The host preference of a Ganoderma lucidum strain for three tree species of Fabaceae family; Cassia nodosa , Cassia fistula and Delonix regia

: Root and butt rot of Cassia nodosa, Cassia fistula and Delonix regia is caused by Ganoderma lucidum . Ganoderma root and butt rot is a lethal disease on C. nodosa, C. fistula, and D. regia trees


INTRODUCTION
The genus Ganoderma includes several wood decaying fungi on living trees as well as dead trunks and stumps, and has been recorded mostly in tropical and temperate countries. Generally, Ganoderma spp. cause extensive heart rots of standing trees by growing in the central, non-living woody tissues. Several studies have been carried out on Ganoderma diseases focusing on economical damage, severity of the disease and host range in many regions such as America, Asia, the Middle East and Europe. Ganoderma lucidum has been reported as the causal organism of the heart rot disease of several tree species; Quercus spp. 1 , Cocos nucifera 2 , Camellia sinensis 3 , Prunus persica 4 , Vitis vinifera 5 . Moreover, Ganoderma diseases have been recorded on 144 hosts in India, and of those G. lucidum has been listed on monitored regularly. Detailed studies were carried out in order to identify the causal agent of the disease using morphological, anatomical and cultural characteristics. Macroscopic and Microscopic studies: Morphological features of basidiocarps, such as shape, size, thickness and margin of pileus were examined. Furthermore, the colour and the texture of the pileus and stipe were also recorded.
Cultural characteristics: Both the pathogen and infested wood chips were cultured on potato dextrose agar medium. Pieces (4x4x2 mm) of pileus and wood chips (4x4x2 mm) were placed on plates on four opposite sides of the plate at right angles, 1cm away from the edge of the plate. Four plates were maintained for each isolate and incubated at 30±1 0 C. Colony characteristics, hyphal features and chlamydospores were examined under the microscope.

Impact of the disease on hosts
Out of 45 live tree species studied, C. nodosa, C. fistula and D. regia were infected by G. lucidum and basidiocarps were observed on the base of the trees. Two single annual badsidiocarps were observed at the base of the live tree trunk, 5-15 cm above ground level as the first sign of the disease on C. fistula. C. nodosa and D. regia, 3-5 single fruit bodies were observed on the live trunk, 5-20 cm above ground level. 5-8 fruit bodies on exposed roots within 1 m radius of the base of trees were also observed. The initial symptom, wilting of the leaves appeared after a few months of the formation of basidiocarps followed by yellowing of the leaves, defoliation, and finally dieback was observed in the crown. The time taken to develop symptoms and death varied with species and the age of the hosts (Table 1). Foliar symptoms appeared on C. fistula five months after observation of basidiocarps and died one month later. Symptoms appeared on C. nodosa and D. regia 10-22 months after exhibiting the first sign of the disease and died 1-2 months later. However, relatively old trees died 8-24 months after the appearance of the first sign of the disease.
Excavated roots of the diseased trees showed rootto-root contact with neighbouring trees. The wood of the butt region of dead trees appeared off-white and had lost its hardness compared to sound wood.

Disease centres
Centre 1 was located on the main access road to the university as a row of trees along the road sides. The row comprises D. regia and Tabebuia rosea with 5m intervals between trees. In addition Jacaranda mimosifolia, Roystonea regia and C. nodosa were also located in the close vicinity. Four D. regia trees ages between 11 to 22 years died due to root and butt rot disease within a 14-year period in a sequential pattern beginning at one end.
Centre 2 was a row of C. nodosa and Mangifera indica trees (at 4 m intervals) along the multi-storey building located on grassland and associated with scattered Bambusa vulgaris, M. indica and Mesua ferrea trees within 3-8m distances. Three C. nodosa trees (ages between 14 to 18.5 years) died along the row over the study period due to the same disease. M. indica trees within the row were not affected by the disease.
Centre 3 was a grass covered courtyard located by the side of a building with scattered C. nodosa, C. fistula, B. vulgaris, Polyalthia longifolia and Leucaena leucocephala trees spaced around 3-5 m distances between the trees. Two trees, C. nodosa and C. fistula aged 15 and 16.5 years respectively died of root and butt rot disease. The distance between these two trees was 7m. Another C. fistula tree located on the other side (7 m away) of the road was not affected. Many other tree species L. leucocephala B. vulgaris and P. longifolia were observed in close vicinity of this centre which were not diseased.
Centre 4 was a row of C. nodosa and Pinus caribea trees along a minor road on the campus. The interval between two trees was about 4 m. Only two Cassia nodosa trees 4 m apart died from this disease in the row. In addition there were Bauhinia variegata, P. longifolia and Plumeria rubra located in the close vicinity, but these tree species were not affected.

Basidiocarps
Few small white button shaped structures appeared singly on the infected trunks, 5-15 cm above the ground level and on exposed roots. Eventually, they elongated into 5-6 cm long, cylindrical to flat, erect immature basidiocarps. It comprised a white top and a reddishbrown glazy stipe. Thereafter, slow growth rate of the basidiocarp was observed and a horizontal hoof-shaped pileus was formed with a stipe attached to the wood. At maturity the pileus was also tightly attached to the wood. The upper surface of the young basidiocarp was glazed yellowish brown with a white margin while the lower surface was white with a glazed reddish brown stipe.
The mature basidiocarp was hoof-shaped, hard, shiny and mahogany-red on top with concentric grooves, and cream to white under surface with a reddish-brown boarder along the pileus margin. The length and width of the mature basidiocarp were 8.0 cm and 9.0 cm respectively. The cross section of the basidiocarp is more or less wedge-shaped while the thickness at the proximal end was 2 cm which thinned out towards the marginal end (0.5 cm). The colour of old basidiocarps changed into dark maroon to black on top and the texture of the pileus changed to be spongy. The underside appeared as ash-brown with a 1.2 cm broad, shiny reddish-brown boarder along the pileus margin.
Hoof-shaped, laterally stipitate basidiocarps were formed singly at the base of the trunk directly attached to the wood. Basidiocarps were observed on exposed lateral roots within 1 m radius of the base of D. regia and C. nodosa whereas on C. fistula, basidiocarps were formed only on the butt region up to 15 cm height of the trunk. The bark where the basidiocarps formed was slightly depressed and cracked.

Cultural and microscopical characteristics
The colony colour, growth characteristics of isolates from pileus and wood chips were more or less the same in this study ( Table 2).
The width of dark brown skeletal hyphae of the context was 4.47-7.45µm. Round to elliptical-shaped pores were observed on the underside of the pileus, with pore length of 158-168 µm and width of 141-152 µm. 4-5 pores/mm were observed on the pore layer. Basidiospores were golden brown, ovate, thickwalled and bitunicate. The length and width of spores ranged from 8.94 to 10.43 µm and 5.99 to7.45µm respectively.

DISCUSSION
Macroscopic, microscopic and cultural characteristics of the Ganoderma isolates were exactly identical with the characteristic features of Ganoderma lucidum complex, which has been described previously by several authors 8,9 . Morphological features of basidiospores of the isolated pathogen were similar to that of G. lucidum strains reported by several authors. Further, the size of basidiospores measured in the current study is comparable with sizes of previous records of G. lucidum; 8.5-13x5.5-8.5 µm 10 , 7-13x5-9 µm 11 ,9-11x6-7 µm 12 7-13x 6-8 µm 13 and 9.9-11.9x6.2-7.7 µm 14 . The basidispores of Ganoderma spp. within the family Ganodermataceae are considered as characteristic taxonomic feature 15 . Hence, the isolated causal organism of root and butt rot of C. nodosa, C. fistula and D. regia was identified as a Ganoderma lucidum strain. Several distinctive strains have been identified in Ganoderma lucidum complex from different parts of the world.
Infected trees survive for varying periods of time, depending on the species and the age of the tree. C. fistula was the most susceptible tree to G. lucidum strain, dying within a relatively short period (6 months) of formation of fruit bodies (signs) while the other two species, C. nodosa and D. regia showed mild susceptibility, and survived for 1-2 year periods after exhibiting the first sign. Generally, younger trees of these species are more susceptible to the disease than older trees.
In the current study, the particular strain of G. lucidum was identified as the causal agent of only three host species, which include two previously recorded species by Sankaran et al. 6 . It is evident that the isolated G. lucidum strain shows host preference, and the decay causing pathogen attacks only C. nodosa, C. fistula and D. regia trees.