Evaluation of Ten Cacao Clones in A Root-Knot Nematode Meloidogyne Incognita Infested Field

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1 Scientia Agriculturae E-ISSN: X / P-ISSN: DOI: /PSCP.SA Sci. Agri. 11 (0), 2015: PSCI Publications Evaluation of Ten Cacao Clones in A Root-Knot Nematode Meloidogyne Incognita Infested Field Okeniyi M.O 1, Adedeji A.R 1, Orisajo S.B 1, Agbebaku E.A 2 1. Crop Protection Division, Cocoa Research Institute of Nigeria, P. M. B. 5244, Ibadan 2. Farming System and Extension Division, Cocoa Research Institute of Nigeria, P. M. B. 5244, Ibadan Corresponding author michael_okeniyi@yahoo.co.uk Paper Information A B S T R A C T This experiment was carried out on the field in a Randomized Complete Received: 8 April, 2015 Block Design (40 X 10m plot) to evaluate cacao clones for resistant in nematode infested field. The twelve cocoa clones evaluated are MXC67, Accepted: 11 June, 2015 T86/2, PA150, LCTEEN, T12/11, T53/5, T101/15, T65/7, ICS1 and AMAZ Based on gall index, nematode reproduction factor and Published: 20 August, 2015 growth parameters, MXC67, T86/2, PA150, T101/15 and T53/5 were susceptible to the nematode. Two other clones, T65/7 and ICS1 were Citation tolerant. A high degree of resistance was exhibited by LCTEEN, T12/11 and AMAZ Compared with F 3 Amazon and Amelonado varieties, Okeniyi MO, Adedeji AR, Orisajo SB, Agbebaku EA Evaluation of Ten Cacao Clones in A Root-Knot Nematode Meloidogyne Incognita Infested Field. Scientia the two most famous cocoa varieties in Nigeria, four clones (LCTEEN, T65/7, ICSI and AMAZ 15-15) were superior to F 3 Amazon and Amelonado. This study showed that some of the clones that when planted Agriculturae, 11 (2), Retrieved from in nematode infested soil will drastically reduce seedling failure (DOI: experienced by farmers. This tolerant and resistant clones identified should /PSCP.SA ) be included in breeding programme for resistance PSCI Publisher All rights reserved. Key words: Resistance, Tolerance, Susceptible, Clones and Meloidogyne incognita MXC67, T86/2, PA150, LCTEEN, T12/11, T53/5, T101/15, T65/7, ICS1 and AMAZ Introduction The cacao plant belongs to the genus Theobroma in the family Malvaceae. Over 20 species of Theobroma are recognized. All cocoa cultivated for the international market belong to the single species Theobroma cacao (L). There are three large and dinstinct groups within the species T. cacao. These are the Criollo, the Trintario and the Forastero Amazonia (Opeke, 2005). Currently, Nigeria is the fourth largest cocoa producing country in the world, coming after Cote d Ivoire, Ghana, and Indonesia and accounting for 6% of world total output (Omololu, 2014). Cocoa occupies a total of over 403,000 hectares of farmland in seventeen states of the country although popular claim is fourteen states (RMRDC Survey report 2004). Meloidogyne spp are the most important nematodes of cacao due to their pathogenicity and wide distribution in cocoa producing regions (Campos & Villain, 2005). It is a common pest of cacao in West Africa (Whitehead, 1969; Asare- Nyako and Owusu, 1979; Fademi et al., 2006; Okeniyi et al., 2009). Symptoms of M. incognita damage on cacao seedlings are dieback, stunting, wilting, chlorosis and reduction in size of the leaves, galling of the root or complete death of the seedlings (Afolami and Caveness, 1983; Orisajo and Fademi, 2005; Orisajo et al., 2007). Although control strategies have been based on the use of chemical nematicides (Afolami, 1993), the need for alternatives to nematicides has stimulated research focusing on sustainable tactics for management of plant parasitic nematodes (McSorley and Poranzinska, 2001). The objective of this research was to evaluate clones of cacao for their reaction to Meloidogyne incognita on the field. Materials and Methods Field experiment was carried out at the research farm of Cocoa Research Institute of Nigeria (CRIN) at Ibadan, Nigeria (Latitude 7.26, Longitude 3.54 and 122m above sea level). The annual rainfall ranges between mm, distributed over 5-7 months from April to October. The average daily temperature range is C.

2 Field Experiment A 40m x 10m plot was used in a Randomized Complete Block Design (RCBD) to test the susceptibility/resistance of the clones (MXC67, T86/2, PA150, LCTEEN, T12/11, T53/5, T101/15, T65/7, ICS1 and AMAZ 15-15) in nematode infested soil. Pre-planting survey was carried out to determine the population of plant parasitic nematodes in the plot using Coyne et al., 2007 tray modification of Baermann technique. Six months old seedlings of all the clones raised in the nursery were transplanted into the nematode infested field. After transplanting, regular visual observations were made for disease symptoms expression. The growth parameters such as plant height, stem girth, and numbers of leaves were recorded. To assess infection, destructive samplings of some seedlings was done. The roots were carefully reed of soil, washed under a gentle stream of tap water, mopped and galls counted using a hand lens at 3-5 X magnification. Root galling was assessed using the 0-5 gall index (Sasser et al., 1984). Nematode eggs were collected from each root system using sodium hypochlorite method (NaOCl) of Hussey and Barker (1973) and counted. Aliquots of 100-cm3 soil samples from each pot were assayed for juveniles of M. incognita using the modified Baermann technique (Coyne et al., 2007). Analysis of variance (ANOVA) and correlation analysis were carried out on data collected and the means were compared using the New Duncan Multiple Range Test (DMRT). Resistance rating was carried out using the quantitative scheme for assigning crop varieties into resistance categories based on crop yield, reproduction factor R, and gall index (GI) (Afolami et al., 2004) Results Effects of M. incognita on the growth and development of cocoa clones Plant growth as expressed by height, number of leaves and stem girth was significantly (P= 0.05) suppressed by M. incognita population in the field. The nematode stimulated a reaction that led to a better growth as expressed by significant height in PA150 this was followed by T53/5. When compared with F 3 Amazon and Amelonado which are being planted by farmers, all the clones performed better with the exception of AMAZ which recorded the least height (Figure 1). Figure 2 shows the effect of M. incognita on mean number of leaf. MXC 67 had the highest number of leaves; this was closely followed by T86/2. All the clones screened compare favourably with the two varieties that served as the control (F 3 Amazon and Amelonado). In terms of stem girth, all the clones compared favourably with F 3 Amazon and Amelonado (Figure 3). However, for the fresh root weight T53/5 recorded the highest weight with AMAZ having the least root weight (Figure 4). For fresh leaf weight, there was no significant difference in leaf weight of MXC67, PA150, T12/11 and T53/5 (Figure 5). T53/5, T101/15, PA150and MXC67 showed a significantly higher weights than the other clones even above F 3 Amazon and Amelonado. Root investigation of PA150revealed a good root development with fine root hair. AMAZ also showed comparatively healthy shoot growth. PA150 expressed good growth with healthy shoot development, having a significantly increased stem girth and total dry matter. Root investigation of PA150 revealed a healthy root system with fine root hairs. AMAZ showed an incipient growth advantage in terms of leaf area, stem girth and good root development which later resulted in leaf drop that gave them unthrifty appearance. The inoculated plant showed drastic effect of root-knot nematode infection at a latter stage of growth by expressing a significant reduction in leaf number, stem girth and total dry matter with a reproduction factor of 1.0 and gall index of 2.0 (Table 2). Amelonado and F 3 Amazon manifested chlorosis six weeks after transplanting. They both manifested stunted growth, and later investigation revealed poor root development with galls in these plants. Nematode infection led to a reaction that stimulated a rapid growth in T86/2 but with reduced leaf number, stem girth and poor root development. Eight weeks after transplanting, vein clearing and narrowing of leaves were observed at an early stage of growth in T53/5. The vein clearing later disappeared and root investigation revealed poor root development with galls. ICS1 also exhibited a better growth in terms of the growth indices in the presence of M. incognita infection as expressed by significantly increased height; leaf number and stem girth. T65/7 perform favorably well in terms of the growth parameters vis-a-vis plant height,leaf number, stem girth and root development. T12/11 had growth advantage in terms of leaf number, height, stem girth and total dry matter, root investigation revealed good root development with fine root hair when compared to the nematode-free plant. Discussions The growth response of T. cacao clones in nematode infested field is presented in Figure 1to Figure 6. Of the ten clones evaluated, LCTEEN and T12/11 gave the best vegetative plant characters such as plant height, number of leaf, stem girth, fresh root weight and fresh leaf weight. LCTEEN and ICS1 gave the highest plant height, which was closely followed 91

3 by MXC67 and T53/5. The least plant height was recorded in Amelonado (Table 1). In terms of number of leaf AMAZ followed by LCTEEN gave significantly better vegetative yield performance than others. In all the clones, evaluated T12/11 had a significantly high stem girth followed by LCTEEN, T53/5 and ICS1, while AMAZ and Amelonado yielded the least stem girth. The fresh shoot weight was higher in T12/11 and followed by ICS1 while the lowest shoot yield was recorded in Amelonado (Fig.2). LCTEEN and T101/15 had the highest fresh root weight; this was followed by T12/11. F3 Amazon and Amelonado yielded the least root weight (Table 2). T12/11 yielded the highest total dry matter followed by T101/15. F3 Amazon, Amelonado and T86/2 yielded the least total dry matter. Table 4 gives the response and rating of ten clones of cocoa using the modified version Afolami et al., (2000) quantitative scheme for assigning crop varieties into resistance categories, LCTEEN, T12/11 and AMAZ were poor host of M. incognita and they exhibit resistant characteristics to the nematode, which was unable to reproduce on these clones. These clones showed good root development in spite of M. incognita infection. The galls were fine and root hairs not destroyed. MXC67, T86/2, PA150, T101/15 and T53/5 were obviously very susceptible to the nematode and were rated as susceptible to M. incognita because the nematode successfully established itself on these clones. T65/7 and ICS1 were rated as tolerant since the nematode was also able to establish itself successfully on the clones. The results of this experiment having employed a combination of root-gall index, nematode reproduction factor and other growth indices showed that of the ten clones tested, five were rated as being susceptible, three were resistant and two being tolerant. This result further corroborated the initial screen-house evaluation by Okeniyi et al., 2009 Of all the clones that were evaluated showed individual variations in growth response when inoculated with M. Incognita eggs, which according to Okeniyi et al., 2009, may in-part be a function of the pest pressures which resulted in an indirect relationship between the various growth indices and the nematodes populations and partly due to genetic expressions of the innate plant characters. The similarities among the tested clones in thier respective host reaction for M. incognita reproduction according to (Nwanguma et al., 2005) reflected the lack of genetic variability among the clones. It thus implies that the uniformity in genetic trait could pose difficulty in obtaining resistant. The relatively low populations of the nematodes in clones AMAZ 15-15, T12/11 and LCTEEN indicated some degree of resistance, which probably explains the no damage of the root system and thus insignificant impact on the growth characters (Okeniyi et al., 2009). AMAZ was superior in total dry matter to all the other clones tested. The superiority of AMAZ 15-15, T12/11 and LCTEEN (resistant clones); T65/7 and ICS1 (tolerant clones) in terms of plant height, leaf number and total dry matter according to Okeniyi et al., 2009, could thus be attributed to the low level of root nematode infection which resulted in enhanced root development and ability to tap nutrients from the soil maximally. It is also a known fact that the ability to withstand nematode attack good plant vigour and high yield are all indices for high level of plant tolerance (Nwanguma et al.,2005). NEMATODE GENERA Table 1. Plant-parasitic nematode extracted from experimental plot at CRIN, Ibadan FREQUENCY OF % FREQUENCY NEMATODE % NEMATODE OCCURRENCE IN 250g RATING IN 250g SOIL* POPULATION IN 250g POPULATION IN 250g SOIL SOI; SOIL** Meloidogyne spp , Pratylenchus spp , Helicotylenchus spp , Paralongidorus spp , Eutylenchus spp , Scutellonema spp , Hemicyclophora spp , Xiphinema spp , Longidorus spp , Anguillulina spp , NOTE: * = n/nx100 (n=number of times individual nematode occurred and N=sample size (60) ** = n/tnx100 (n=individual nematode in all the sample and TN=Total population of all the nematode extracted in the sample. 92

4 Figure 1. Influence of M. incognita on mean plant height of cacao seedlings 24 weeks after transplanting. Figure 2. Influence of M. incognita on mean number of leaf of cacao seedlings 24 weeks after transplanting. Figure 3. Influence of M. incognita on mean girth of cacao seedlings 24 weeks after transplanting. 93

5 Figure 4. Influence of M. incognita on mean fresh root weight of cacao seedlings 24 weeks after transplanting. Figure 5. Influence of M. incognita on mean fresh leaf weight of cacao seedlings 24 weeks after transplanting. Figure 6. Influence of M. incognita on mean fresh shoot weight of cacao seedlings 24 weeks after transplanting. 94

6 Table 2. Resistance Rating of ten clones of cacao using the quantitative scheme for assigning crop varieties into resistance categories based on dry matter, reproduction factor and gall index (Afolami et al., 2004). Mean Gall Reproduction Resistance No of Index factor Category Clones Gall (GI) (R) MX Susceptible T8/ Susceptible PA Susceptible LCTEEN Resistant T12/ Resistant T101/ Susceptible T53/ Susceptible T65/ Tolerant ICS Tolerant AMAZ Susceptible F 3 AMAZON Susceptible AMELONADO Susceptible GI=Gall index where 1=1-2 galls, 2=3-10 galls, 3=11-30 galls, galls, 5=>100 galls (Taylor and Sasser, 1978) R=Nematode Reproduction factor-final number of juveniles and eggs Conclusion The result of this study clearly showed that in spite of the resistance of all these clones to Sahibergella singularis and black pod disease (Otuonye et al., 2007), only three of them are resistant and two of the clones are tolerant to M. Incognita infection. It is therefore evident that breeding for resistance can no longer neglect the effect of nematodes. References Afolami SO, Atungwu JJ, Odeyemi IS, Orisajo SB Going beyond gall index in studying and reporting resisitance to root-knot nematodes. Nigerian Journal of Plant Protection. 21 : Afolami SO, Caveness FE The Frequency of occurrence and geographical distribution of plant parasitic nematodes associated with Theobroma cacao in Nigeria. Turrialba 33(1) : Afolami SO, Ojo AA Screning of Theobroma cacao germplasm for resistance against a root-knot nematode, Meloidogyne incognita in Nigeria. In Proceeding of the 9 th International Cocoa Research Conference. Lome Togo, pp Afolami SO The Effect of Basamid Granular (Dazomet) on nematodes parasitic on cacao seedlings in the nursery. In: Proceedings, 11th International Cocoa Research Conference, Bahia, Brazil. pp Afolami SO Suggestions for the improvement of current methods of studying and reporting resistance to root-knot nematodes. International Journal of Nematology,10(1): Asare-Nyako A, Owusu GK Meloidogyne incognita infection of cocoa seedlings. In:Proceedings, 7th International Cocoa Research Conference, Douala, Cameroun Nov pp Campos VP, Villain L Nematode parasites of coffee and cocoa. In: Luc M, Sikora RA, Bridge J (Editors), Plant Parasitic Nematodes in Subtropical and Tropical Agriculture,2nd Edition, Wallingford, UK: CABI Publ. pp Coyne DL, Nicol JM, Claudius-Cole B Practical plant nematology: a field and laboratory guide. SP-IPM Secretariat, International Institute of Tropical Agriculture (IITA), Cotonou, Benin. 84pp. Fademi OA, Orisajo SB, Afolami SO Impact of plant parasitic nematodes on cocoa production (in Nigeria) and outlook for future containment of the problem. In Proceedings 15th International Cocoa Research Conference, San Jose, Costa Rica. FAOSTAT online database Ferris H, Matute MM Structural and functional succession in the nematode fauna of a soil food web. Applied Soil Ecology 23: Gullino ML, Camponogara A, Gasparrini G, Rizzo V, Clini C, Garibaldi A Replacing methyl bromide for soil disinfestations: The Italian experience and implications for other countries. Plant Disease 87: Hussey RS, Barker KR A Comparison of methods of collecting inocula for Meloidogyne spp. including a new technique. Plant Disease reporter 57: Luc M, Guiran G. de Les nematodes associes aux plantes de L Quest Africain. Liste preliminaire. Agron. Trop. Nogent, 15 (4) McSorley R, Poranzinska DL Elements of sustainable agriculture. Nematropica 31: 1-9.Mohanty S, Paikaray NK, Rajan AR, Availability and uptake of phosphorus from organic manures in groundnut (Arachis hypogea L.)-corn (Zea mays L.) sequence using radio tracer technique. Geoderma 133: Orisajo SB, Dongo LN Nematicidal potential of some indigenous plant extracts against rootknot nematode on cacao. African Scientist 6(4): Orisajo SB, Fademi OA Influence of neemfortified cocoa pod husks soil amendment on Meloidogyne incognita in cocoa. African Scientist 6(3): Orisajo SB, Okeniyi MO, Fademi OA, Dongo LN Nematicidal effects of water leaf extracts of Acalypha ciliata, Jatropha gossypifolia, Azadirachta indica and Allium ascalonicum on Meloidogyne incognita infection on cacao seedlings. Journal of Research in Bioscience 3(3): Otuonye AH, Adeoti AYA, Agbeniyi SO, Aikpokpodion PO, Enikuomehin O, Popoola TO Evaluation of the susceptibility of local cocoa germplasms to Phytophthora pod rot disease using the leaf discs technique. Journal of Food, Agriculture and Environment Vol. 5 (3&4): Sasser JC, Carter CC, Hartman KM Standardization of host suitability studies and reporting of resistance to root-knot nematode. USAID (United States Agency for International Development), Raleigh, North Carolina, U.S.A., 7pp. Taylor C, Taylor L Future trends in cocoa industry - a perspective. Coffee and Cocoa International 33(3):