑡𝑒𝑑 Ratio of root infestation (%) = . 100 𝑇𝑜𝑡𝑎𝑙 𝑜𝑓 𝑟𝑜𝑜𝑡 𝑠𝑢𝑟𝑣𝑒𝑦𝑒𝑑 18.104.22.168 Extraction with perineal pattern females Females of Meloidogyne were dissected out from the root galls by using forceps under compound microscope and transferred 20 females to 0,9% NaCl. 22.214.171.124 Cut perineal patterns Females were cut with a sharp knife and the body content was cleared away from the internal surface of the cuticle. The cuticle of the posterior third of the body was trimmed down to a small piece containing vulva, anus and tail tip. Perineal patterns were sealed with a coverslip by nail varnish 126.96.36.199 Making specimens How to make temporary and long-term specimens was been according to Bezooijen (2006) and Ravichandra (2010). 5
188.8.131.52 Examinations of morphological features Morphological features included the perineal pattern, stylet morphology, and distance from the base of the stylet knobs to the dorsal esophageal gland, length of second-stage juveniles, males, females by Bel-capture ruler. 184.108.40.206 Identification root-knot nematodes by molecular Root-knot nematodes were identified by molecular basing on primer of tropical root-knot nematodes group. 2.4.2 Biological and ecological characteristics of root-knot nematodes (M. incognita) detected on eggplant 220.127.116.11 Culturing and purifying species Root-knot nematodes (M. incognita) were cultured and purified on tomato roots in B5Gamborg semi-liquid media. 18.104.22.168. Studying morphological and biological characteristics (life cycle, ratio of hatching of M. incognita) in the laboratory Morphology characteristics and development stages of M. incognita were observed at temperature about 24±1oC with 3 different soil moisture at 30 - 40%, 40 - 50%, 50 - 60%. Roots were stained to describe morphology characteristics and development stages. Roots were stained by fuchsin acid. Ratio of hatching was determined by counting the number of the second-stage juveniles formed on the total of hatching eggs in 3 different environments including distilled water; extracts of 1-month eggplant roots; 6-month eggplant roots. 22.214.171.124 Studying ecological conditions affected to density of root knot nematodes Preparing free-disease plants Seeds are sterilized and sown on sterilized substrates in the greenhouse. Nematode inoculation in pots in the greenhouse The second-stage juveniles of M. incognita were extracted from tomato roots cultured in the laboratory. Initial population of M. incognita was inoculated about 2000 individuals/pot. Effect of soil moisture, rainfall, temperature on M. incognita in the field The experiment was conducted in the open field grown eggplant TN 252 Green King variety in Suoi Thong B village, Da Ron communes, Don Duong district in Lam Dong provine. Secondary data of temperature and rainfall during 2014-2017 was collected from meteorological stations of Thanh My, Don Duong and Da Lat. Soil moisture is determined by weigh method. The density of the second-stage juveniles was determined a 30-day period during three continuous seasons to assess correlation between the density of the second-stage juveniles of M. incognita and rainfall, temperature and soil moisture. Effects of soil conditions (particle composition and organic content in soil) on density of root-knot nematodes Analysis of soil texture Analysis of soil texture was deternined according to Bouyoucos (1962) and named based on soil triangle. Correlation of sand content and density of second-stage juveniles in the soil was assessed. Effect of different organic amendments on root- knot nematodes M. incognita parasitised on eggplant Experiments were carried out in the green house. Meterials included organic amendments such as chicken manure, pig manure, goat manure, cow manure, commercial organic fertilizer and eggplant “TN252 Green King”. Comparison between the 5 treatments with control was 6
carried out, each repeated 3 times in clay pots. Root-knot nematodes (M. incognita) were inoculated at 3 weeks planting. Soil and root samples were taken from 6 planting pots and mixed to collect mixture samples. Experimental measures included density second-stages juveniles in soil and roots, ratio of root galls, number of galls, flowers, fruits and eggplant yield. Evaluation of eggplant varieties on root-knot nematodes The experiment was carried out in pots, in sandy clay, including six eggplant varieties including Thailand No.1, TN252 Green King, F1-033 local, black fruit NV123, Runako and Com Xanh, repeated 3 times. Inovation second-stages juveniles of M. incognita after 3 weeks planting. Soil and root samples were mixed from 6 pots to a mixture sample. Experimental measures were density of the second-stages juveniles in soil and roots, root-gall index, number of flowers, fruits and ratio of fruiting. 2.4.3 Control methods root-knot nematodes (Meloidogyne incognita) detected on eggplant following by integrated nematodes management in Lam Dong These experiments was conducted using eggplant ‘TN 252 Green King’ in the open field in Da Ron, Don Duong in Lam Dong. The experiments were repeated 3 times. Planting density was about 17,000 plants per hectare. Care and irrigation were the same in all treatments. Experimental measures included density of the second-stage juveniles in soil at the time of 30 days, 60 days, 90 days planting and the end of harvesting; density of the second-stages juveniles in roots; effective control of root-knot nematode calculated by Henderson - Tilton formula; ratio of root infected, root gall index and eggplant yield. 126.96.36.199 Cultivation methods to control root-knot nematodes Experimental treatments included: (i) crop rotation between eggplant with tung ho, cabbage, chili peppers, sweet corn, (ii) intercropped with the common bean Phaseolus vulgaris and eggplant. 188.8.131.52 Physical methods to control root-knot nematodes Experimental treatments included: (i) soil solarization (after harvesting the previous crop, plowing and drying the soil for 4 weeks); (ii) Keep soil dried (after harvesting the previous crop, plowing, every 7 days upsetting soil, keep soil dry out and soil moisture lower than 35% in 4 weeks); (iii) Covered with dark plastic (after harvesting the previous crop, deep plowing at least 25cm, irrigating and covering by plastic for 4 weeks); (iv) Burning soil (after harvesting the previous crop for 2 weeks, plowing the soil, making rows, covering a thin layer of 5-7cm husk into the planting groove, then a layer of 5cm thick soil, burning anaerobic, irrigating to soil moisture after 48 hours); and (v) Control (after harvesting the previous crop, did not using any method in soil, keeping for 4 weeks, plowing soil and making row for ready to plant). 184.108.40.206 Biologicial methods to control root-knot nematodes The experimental treatments included: Jianon Chitosan super (Chitosan); Vineem 1500 EC (neem - azadirachtin); Abuna 15GR (saponin); Biosune one (Trichoderma harzianum, Trichoderma viride and other microorganism) and control. 220.127.116.11 Chemiscial methods to control root-knot nematodes The experimental treatments included: Tervigo 020SC (Abamectin), Cazinon 10GR (Diazinon), Vifu-super 5GR (Carbosulfan), Map Logic 90WP (Clinoptilote) and control. 2.4.4 Building a model of control root-knot nematodes (Meloidogyne incognita) according to integrated nematodes management detected on eggplant in Lam Dong The previous crop was Tung Ho. After harvesting, cleaning the fields, plowing and drying for 4 weeks, then making rows. Cow manure putting down with an amount of 40m3/ha were 7
incubated with 10 kg/ha probiotics containing T. harzianum. Irrigating and using plastic covered on rows, keeping them for 7 days to allow fungi multiplied biomass rapidly, then planting after 7 days. After 10 days planting, eggplant was putted down chicken manure with amount of using about 300kg/ha. The area of the model was 500 m2 compared to farmers' handling. Evaluation of economic efficency of the model and control methods Calculation of production costs included costs for fertilizers, pesticides and labor. Calculating total income based on yield and price which was assumed about 2,000 VND per kilogram (price assumed if profit was the lowest), the efficient economic was calculated as following: Total income (thousand VND) = selling price per kg x yield Total costs = cost of seed + cost of labor + cost of fertilizer + cost of plant protection + cost of irrigation Net income = Total income - Total costs 2.5 Data analysis Data were collected and analysed of variance (ANOVA) by Microsoft Excel 2013 software, IBM SPSS Statistics Version 22 software and MSTATC. The mean comparison was done through LSD test and Duncan test with significance level of ≥95% (p≤0.05) CHAPTER 3. RESEARCH RESULTS AND DISCUSSION 3.1 Cultivation methods and control nematode parasitited on eggplant of farmers in the studied area 3.1.1 Survey of traditional cultivation of eggplant farmer in the studied area Cultivation and control methods root-knot namtodes detected on eggplant of farmers in the studied area Table 3.1&3.3 The area and yield of eggplant varieties were grown widely in open field (Lam Dong, 4/2014-6/2017) Eggplant varieties Ratio Density Yield Infestation Yield Season of of (plants/ha) (tons/ha) level (%) losses the highest grown (%) nematode (%) populations TN252 Green King 56.47 17,000 92.72 79.16 27.13 Rainy season Runako 12.94 25,000 35,90 63.63 20.37 Rainy season Oval fruit Japanese 11.76 25,000 36,60 48.00 20.41 Rainy season Thailand No.1 8.24 19,000 74.43 71.43 25.34 Rainy season Black fruit VN123 5.88 22,000 72.00 40.00 15.11 Rainy season F1-033 local variety 3.53 19,000 78.33 33.33 Trivial losses Com Xanh 1.18 25,000 35.00 0 0 Rainy season There was 7 varieties planted in Lam Dong, TN252 Green King was the most planted variety, accounting for 56.47% of the total, average yield was 92.72 tons/ha and density was 17,000 plants/ha, followed by the Runako variety accounted for 12.94% of total, average yield was 35.90 tons/ha, density was 25,000 plants/ha. The lowest ratio belonged to Com xanh variety, just only 1.18% and the yield was just about 35.00 tons/ha. Only 34.11% farmers checked pest and disease status of nursery stock and only checked the surface parts such as fungal disease on leaves or stems. If the nursery stock was not managed 8
soil well, crops would suffered from plant parasitic nematodes, then spread nematodes to the field. The variety of TN252 Green King was the highest ratio of plant detected by nematodes (79.16%), followed by Thai No.1 variety (71.43%), next to Runako (63.63%) and the lowest ratio was the Com Xanh (0%, n = 1). Meanwhile, variety TN252 Green King showed the highest yield losses (27.13%), variety Thailand No.1 (25.34%). Local variety and Com Xanh yield were decreased negligibly. Soil treatment and eggplant parasitic nematode control methods of farmer Table 3.4 Treatment of soil by farmers in Lam Dong (4/2014-6/2017) Treatment methods Ratio application (%) Dose Lime 100.0 950 kg/ha Chemical methods 62.35 Biological methods 5.88 Physical methods 67.06 Only used lime 11.76 All surveyed farmers used lime before planting to treat soil for controlling pathogens and some insects caused disease of eggplant, accounted for 100% for surveyed farmers. Chemical methods were also commonly used, accounting for 62.35%. Commercial chemicals were used such as Nokaph 10GR (30.82%), Map Logic 90WP (5.88%), Basudin 10% granular (8.0%) and Binhtox 1.8EC (17.64%). Up to 67.06% of surveyed farmers used physical methods such as fallow, did not plow, covering with dark nylon. Only 5.88% of surveyed farmers used biological methods for controlling disease in soil. About 11.76% of surveyed farmers did not use any treatment methods except liming. According to farmers, chemical methods were the most effective brought the highest profits. 76.46% of surveyed farmers recognized symptoms of stunted plant, roots galls, low uptake nutrition, they changed the ways to provide fertilizer for eggplant by diluting fertilizer then irrigating directly to roots or spraying on the foliar instead of putting down into soil as before. Only 5.88% of surveyed farmer using Tervigo 020SC sprayed on their roots to control root parasitic nematode.
Fertilizers used and feritilizer dosage used in eggplant cultivation Table 3.6 Using fertilizer of eggplant farmers (Lam Dong, 4/2014-6/2017) Type of Ratio of Dose Frequency Ways to use fertilizer use (%) Manure 100.00 Before planting Scattering between rows 0.3 - 1.0 Every ten days Scattering around root tons/ha NPK 100.00 0.2 Every ten days Irrigating around root after 5 tons/ha months planting Foliar 94.11 3 l/ha Every month, after Spraying on leaves after 6 fertilizer 60 planting months planting Nitrogen 94.11 0.1 - 0.2 Within 1 - 2 months Scattering around root tons/ha after planting Phosphorus 100.00 0.5 - 0.7 Once before Scattering around root tons/ha planting Potassium 100.00 0.1 - 0.2 Basal fertilizer, Scattering around root and tons/ha harvest period between rows Survey results show that 100% of eggplant farmers used organic fertilizers and NPK, phosphate and potassium fertilizers. However, fertilizer types and dosage were different. Up to 94.11% of surveyed farmers used foliar and nitrogen fertilizers to put down for eggplant. 3.1.2 Symptoms and the major eggplant pests and diseases in Lam Dong Common eggplant diseases included green wilt, Verticillium wilt, anthracnose and mosaic disease. All diseases exposed symptoms on leaves, fruits, but none symptoms on roots and none galls. Eggplant insects included leaves and fruit insects and thrips palm. These insects damage on leaves, stems and fruits of plants, but not detected in root system. There were seven genus nematodes infected eggplant root system including Helicotylenchus, Tylenchus, Meloidogyne, Criconemella, Pratylenchus, Rotylenchulus and Longidorus. In which, Helicotylenchus were encountered with 100% of eggplant fields. Next, genus of nematodes was Tylenchus, achieved 91.76%, following Meloidogyne with 83.52%. Longidorus was the least parasitic nematode only 2.35%. The highest nematode density belonged to Meloidogyne (667 individuals/50cm3 soil), followed by Helicotylenchus with 570 individuals/50cm3 soil. Density of Rotylenchulus was the lowest with only 16 individuals/50cm3. This proved that root-knot nematodes played an important role for yield losses. Typical symptom of root-knot nematodes was the appearance of numerous galls on the roots. With the serious damage, galls were enlarged, even forming large swollen on the main roots. Symptoms of root-knot nematodes above-ground were differently recognized, especially when plants were young. When eggplant was 3-4-month age, root gall index was approximately 6, symptoms were recognized by stunted, yellow leaves, small and fewer fruits. Below-ground symptoms exposed in the period of about 6-8 months after planting. This was the time that eggplant have been ready for harvest, growth and resistance of the plant were reduced, root system was old, less forming new roots and lower uptake nutrition.
3.1.3 Density of root -knot nematodes (Meloidogyne spp.) detected on eggplant in Lam Dong Table 3.9 Second-juvenile stages density of Meloidogyne sp. in soil and roots and infestation level (Lam Dong, 4/2014-6/2017) Density of second-juvenile stages Average of Average in Average in year rainy season dry season 3 Density of J2 in soil (individual/50cm in soil) 1455 1750 827 Density of J2 in roots (individual/5 g root) 728 939 331 Damage level according to farmer estimated (%) 4.04 5.13 2.85 Yield losses (%) 12.41 14.89 7.19 During the year, density of second-juvenile stages in soil was 1455 individuals/50cm3, in roots was 728 individuals/5g of roots. In the dry season, nematodes density was lower than average of year, the density of the second-juvenile stages was just only 827 individuals/50cm3 in soil lower than 628 individuals/50cm3 in soil that of average year and 331 individuals/5 g roots in roots lower than that of 397 individuals/5g in roots of average year.
Figure 3.12 Correlation between damage thresholds and yield crop (Lam Dong, 4/2014-6/2017) Damage level of root knot nematodes was higher, eggplant yield losses was more, they have been non-linear correlation according to the cubic equation (y = 0.02 + 3.09x-2.41x2 + 0, 49x3). In eggplant field, if root gall index of root knot nematode was 1-3, the yield would be not reduced. When symptoms of root-knot were recognized visibly on roots such as stunting, fertilizer was used much higher, flowers and fruits would be reduced, yield losses would be higher. Meanwhile, eggplant roots were more than 60% of root galls, approximately 6 root-gall index, estimated yield losses up to 40 - 50%. Density of the second-stage juveniles of root-knot nematodes in soil ranged from 500 individuals/50cm3 to 1500 individuals/50cm3 in soil. In the 71 surveyed samples, there were 6 soil samples with density of second-stage juveniles ranged of 2000 - 3000 individuals/50cm3, 4 soil samples with density higher than 3000 individuals/50cm3.
3.1.4 Surveying, collecting and identifying root-knot nematodes (Meloidogyne spp.) detected on eggplant The surveyed results showed that, root-knot nematodes were detected in the 83.52% of the soil and root samples collected. There were 2 species presented in the 71 soil and root samples. Morphological characteristics were described. 48 out of 71 collected samples were the same morphological characteristics of Meloidogyne incognita, 17 out of 71 collected samples were the same species of M. javanica, 6 samples were occurred mixture of the two species.
Figure 3.18 The frequency of Meloidogyne species (percentage) in Lam Dong (4/2014 - 6/2017) The surveyed results of root-knot nematodes species showed that occurence of M. incognita was 67.71%, followed by M. Javanica, accounted to 23.94% and mixture of the two species only 8.45%. Table 3.12 Incidence of root knot nematodes, root gall index and density of root-knot nematodes detected on eggplant (Lam Dong, 4/2014-6/2017) Location study Incidence Root gall Density of J2 Root-knot nematodes species (%) index individials/50cm3 Don Duong 88.09 4.91 789 M. incognita, M. javanica Duc Trong 84.85 4.03 547 M. incognita, M. javanica Da Lat 60.00 3.12 356 M. incognita, M. javanica In of three studied location, the highest incidence of root-knot nematodes was in Don Duong (84.85%), the lowest was in Da Lat (only 60%). The density of second-stage juveniles in soil and root gall index were the highest in Don Duong, with 789 individuals/50cm3 and 4.91, respectively, otherwise, the lowest figures were in Da Lat, proved by 356 individuals/50cm3 and 3.12 respectively. All of three studied areas, two species occurred were M. incognita and M. javanica, but infectious fields in Don Duong and Duc Trong with the mixtures of M. incognita and M. javanica was higher than that in Da Lat. Identification results showed that DNA samples were suitable for M. ethiopica, M. arenaria, M. enterolobii and M. javanica on eggplant. 4 samples were acceptable Mi2F4/Mi1R primers, M. incognita were identified. Generally, 71 out of 85 collected samples were found Meloidogyne species with M. incognita and M. javanica. Out of two, M. incognita was more frequent played an important damage role with 67.61%, M. javanica was lower frequent with 23.94%. Mixture of them was only 8.45%. 3.2 Biological and ecological characteristics of Meloidogyne incognita 3.2.1 Morphological characteristics of Meloidogyne incognita Life cycle of M. incognita undergwent the first moult in the eggs to develop to the firststage juveniles (J1) to the second-stage juveniles (J2) to third-stage juveniles (J3) to fourth-stage 12
juveniles (J4) then to mature stage (adult females and males). Specific characteristics of them were showed as follows: Eggs: Egg shape was oval, contained in egg mass of the female. Eggs were laid by females and usually found on the surface of galled roots sometimes inside plant tissue. The first-stage juveniles: embryogenesis proceeds to the first-stage juveniles with a blunt tail tip, vermiform, curl up which mounted one in the egg. Then, they developed to second-stage juveniles depended on environmental condition. The second-stage juveniles: Second-stage juveniles of M. incognita were movable vermiform, tapered head with stylet. The tail part is transparent. The third-stage juveniles: Under favorable environment, J2 hatched in roots to J3 which were tapered at the end of tails, bulged in the middle. Their head were without stylet, short tail. The fourth-stage juveniles: They were avocado shape, lack a functional stylet, short - small clear tail. They did not feed. Adult stage Females: Adult females were pear shape, sedentary, head with stylet, feed roots. Males: Male were mobile, remain vermiform, leaved galls and entered soil. They have spicules at the end of the tail. Stylet of male is blunt, set off. Table 3.13 Size of some stages of M. incognita Grown stage Length (µm) Width (µm) Stylet length (µm) DEGO (µm) Eggs 77.77 32.15 Second-stage juveniles 385.13 10.64 2.62 Females 627.17 404.17 15.34 2.90 Males 1595.53 26.95 2.50 Note: DEGO (dorsal esophageal gland orifice) Morphology of M. incognita has undergone different stages of development. Egg stage was the smallest size. The second-stage juveniles, females and males were significant differences length, width, tail, stylet and dorsal esophageal gland orifice (DEGO). The second-stage juveniles and males were vermiform, but the length of males was over 5 times than that of the second-stage juveniles, males stylet was over 2.5 times than that of the second-stage juveniles, DEGO of males was lower than that of the second-stage juveniles. Females length was shorter than that of males, longer than that of the second-stage juveniles. 3.2.2. Biological characteristics of Meloidogyne incognita 18.104.22.168 Life cycle, characteristics of the sexual phases of M. incognita Life cycle of M. incognita started the second-stage juveniles infected into root, underwent some moults such as the third-stage juvenile, the fourth stage juveniles and adults, then hatched a new infectious second-stage juveniles when its life cycle completed. Table 3.14 Development time and ratio of root inoculom of the second-stage juveniles Experiment Development time (days) Soil moiture Ratio of root (%) inoculum(%) Shortest Longest Everage ns c 1 7 9 8.04 36.13 85.68c b 2 7 9 8.00 46.77 95.46a 3 7 9 8.04 57.11a 93.31b Average 8.04 46.67 91.48
Development time of the second-stage juveniles in eggplant roots averaged 8.01 days and there was no difference between different moisture thresholds. Ratio of root inoculum of the second-stage juvenile was different at three soil moisture thresholds. At the average soil moisture (36.13%), ratio of root inoculum of the second-stage juveniles was the lowest, only 85.68%, the highest at the soil moisture of 46.77% reaching 95.46%, soil moisture of 57.11% achieved 93.31% . Table 3.20 Life cylce of M. incognita Development stage Time of development stage (days) Average (days) Experiment 1 Experiment 2 Experiment 3 Second-stage juveniles 8.04 8.00 8.04 8.03 Third-stage juveniles 4.00 4.00 3.93 3.98 Fourth-stage juveniles 2.13 2.07 2.20 2.13 Females 8.00 8.00 8.07 8.02 Egg and first-stage juveniles 5.27 4.80 4.93 5.00 Life cycle 27.44 26.87 27.17 27.16 Males 19.00 20.00 19.00 19.25 o Temperature ( C) 24±1 24±1 24±1 24±1 Soil moiture (%) 36.13 46.77 57.11 46.67 In three different soil moisture at 36.13%, 46.77% and 57.11%, the average development time of the third-stage juveniles was 3.98 days, the fourth-stage juvenile was only 2.13 days, adult female was 8.02 days. Time of development from fourth-stage juveniles to male was 19.25, there was not significant difference among 3 different soil moisture. Life cycle of the M. incognita root knot nematode was 27.16 days at temperature at 24±1oC and soil moisture at 46.67%. The development time of M. incognita did not differ significantly between 3 different soil moisture. Development time of the second-stage juveniles and females were the longest with 8.03 and 8.02 days, respectively. Development time of the fourth-stage juveniles was the shortest with only 2.13 days, followed by the third-stage juveniles was 3.98 days while eggs and the first-stage juveniles was for 5 days. Development time from the fourthstage juveniles to males was 19.25 days. Time from the second-stage juveniles to males at conditions of 24oC ± 1oC, 46.67% was 33.39 days. 22.214.171.124 Ratio of hatching egg of M. incognita Table 3.21 Effect of environment condition to ratio of eggs hatch of M. incognita STT Environment for eggs hatch Ratio of hatching (%) 1 Distilled water 69.30c 2 Root extraction of one-month root 90.72a 3 Root extraction of over-month root 83.00b The eggs hatch ratio of M. incognita was the highest in extract of one-month eggplant roots, reaching 90.72%, followed by in extract of six-month eggplant roots with 83.00% and much higher than that of distilled water with only 69.30%. 3.2.3 Effect of ecological conditions on Meloidogyne incognita 126.96.36.199. Effect of soil texture on density of the second-stage juveniles in soil
Table 3.22 Effect of soil-particle on the second-stage juveniles detected on eggplant in Lam Dong (4/2014-6/2017) Clay content Limon content Sand content (%) Soil types Density of J2 (%) (%) (individuals/50cm3) 54.00 28.54 17.46 Sand 222 40.99 40.80 18.21 Silty clay 365 18.72 58.95 22.33 Silty loam 513 35.99 32.19 31.82 Clay loam 547 23.14 37.74 39.12 Loam 593 29.13 19.39 51.48 Silty clay loam 782 38.53 8.25 53.22 Sandy clay 891 17.34 21.05 61.61 Sandy loam 1735 Correlattion (r) between of sand and J2 0.730 Soil types affected differently on density of the second-stage juveniles. While clay and limon content were not correlated with density of second-stage juveniles in soil, sand content was correlated with density of the second-stage juveniles. Out of all soil types, density of the secondstage juveniles in clay was the lowest, average of 222 individuals/50cm3 in soil, followed by silty clay with average density of 365 individuals/50cm3 in soil, silty loam with 513 individuals/50cm3 in soil. Density of the second-stage juveniles in sand loam was the highest, achieved 1735 individuals/50cm3. Density of the second-stage juveniles in loam was lower than in sand but higher than in clay. 188.8.131.52. Effect of rainfall, air temperature and soil moisture on M. incognita in soil grown eggplant in Lam Dong Table 3.23 Correlation between soil moisture, air temperature and rainfall to the density of M. incognita in Lam Dong (6/2014-5/2017) Density of J2 Soil moisture Rainfall Temperature 3 (individual/50cm ) (%) (mm) (oC) Density of r 1 0.672* 0.678* 0.516 J2 Sig. (two ways 0.017 0.015 0.086 (individuals/ factor) 50cm3) N 12 12 12 12 * ** Soil r 0.672 1 0.812 0.598* moisture Sig. (two ways 0.017 0.001 0.040 (%) factor) N 12 12 12 12 * ** Rainfall r 0.678 0.812 1 0.687* (mm) Sig. (two ways 0.015 0.001 0.014 factor) N 12 12 12 12 Temperature r 0.516 0.598* 0.687* 1 o ( C) Sig. (two ways 0.086 0.040 0.014 factor) N 12 12 12 12 *. Correlation at p= 0.05 (two ways factor) **. Correlation p=0.01 (two ways factor)
There was a close linear correlation between three factors: rainfall, soil moisture and temperature and also between these factors and the density of the second-stage juveniles in soil. Rainfall and soil moisture have a very close correlation (r = 0.812). Soil moisture, rainfall are correlated with density of the second-stage juveniles in the soil (0.5 0.678, respectively. Soil moisture and rainfall were close correlation with density of the secondstage juveniles in soil. The results proved that soil moisture and rainfall affected on density of second-stage juveniles of root-knot nematodses in soil. 184.108.40.206 Density variation of M. incognita in a growing season of eggplant in Lam Dong
Figure 3.33 Density of the second-stage juveniles of M. incognita in a growing season of eggplant in Lam Dong (crop 2015 - 2017) At 30 days before planting, density of M. incognita was low only 225 individuals/50 cm3 in soil, but then, at the time of planting, density of second-stage juveniles increased. They increased fastly at 30 days and 60 days after planting, achieved 3051 and 2975 individuals/50cm3 in soil, respectively. Then, density of root knot nematodes decreased sharply, tended to stabilize at 90 days, 120 days and 150 days after planting. 180 days after planting, density of M. incognita in soil gradually decreased. At the end of the crop, density of root-knot nematodes in soil is reduced to 476 individuals/50cm3. 220.127.116.11 Effect of different organic amendment on root knot nematodes detected on eggplant Table 3.24 Effect of organic amendment on the second-stage juveniles (J2) M. incognita in soil, roots, root knot nematodes, ratio and number of galls detected on eggplant (Lam Dong, 20142015) Treatments Density of J2 Density of J2 in Ratio of root Number of in soil root infestation galls (galls/ (individuals/50cm3soil) individuals/g) (%) root) cd c c Chicken manure 1634.0 266.3 93.93 7.67 c Pig manure 2967.3 a 1055.0 b 100.00 a 18.67 b c b ab Goat manure 1859.3 1043.0 98.53 15.67 b Cow manure 1360.7 d 1377.0 b 100.00 a 19.00 b Organic fertilizer (%) 1825.3 c 309.7 c 95.27 bc 8.67 c Control 2562.0 b 1989.7 a 100.00 a 22.67 a
Density of second-stage juveniles of M. incognita in soil was the lowest in cow manure treatment (1360 individuals/50cm3 soil), followed by chicken manure (1634 individuals 50cm3 soil), organic fertilizer 73% (1825 individuals/50cm3 soil), goat manure (1859 individuals/50cm3 soil) and the highest density of second-stage juveniles was in pig manure treatment (2967 individuals/50cm3 soil). In roots, density of second-stage juveniles was the lowest in chicken manure treatment (266 individuals/5g roots) and commercial organic fertilizer 73% organic content (309 individuals/5g roots). Ratio of root infestation in chicken manure treatment and organic fertilizer treatment were significant difference from other treatments. Number of root infestation was the highest in the control (22.67 galls/root) and significant difference from in pig manure (18.67 galls/root), goat manure (15.67 nodules/roots) and cow dung (19.00 notes/root). number of galls in chicken manure treatment and commercial organic fertilizer treatment were the lowest, not statisticial difference, only 7.67 galls/root and 8.67 galls/roots, respectively. 18.104.22.168 The reaction of eggplant varieties with root knot nematodes Table 3.25 Density of the second-stage juveniles of M. incognita in soil (Lam Dong, 2017) Density of second-stage juveniles in soil (individuals/50cm3 soil) Varieties Inoculated 20 NSN 40 NSN 80 NSN 100 NSN ED ab c a Thai Lan No.1 2000 577 1095 1431 1955 a 1123 ab TN 252 Green king 2000 727 a 2433 a 1324 a 1697 abc 1285 a ab b b c F1-033 local variety 2000 561 1524 1044 1443 1179 ab Black fruit NV123 2000 326 bc 971 c 1262 a 1763 ab 1023 b Runako 2000 241 c 982 c 1062 b 1563 bc 1253 a Com Xanh 2000 357 bc 945 c 1344 a 1615 bc 1259 a Note: NSN: days after inoculating; ED: end of crop. Second-stage juveniles density decreased at 20 days after inoculating and increased at 40 days, 80 days, 100 days after inoculating. Density of the second-stage juveniles in soil was highest in TN252 Green King, followed in Thai No.1, balck fruit NV123. It was the lowest in Runako and Com Xanh. Table 3.26 & 3.27 Density of M. incognita in roots, infestation level at 150 days after inoculation, number of flowers, fruits and proportion of fruiting (Lam Dong, 2017) Varieties Density of J2 in root Root gall Flowers Number Ratio of (individuals/5g roots) index of fruits fruiting (%) ab a b Thai Lan No.1 1228 5.67 14.33 11.00 b 78.11 a a a ab b TN 252 Green king 1348 6.67 18.00 10.33 57.80 b F1-033 local variety 939 d 4.00 b 15.67 b 11.67 b 76.90 a Balck fruit NV123 1008 cd 5.67 a 13.67 b 10.00 b 71.54 ab Runako 1121 bc 5.67 a 22.67 a 16.67 a 75.68 a cd a b b Com Xanh 981 6.33 13.33 10.67 79.04 a Density of M. incognita in roots and root galls index in F1-033 local varieties were the lowest. Otherwise, density of second-stage juveniles in roots and root galls index in TN252 Green King were the highest but not significant difference from Thai Lan No.1. The highest flowers in Runako (22.67 flowers/plant) was not significant difference, compared to TN252 Green King (18.00 flowers/plant). The average numbers of fruits in Runako was also the highest (16.67 fruits/plant) completely different from other varieties. Ratio of fruiting of Com Xanh (79.04%), Thailand No.1 (78.11%), local variety (76.79%) and Runako (75.68%) were higher than that of TN252 Green King. 17
3.3 Integrated nematodes management of root knot nematode in open field 3.3.1 Cultivation methods to control root knot nematode detected on eggplant Table 3.28 Effect of cultivation methods on density of second-stage juveniles of M. incognita detected on eggplant in Lam Dong (2014-2016) Treatments Density of second-stage juveniles Efficiency (%) in soil (individuals/50cm3) TT 30N 60N 90N ED 30N 60N 90N ED d a a Rotation with Tung Ho 631 607 689 816 462 5.73 22.71 34.03 6.82a c Rotation with Chinese 822 767 1015 1927 590 8.56 12.60d -19.59d 4.46c cabbage Rotation with chili 630 975 1067 1601 441 -51.65e -19.88e -29.64e 2.54d pepper Intercroping with 805 624 922 1420 576 24.04a 18.93b 10.01c 4.75b French bean Rotation with sweet 958 762 1164 1261 686 22.06b 14.00c 32.85b 4.69b corn Eggplant 780 796 1102 1529 586 LSD0,05 0.765 0.765 0.399 0.2113 Note: TT: before planting; N: days after planting; ED: end of crop
Density of second-stage juveniles in soil in most of experimental treatments decreased dramatically at 30 days after planting, increased at 60 days and at 90 days after planting then decreased at the end of harvest. Rotation eggplant with different crops showed that density of the second-stage juveniles in soil decreased significantly at 30 days planting. The highest efficiency in Tung Ho treatment was 34.03%, followed by sweet corn reaching 32.85% and intercropping with French beans 24.04%. Crop rotation of eggplant with chlili pepper has negative efficiency at 30 days, 60 days and 90 days planting. The results indicated that rotation eggplant with chili peppers was not effective for control root-knot nematodes (M. incogonita) detected eggplant. Bảng 3.29. Effect of cultivation methods on root infection of root knot nematode detected eggplant in Lam Dong (2014-2016) Treatments Density of J2 in root Ratio of root Root gall Yield (individuals/5 g) infestation (%) index (tons/ha) Rotation with Tung Ho 489e 37.13e 2.67e 109.00a Rotation with Chinese 1032b 69.29b 4.33b 104.30ab cabbage Rotation with chlili 1136a 73.40a 4.67a 93.00c peper Intercroping with French 833d 47.94d 3.97c 102.30b bean Rotation sweet corn 933c 49.83c 3.67d 103.00b a a b Specializing eggplant 1111 72.15 4.33 94.67c LSD0,05 35.03 1.311 0.2905 3.651 Density of second-stage juveniles in roots was the highest in rotation between chili pepper treatment and eggplant (1136 individuals/5g roots), decreased slightly to 1111 individuals/5g roots in specializing eggplant treatment, the lowest figure belonged to rotation between Tung Ho and eggplant (498 individuals/5g roots). The ratio of root infestation and root gall index were the lowest in the rotation between Tung Ho and eggplant with 37.13% and 2.67, respectively. 18
Summary, the rotation eggplant with Tung Ho reduced density of root knot nematodes in roots, ratio of root infestation, root gall index and increased yield of eggplant, followed by intercropping with French beans, rotation with sweet corn and Chinese cabbage. Rotation between chili peppers and eggplant and specializing eggplant increased ratio of root infestation, root gall index and reduced eggplant yield. 3.3.2. Physical methods for control root-knot nematodes dettected on eggplant Table 3.30 Effect of physical methods on density of M. incognita detected on eggplant in Lam Dong (2014 - 2015) Density of second-stage juveniles in Efficiency (%) soil (individuals/50cm3) Treatments TXL 30N 60N 90N ED 30N 60N 90N ED d b a Soil 1232 1315 1667 2560 1164 3.15 4.06 4.09 4.40c solarization Keeping soil dried 1321 1086 1862 2723 1195 25.40c 0.06c 4.87a 8.47b Covering plastic 1329 902 1617 2858 1175 38.41b 13.73a 0.74b 10.54a Burning 1389 760 1883 2996 1370 50.35a 3.88b 0.44b 0.20d Control 1116 1230 1574 2418 1103 LSD0,05 3.245 1.88 1.286 4.147 Note: TXL: before applying; N: days after applying; ED: end of crop
The density of the second-stage juveniles in soil decreased at 30 days planting, then increased at 60 days and 90 days planting and fell sharply at the end of the crop. After 30 days of treatment, the most efficiency of root knot nematodes was in burning method, reaching 50.35%, followed by covering plastic, reaching 38.41%. The lowest efficiency was in soil solarization only 3.15%. Table 3.31 Density of second-stage juveniles (M. incognita) in root, ratio of root infestation, root gall index and eggplant yield in lam Dong (2014-2015) Density of J2 Ratio of root Root gall index Yield Treatments in root infestation (%) (tons/ha) ab b bc Soil solarization 611 62.03 5.67 102.70a Keeping soil dried 512b 59.40b 6.00bc 103.30a a b b Covering plastic 758 63.37 6.33 102.30ab Burning 484b 53.73b 5.33c 98.33b Control 735a 81.67a 8.00a 103.30a LSD0,05 167.6 10.14 0.977 4.256 Density of the second-stage juveniles in roots was the highest in covering plastic treatment and control, 758 individuals/5g roots and 735 individuals/5g roots, respectively. The figure was the lowest in burning treatment (484 individuals/5g roots), different from soil solarization (611 individuals/5g roots) and keeping soil dried (512 individuals/5g roots). The ratio of root infestation and root gall index of control and covering plastic treatment were higher than that of burning and soil solarization. Yield of using burning method (98.33 tons/ha) was lower than keeping soil dried (103.30 tons/ha and control (103.30 tons/ha).
3.5.3 Biological methods to control root-knot nematodes detected on eggplant Table 3.32 Effect of biological methods on density of root-knot nematodes detected on eggplant in Lam Dong (2014-2015) Treatments Density of J2 in soil (individuals/50cm3) Efficiency (%) TXL 30N 60N 90N ED 30N 60N 90N ED Chitosan 382 570 988 845 1549 -24.73 -17.40 41.77 -108.75 Vineem 835 649 1049 1067 1098 35.03 42.97 66.36 32.30 Abuna 511 1640 789 1083 1088 -168.28 29.91 44.21 -9.61 Biosun 584 662 557 1744 1592 5.24 56.71 21.39 -40.34 Biofumigation 714 481 1199 1983 1401 43.69 23,77 26.89 -1.01 Control 591 707 1302 2245 1148 Note: TXL: before treatment; N: days after applying; ED: end of crop
Density of M. incognita decreased at 30 days, 60 days and 90 days after applying in all treatments comparing to the control. Efficiency of Jianon chitosan super and Abunar 15GR treatment were negative efficiency after 30 days applying and the end of harvetsing. The highest effective methods to control of root-knot nematodes was in Vineem after 90 days applying, achieved 66.36%, next Biosun one after 60 days treatment (56.71%), then bio-fumigation (43 , 69%) after 30 days processing. Table 3.33 Effect of biological methods on density of the second-stage juveniles in roots, ratio of root infestation, root gall index and yield of eggplant in Lam Dong (2014-2015) Treatments Density of J2 in roots Ratio of root gall Root gall Yield (individuals/5 g root) (%) index (tons/ha) Chitosan 1596b 60.0bc 5.33c 108.00a Neem 796e 68.37b 5.67bc 102.70b Saponin 1400c 69.13b 6.33b 92.000e d b bc Biosun one 974 66.18 5.67 99.00c Biofumigation 874e 53.61c 5.00c 95.67d Control 2004a 78.03a 7.67a 95.67d LSD0,05 95.5600 8.3240 0.7776 3.2970 Density of the second-stage juveniles (M. incognita) in eggplant roots was the highest in control (2004 individuals/5g roots), followed by Jianon chitosan super treatment (1596 individuals/5g roots), Abuna 15GR (1400 individuals/5g roots). The lowest density of secondstage juveniles was in Vineem (796 individuals/5g roots) and biofumigation (874 individuals/5g roots). Ratio of root infestationand and root galls index in biofumigation treatment were the lowest, 53.61% and 5.00, respectively. These figures of control were the highest up to 78.03% and 7.67, respectively. While yield of eggplant in Jianon chitosan reached the highest value (108.00 tons/ha), followed by Vineem (102.70 tons/ha), the lowest in Abuna 15 GR only 92.00 tons/ha.
3.3.4 Chemical methods to control root-knot nematodes detected on eggplant Table 3.34 Effect of chemical methods on density of root-knot nematodes detected on eggplant in Lam Dong (2014-2015) Treatments Density of J2 in soil (individuals Efficiency (%) /50cm3) TXL 30N 60N 90N C.V 30N 60N 90N C.V a a a Tervigo 1419 772 953 1559 2638 43.77 51.15 -3.18 -0.45a d b Map logic 1220 1061 2061 1407 2538 10.12 -22.86 -8.31b -12.39b Cazinon 1171 757 2230 1638 3300 33.19b -38.49c -31.37c -52.57c Vifu-super 990 830 1952 1588 3155 13.35c -43.39c -50.64d -72.19d Control 1112 1076 1529 1184 2058 LSD0,05 2.978 3.552 1.338 2.318 Experimental results show that density of M. incognita in eggplant field during 2014-2015 decreased grammatically after 30 days applying then increase dramatically after 60 days, 90 days planting and at the end of the harvesting. All experimental treatments were effective to control root-knot nematodes after 30 days treatment but efficiency was not extended from 60 days to the end of harvest. Efficiency of Tervigo 020SC was higher than that of Map Logic 90WP and Vifusuper 5 GR. Table 3.35 Effect of chemical methods on density of second-stage juveniles in roots, ratio of root infestation, root gall index and yield of eggplant in Lam Dong ( 2014-2015) Density of J2 in roots Ratio of root Root gall Yield Treatments (individuals/5 g root) infestation (%) index (tons/ha) Tervigo 541c 38.27b 3.67c 104.00a Map logic 681c 73.54a 6.33b 88.33c b a b Cazinon 867 68.32 6.33 99.33ab Vifu-super 1184a 83.03a 7.00ab 88.67c Control 1019b 87.69a 7.33a 95.67b LSD0,05 154.4 20.02 0.9967 5.395 Density of the second-stage juveniles of M. incognita was highest in Vifu-super (1184 individuals/5g roots), followed by control (1012 individuals/5g roots). That value was the lowest in Tervigo treatment (541 individual/5g roots). Ratio of root infestation the Tervigo was the lowest, only 38.27%, different from all of treatments. Root gall index was the highest in control (7.33). The lowest root gall index was in Tervigo, only 3.68. Eggplant yield was the highest in Tervigo (104.00 tons/ha) higher than that of control (95.67 tons/ha), Map logic (88.33 tons/ha) and Vifu-super (88.67 tons/ha). Generally, in all of methods to control root-knot nematodes (M. incognita), the most effective method was biological methods, followed by physical methods, chemical methods and the lowest effective was cultivation methods. Biological methods, physical methods and cultivation methods were effective to control root-knot nematodes without affecting environment and humans, safety and low-cost. Therefore, this study tended to use integrated nematode management by combination between cultivation, physical and biological methods to control root-knot nematodes in 2 models in Don Duong and Duc Trong in Lam Dong province.
3.4. Solutions to control root knot nematodes (Meloidogyne incognita) on detected eggplant towards integrated management in Lam Dong 3.4.1 Effect of total management solutions to control of root-knot nematodes on nematode population density M. incognita Table 3.36 Density of second-stage juveniles of M. incognita and efficiency in Lam Dong (2016-2017) Model Density of J2 in soil (individuals/50cm3) Efficiency (%) TXL 30N 60N 90N C.V 30N 60N 90N C.V Don Duong 1031 708 458 701 1103 36.68 66.63 30.01 24.50 Control 1017 1103 1295 1129 1441 Duc Trong 859 624 523 1010 1061 35.13 58.55 26.68 18.47 Control 868 972 1275 1392 1315 CV (%) 6.12 5.27 5.19 6.79 7.51 Density of second-stage juveniles in soil in two models decreased rapidly after 30 days and 60 days applying, lower than that in control. Efficiency in both models was high, reached 66.63% in Don Duong and 58.55% in Duc Trong after 60 days applying. Efficiency to control root knot nematode extended from the beginning to the end of crop. 3.4.2 Effects of integrated nematode management on density of root-knot nematodes in roots, ratio of root gall, root gall index and yield of eggplant Density of root-knot nematodes in roots, root infestation, root gall index in both models were lower than that in control. Density of the second juveniles of M. incognita in roots of both models in Don Duong and Duc Trong were lower than that in control, proved by 107 individuals/5g roots, 218 individuals/5g roots, respectively. Table 3.37 Effect of integrated nematode management on root damage and yield of eggplant in Lam Dong (2016-2017) Model Density of root-knot nematodes Ratio of root Root gall Yield (individuals/5 g root) infestation (%) index (tons/ha) Don Duong 107d 12.53d 1.93b 116.33a Control 727a 37.32b 4.29a 91.67b c c b Duc Trong 218 22.68 2.28 112.33a Control 624b 41.51a 4.25a 95.67b LSD0,05 49.29 3.706 0.4801 4.587 Ratio of root infestation in the model of Don Duong (12.53%) and Duc Trong (22.68%) were low compared to the control of Don Duong (37.32%) and Duc Trong (41.51%). Root gall index of eggplant roots in Don Duong model (1.93) and Duc Trong model (2.28) were lower than that in both controls. Eggplant yield in both models was also higher than that in the control, reached the highest with 116.33 tons/ha in Don Duong model, higher than 24.66 tons/ha compared to the control model.
3.4.3 Economic efficiency of the model Table 3.38 Economic efficiency of integrated nematode management to control root-knot nematodes on eggplant in Lam Dong (2016-2017) Model Don Duong Control Duc Trong Control
Total cost (1,000 VND)) 125,820 125,500 123,520 123,800
Yield (kg/ha) 116,330 91,670 112,330 95,670
Net income (1,000 VND) 107,240 57,640 99,240 65,640
Efficiency (1,000 VND) 49,600 33,600 -
It is estimated that yield of the model increased, net income of models was higher than traditional method application to control root-knot nematodes. Model applied integrated nematode management in Don Duong, the net income was achieved 107,240 thousand VND (107,240,000 VND/ha) higher 49,600,000 VND/ha compared to the control. The model in Duc Trong was higher 33,600 thousand VND (33,600,000 VND/ha) compared to the control. The results show that economic efficiency of application of integrated nematode management was higher than that of farmers’application. Generally, application of integrated nematode management on eggplant to control root-knot nematodes was reduced root gall index, increased yield compared to farmers control methods. CONCLUSIONS AND RECOMMENDATIONS Conclusions 1. In this study two species of root-knot nematode parasitised on eggplant including M. incognita and M. javanica were identified. M. incognita was more common species, accounting for 67.61% while M. javanica accounted for 23.94% and mixture of two species made of 8.45%. 2. Life cycle of M. incognita culturing on eggplant root in sandy (70% sand, 20% limon, 10% clay) at temperature 24 ± 1oC was 27.16 days. At the humidity threshold of 46.77%, the inoculation level of second-stage juveniles into eggplant roots reached 95.46%, higher than that at humidity threshold of 57.11% (93.31%) and 36.13% (85.68%). Ratio of second-stage juveniles of M. incognita hatched from egg was highest in the extract of one-month eggplant roots (90.72%), higher than in the extract of six-months eggplants roots while the lowest ratio was in distilled water (69.30%). . 3. Sand content, soil moisture and rainfall had strong correlation with the density of second-stage juveniles in soil. The density of second-stage juveniles increased proportionally with the sand content. Density of M. incognita increased in rainfall from 0 to 160mm/month and soil moisture ranging from 30 to 55%, otherwise, density of M. incognita decreased proportionally with rainfall over than 160mm/month and humidity over 55%. Chicken manure, organic fertilizer declined density of M. incognita detected on eggplant. Eggplant TN252 Green King was the most susceptible by M. incognita. 4. Integrated nematodes management to control root-knot nematodes: Rotation between eggplant and Tung Ho was the most effective cultivation methods with efficiency of M. incognita reached 34.03%, ratio of root infestation (37.13%), root gall index (2.67). While burning was the most effective in physical methods to control root-knot nematodes with efficiency achieved 23