Doctor of Philosophy in BOTANY

STUDIES ON HISTOPATHOLOGICAL TRANSFORMATIONS INDUCED BY MELOIDOGYNE INCOGNITA IN BITTERGOURD, MOMORDICA CHARANTIA ABSTRACT THESIS SUBMITTED FOR THE DEGREE OF Doctor of Philosophy IN BOTANY BY MOHD. YAQUB BHAT T'S^Sl DEPARTMENT OF BOTANY ALIGARH MUSLIM UNIVERSITY ALIGARH (INDIA) 1999 I Acc. N- )*' '"'hat Univc^ ABSTRACT Momordica charantia, an important vegetable, belonging to the family Cucurbitaceae, is grown during summer throughout India. Often, the roots of plants are attacked by the root -knot nematode, Meloidogyne incognita that causes stunting, wilting, and yellowing of above ground parts and galling on the roots. Deformations in the galled roots affect absorption of water and minerals by the root and transport of these substances towards the shoot. The plant, at the initial stages of infection, shows wilting symptoms, the plant growth retards, the leaves become chlorotic, and finally the plant dies. The ultimate impact of the disease is the yield loss that may reach upto fifty percent. The experiments were carried out in various stages. In one phase the infected plants were compared with uninfected control plants. In the other phase disease controlling measures were applied and their effects on plants and nematode were studied. The overall growth of infected plants when treated with Paecilomyces lilacinus increased. The plant growth increased to a greater extent when the fungus was applied into the soil one week before nematode inoculation, when compared with nematode inoculated and untreated plants. In Tj plants, the lengths and weights of roots and shoots increased to such an extent that the plant growth became almost equal to { 2 } that of control plants. The fungus was found spreading in the soil, around the root surface, and in the inner tissues of the root. The fungus after penetrating into the root tissues grew inter-and intracellularly. It preferred the lumen of vessel elements, during intracellular development. The hyphae were observed in the giant cells and also in the body of the mature female. It damaged and destroyed eggs and egg masses and ultimaterly checked secondary infection. A decreasing trend in plant growth parameters was observed as the time of application of the fungus increased. In comparison to nematode inoculated and untreated plants, the treated plants exhibited an increase in number of branches, number of flowers, and leaf area. In the vascular tissue, less disruption was observed in the treated plants. In the second experiment, aldicarb a non-fumigant systemic nematicide was used to study its effect on the development of the nematode and on the plant growth. The nematicide was applied simultaneously at the time of inoculation and at an interval of one week to four weeks after nematode inoculation. The Tj plants in which nematicide was applied simultaneously alongwith nematode inoculation exhibited an enormous increase in plant growth as compared to nematode inoculated_and untreated olants. In T 3 plants the lengths and weights of roots and shoots, number of branches, leaf area, number of flowers increased. In these plants size of the gall, number of the galls, number of mature females, number of egg masses { 3 } were lower. When the application time increased from one week to four weeks, the growth of the plant decreased, number of galls, size of galls, number of mature females and number of egg masses increased. The anatomical studies of galled roots revealed most disruption in T^ plants and least disruption in T^ plants. The galls of T3 plants contatined less amount of abnormal xylem and abnormal pl^^m as compared to other treatments. The third experiment was aimed at knowing the effect of different inoculum levels of the nematode on the growth of the plant, on the formation of the galls, on the development of the nematodes, and on the formation of abnormal tissues inside the galls. An initial population density of Meloidogyne incognita affects different host plants differently. Low population level either may not affect a plant, or it may be beneficial or harmful. Momordica charantia responded differently to different population densities. At the lowest inoculum level there was not any remarkable decline in growth of plant as compared to control. At 50 Jj level, a slight but non-significant increase was observed. At higher initial inoculum levels the growth decreased significantly. Reduction in lengths and weights of both roots and shoots was maximum at 5,000 J„ the highest inoculum level. The galls were scanty and very small at lowest (Pi=05 Jj) initial inoculum level. The gall number and the gall size increased from lower to higher inoculum levels wi^h the maximum at highest inoculum level. { 4 } The number of mature females recovered from plants at Pi = 05 J^ was low that increased to maximum at Pi = 5,000 J^. However, their size decreased as the inoculum level increased. At lowest inoculum level one nematode was enough to cause the formation of giant cell complex. While at higher inoculum levels more nematodes were found causing multiple giant cell complexes. The average size of giant cell was larger at lower Pi and smaller at higher inoculum level. The giant cell cytoplasm was more dense at lower inoculum level than at higher inoculum level. Abnormalities in the orientation and structure of xylem and phloem were few at lower initial inoculum level and more at higher inoculum levels. At lowest Pi the number of giant cells around the head of the mature female was 6 to 8. The gaint cells were larger in size enclosed dense cytoplasm as compared to those found in the plant at high inoculum level. The amount of abnormal xylem and phloem was more at higher inoculum level than at lower inoculum level. Out of four primary inoculum levels (O5J2, SOJj, SOOJj and 5,000J2) considered, the galling was scanty and also the size of the gall was very small at the lowest inoculum level. The number of mature females recorded was small, at lowest inoculum level, however, their size was large as compared to the other inoculum levels. At this level the number of egg masses obtained was large. By the increase in primary inoculum level, the plant growth gradually reduced, number and size of the gall increased, number of { 5 } mature females per gram root increased but the size of the mature female decreased. The number of egg masses per plant also increased. Greatest reduction in length and weight of plant was observed at the highest initial inoculum level. The number and the size of the gall, the number of mature females per gram root and the number of egg masses per plant were maximum at the highest inoculum level. The size of the mature female decreased at higher inoculum levels. In another experiment different varieties of bittergourd available in the market were examined to find out any Meloidogyne incognita resistant variety. This experiment was conducted because previously tested resistant variety is not available. The six varieties viz Faizabadi, Jhalarwali, PDM, Jaunpuri, Baramasi and Aligarh local obtained from different seed sources were evaluated for this purpose. Out of the six different varieties of Momordica charantia selected, Aligarh local and Baramasi exhibited highest reduction in length and weight at various initial inoculum levels, as compared to control. The number and size of the galls and the number of the mature females were more in these two varieties. The varietv Faizabadi exhibited least effect of Meloidogyne incognita on plant growth. The gall number was lowest in this variety. In Jhalarwali variety the reduction in plant growth was more than Faizabadi but lower than other four varieties. It produced more galls than Faizabadi. The plant growth of nematode infected PDM and Jaunpuri plants was lower than Faizabadi and Jhalarwali but higher ( 6 } than Baramasi and Aligarh Local. In the variety Faizabadi no galling was observed and the galls if produced were small. In these galls, the giant cells were small, and contained little cytoplasm. The nematode either died or did not reach to mature stage. The galled regions were not distorted. Abnormal xylem and phloem was scarce. In other varieties number and size of galls, size of giant cells increased, being maximum in Aligarh local followed by Baramasi, Jaunpuri, PDM and Jhalarwali. In the last experiment Meloidogyne incognita infected roots were examined from the day one to the 30th day, after inoculation. At regular intervals of time anatomical studies were carried out to investigate sequential changes in the formation of giant cells, in the development of the nematode, and in the formation of hypertrophic and hyperplastic tissue and abnormal vascular elements. The characteristic feature in Meloidogyne induced galls is the formation of discrete, abnormally large giant cells. In growing roots, the juveniles of M. incognita induced giant cell in provascular elements, especially from the cells which develop into primary phloem. Soon after penetration, the juveniles entered the procambium zone and incited hypertrophy and hyperplasia not only in sieve tube transforming cells but also in the nearby cells. A large number of cells around the nematode head enlarged and many of them divided leading to the formation of the gall. The affected cells comprised of the cells of { 7 } cortex, endodermis, pericycle, conjuctive tissue, xylem, phloem and pith parenchyma. The juveniles of Meloidogyne incognita penetrated at or behind the root tips of Momordica charantia. They migrated intercellularly, in the inner tissue, by separating the cell walls. The giant cells were induced in the region of undifferentiated phloem within 48h of inoculation. Maximum number of nuclei and highly dense cytoplasm was noticed after 12 days of inoculation. Decrease in number of nuclei and increase in vacuolation was found after 18 days of inoculation. Smaller giant cells became empty and changed into vessel like elements by the deposition of lignified secondary wall material.

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