31
4.1.1.2 Pathogenicity test
To ensure that endophytic fungi isolates were non pathogenic to the host plants, pathogenicity test was conducted to determine pathogenic and
non-pathogenic isolates. Pathogenicity test was done by growing tomato seeds on pure colonies of fungi, results indicated that tomato seeds did not germinate on
pathogenic colonies while the seeds germinated on the non pathogenic colonies. Seeds were also germinated on PDA media to act as control which indicated that
seeds germinated better on non-pathogenic isolates than in PDA media this is a clear indication that endophytic fungi promote growth in plants as indicated in
Figure 4 From the pathogenicity test, 12 potential isolates of endophytic fungi were obtained and identified macroscopically and microscopically Appendix 7.
Figure 4. Pathogenicity test based on germination of tomato seeds on pure colonies of endophytic fungi and PDA media as control
a Pathogenic colony b non-pathogenic colony and c Tomato seeds germinated on PDA media as control
4.1.2 Colonisation Test
Colonisation is considered an important prerequisite of an effective antagonistic agent, re-isolation was done from the roots to determine the ability of
the endophytic fungi to colonise tomato plant roots, on the other hand no endophytic fungi re-isolation resulted from the control treatments since the control
plants were not inoculated with endophytic fungi Figure 5. The results indicated higher frequency of reisolation of endophytic fungi in
following order; isolate XP 9, Trichoderma pseudokoningii, Trichoderma hamatum, Nigrospora
sp, Fusarium oxysporum and Fusarium chlamydosporum by 100 followed by Chrysosporium sp and sterile black 1 that had percentage
b c a
32 frequency of reisolation value of 86 and 71 respectively, While the percentage
frequency of reisolation was very low in Sterile black 2, Torula sp, Fusarium sp3 and Ulocladium sp with the value of 57, 57, 57 and 43 respectively
Table 2.
Figure 5. Colonisation test a Reisolation of endophytic fungi on PDA media from tomato plant roots of endophytic fungi treated plants
b Re-isolation on PDA media of control tomato plant roots not treated with endophytic fungi
4.1.3 Antagonistic Effect of Endophytic Fungi Against RKN In-planta
All the endophytic fungi isolates showed high significant effect in suppression of the number of root galls and egg mass formation. Root gall was
highly suppressed by Nigrospora sp, Trichoderma pseudokoningii, isolate XP 9 , Table 2. Colonisation ability of endophytic fungi
Treatment Number of root
samples used for reisolation
Number of root samples where colonies that
emerged after reisolation Frequency
of reisolation Control 7 0 0
Torula sp
7 4 57 Sterile black 2
7 4
57 Sterile black 1
7 5
71 F. chlamydosporum
7 7 100
T. hamatum 7 7
100 F. oxysporum
7 7 100
Chrysosporium sp
7 6 86 T. pseudokoningii
7 7 100
Nigrospora sp
7 7 100
Isolate XP 9 7
7 100
Ulocladium sp
7 3 43 Fusarium
sp 3 7 4 57
a b
33 Trichoderma hamatum
and positive control carbofuran by 99.90, 99.82, 99.79, 99.47, 100 respectively, egg mass formation was highly suppressed
in by Trichoderma pseudokoningii, Trichoderma hamatum, Nigrospora sp, isolate XP 9, and positive control carbofuran by100 Table 3.
Table 3. Effect of endophytic fungi on number of root galls and egg masses
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05,
according to Duncan’s Multiple Range Test
4.1.4 Effect of Endophytic Fungi on Plant Growth 4.1.4.1 Effect on height and stem diameter of RKN inoculated plants
The data indicated that plant heights varied depending on different endophytic fungi used, the most effective endophytic fungi in increasing plant
height were Nigrospora sp, Chrysosporium sp, Fusarium chlamydosporum and Trichoderma pseudokoningii
, the least significant species were Ulocladium sp, and Fusarium sp 3 Table 4. The data also shows that stem diameter varied
depending on he endophytic fungi used, most effective endophytic fungi in increasing the stem diameter were Trichoderma pseudokoningii, Fusarium
oxysporum, Nigrospora sp, Torula sp, isolate XP 9 and Trichoderma hamatum
while the least significant species were Ulocladium sp, and Fusarium sp3 Table 5.
Treatment Number of
root galls
Reduction Number of egg
masses Reduction
Control Nematodes 331.67 ± 68.99 a
95.42 ± 37.21a Ulocladium
sp 75.75 ± 12.54 b
77.16 26.00 ± 9.03 b
72.75 Fusarium
sp 3 62.08 ± 10.39 b
81.28 20.92 ± 8.39 bc
78.07 Torula
sp 37.83 ± 13.09 c
88.60 15.58 ± 5.90 c
83.67 Sterile black 2
35.42 ± 11.97 c 89.32
13.42 ± 5.89 c 85.93
Sterile black 1 21.33 ± 6.54 cd
93.56 14.50 ± 4.62 c
84.80 F.chlamydosporum
15.25 ± 3.79 de 95.40
4.42 ± 1.78 d 95.36
Fusarium oxysporum 8.83 ± 3.01 de
97.34 1.83 ± 1.85 d
98.08 Chrysosporium
sp 5.25 ± 2.83 de
98.41 0.25 ± 0.45 d
99.73 T. hamatum
1.75 ± 2.45 e 99.47
0.00 ± 0.00 d 100
Isolate XP 9 0.67 ± 1.23 e
99.79 0.00 ± 0.00 d
100 T.pseudokoningii
0.58 ± 0.99 e 99.82
0.00 ± 0.00 d 100
Nigrospora sp
0.33 ± 0.65 e 99.90
0.00 ± 0.00 d 100
Control Carbofuran 0.00 ± 0.00 e
100 0.00 ± 0.00 d
100
34 Table 4. Effect of endophytic fungi on plant height of RKN inoculated plant
Treatment Mean plant height cm
8 th weeks 9 th weeks
10 th weeks 11 th weeks
Nigrospora sp
63.08 ± 2.35 a 66.17 ± 2.29 bc
69.42 ± 2.43 bc 72.75 ± 2.05 bc Chrysosporium
sp 63.00 ± 3.02 a
72.08 ± 2.90 a 75.50 ± 2.91 a
79.33 ± 3.31 a Fusarium chlamydosporum
62.25 ± 4.52 a 68.75 ± 5.23 ab
73.00 ± 4.86 ab 76.25 ± 4.92 ab Trichoderma pseudokoningii
61.17 ± 4.35 ab 64.67 ± 4.64 bcd
67.00 ± 4.05 cd 69.67 ± 3.85 cd Torula
sp 60.83 ± 3.43 ab
66.00 ± 4.00 bc 68.67 ± 4.89 c
72.83 ± 5.08 bc Sterile black 2
60.58 ± 2.15 ab 64.17 ± 3.07 cd
67.25 ± 3.44 cd 70.67 ± 3.06 cd Trichoderma hamatum
60.50 ± 3.90 ab 62.92 ± 4.03 cd
66.00 ± 4.26 cd 69.17 ± 4.28 cd Sterile black 1
60.08 ± 2.78 ab 65.50 ± 3.26 bcd
67.83 ± 3.24 cd 70.67 ± 2.93 cd Isolate XP 9
58.17 ± 3.99 b 61.42 ± 4.96 d
64.25 ± 4.96 d 67.75 ± 4.83 d
Fusarium oxysporum 57.67 ± 2.27 b
65.67 ± 5.35 bcd 68.67 ± 5.14 c
71.83 ± 4.86 cd Fusarium
sp 3 42.33 ± 5.33 c
46.25 ± 6.22 e 47.08 ± 6.64 e
47.75 ± 6.93 e Ulocladium
sp 42.00 ± 7.25 c
48.83 ± 7.26 e 50.17 ± 7.17 e
50.67 ± 7.48 e Control Carbofuran
33.50 ± 7.71 d 37.92 ± 7.59 f
39.42 ± 7.08 f 39.83 ± 7.16 f
Control Nematodes only 25.50 ± 1.68 e
26.08 ± 1.73 g 27.08 ± 1.78 g
27.58 ± 1.78 g
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05, according to Duncan’s Multiple Range Test
34
35 Table 5. Effect of endophytic fungi on stem diameter of RKN inoculated plants
Treatments Mean stem diameter cm
8 th Week 9 th Week
10 th Week 11 th Week
Trichoderma pseudokoningii 0.90 ± 0.13 a
0.98 ± 0.09 a 0.98 ± 0.09 a
0.98 ± 0.089 a Torula
sp 0.87 ± 0.11 ab
0.91 ± 0.10 abc 0.94 ± 0.1 abc
0.95 ± 0.10 ab Fusarium oxysporum
0.85 ± 0.09 ab 0.94 ± 0.20 ab
0.95 ± 0.10 ab 0.95 ± 0.10 ab
Nigrospora sp
0.84 ± 0.12 ab 0.89 ± 0.12 bc
0.90 ± 0.12 abc 0.90 ± 0.11 abc
Sterile black 1 0.83 ± 0.08 abc
0.91 ± 0.11 abc 0.93 ± 0.10 abc
0.93 ± 0.10 abc Isolate XP 9
0.81 ± 0.12 abc 0.87 ± 0.11 bc
0.90 ± 0.12 abc 0.90 ± 0.12 abc
Trichoderma hamatum 0.82 ± 0.09 abc
0.93 ± 0.15 abc 0.93 ± 0.15 abc
0.93 ± 0.15 abc Chrysosporium
sp 0.82 ± 0.05 bc
0.91 ± 0.05 abc 0.95 ± 0.0 ab
0.96 ± 0.049 a Fusarium chlamydosporum
0.81 ± 0.06 bc 0.85 ± 0.07 c
0.87 ± 0.07 bc 0.87 ± 0.07 bc
Sterile black 2 0.76 ± 0.05 c
0.84 ± 0.12 c 0.85 ± 0.12 c
0.86 ± 0.12 c Control Carbofuran
0.60 ± 0.05 d 0.60 ± 0.05 d
0.60 ± 0.05 d 0.61 ± 0.06 d
Ulocladium sp
0.57 ± 0.05 de 0.57 ± 0.05 d
0.57 ± 0.05 d 0.57 ± 0.05 d
Fusarium sp 3
0.57 ± 0.05 de 0.57 ± 0.05 d
0.57 ± 0.05 d 0.57 ± 0.05 d
Control Nematodes only 0.51 ± 0.07 e
0.54 ± 0.07 d 0.54 ± 0.07 d
0.55 ± 0.06 d
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05, according to Duncan’s Multiple Range Test
35
36
4.1.4.2 Effect on plant fresh and dry weight of RKN inoculated plants
The data indicated that plant fresh weight varied depending on the different endophytic fungi treatments used with the most effective being Trichoderma
pseudokoningii, isolate XP 9 and Nigrospora sp leading to increased plant fresh
weights by 126.83 g, 124.50 g and 124.08 g respectively, while least mean plant fresh weights were observed in Ulocladium sp, and Fusarium sp3 that is 57.42 g and 56.17
g respectively. The data also indicated that plant dry weight varied depending on different species of endophytic fungi, with the most effective being Nigrospora sp,
isolate XP 9, Trichoderma pseudokoningii, Fusarium oxysporum, with dry weight of 42.75 g, 41.50 g, 41.33 g, 39.17 g respectively while the least significant endophytic
fungi treatments in increasing plant dry weight were Ulocladium sp, and Fusarium sp3 with mean dry weights of 18.67 g and 17.00 g respectively Table 6.
Table 6. Effect of endophytic fungi on plant fresh and dry weight Treatment Fresh
weight g Dry weight g
Trichoderma pseudokoningii 126.83 ± 16.74 a
41.33 ± 3.70 ab Isolate XP 9
124.50 ± 13.07 a 41.50 ± 1.93 ab
Nigrospora sp
124.08 ± 11.78 a 42.75 ± 1.96 a
Fusarium oxysporum 113.33 ± 13.44 b
39.17 ± 6.26 abc Sterile black 1
104.67 ± 9.72 bc 36.58 ± 5.68 c
Chrysosporium sp
101.17 ± 11.02 c 37.83 ± 4.15 bc
Torula sp
101.00 ± 7.41 c 36.08 ± 4.58 c
Fusarium chlamydosporum 99.08 ± 14.58 c
39.08 ± 3.63 abc Trichoderma hamatum
98.25 ± 16.22 c 37.92 ± 4.50 bc
Sterile black 2 84.25 ± 8.32 d
36.08 ± 4.54 c Ulocladium
sp 57.42 ± 10.66 e
18.67 ± 4.70 d Fusarium
sp 3 56.17 ± 9.75 e
17.00 ± 4.79 e Control Carbofuran
39.83 ± 7.16 f 20.83 ± 3.46 d
Control Nematodes only 29.33 ± 4.62 g
10.58 ± 2.43 f
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05,
according to Duncan’s Multiple Range Test
37
4.1.4.3 Effect on number of fruit and root length of RKN inoculated plant
The data on mean number of fruits indicated that the number of fruits were significantly increased by Nigrospora sp, isolate XP 9, Trichoderma pseudokoningii,
Chrysosporium sp, Fusarium chlamydosporum, and Trichoderma hamatum with
mean number of fruits of 13.08, 12.42, 12.33, 11.83, 11.33, and 11.17, respectively in that order, least mean number of fruits were observed in Ulocladium sp, and
Fusarium sp 3 that is 4.00 and 3.92 respectively Table 7.
The data also indicates that all isolates used were highly significant in increasing root length development in comparison to the control treatments, whereby
Nigrospora sp, isolate XP 9, Trichoderma pseudokoningii, Fusarium oxysporum, and
Fusarium chlamydosporum had the highest mean root length of 42.75 cm, 41.50 cm.
41.33 cm and 39.17 cm , respectively, least root length was observed in treatments with Ulocladium sp, and Fusarium sp3 that is 18.67 cm and 17.00 cm respectively
Table 7 and appendix 5. Table 7. Effect of endophytic fungi on number of fruits and root length
Treatments No. of fruits
Root length cm Nigrospora
sp 13.08 ± 1.24 a
42.75 ± 1.96 a Isolate XP 9
12.42 ± 2.43 ab 41.50 ± 1.94 ab
Trichoderma pseudokoningii 12.33 ± 1.67 ab
41.33 ± 3.70 ab Chrysosporium
sp 11.83 ± 1.90 abc
37.83 ± 4.15 bc Fusarium chlamydosporum
11.33 ± 2.87 abcd 39.08 ± 3.63 abc
Trichoderma hamatum 11.17 ± 2.52 abcd
37.92 ± 4.50 bc Torula
sp 10.42 ± 2.15 bcd
36.08 ± 4.58 c Sterile black 2
10.08 ± 2.58 cd 36.08 ± 4.54 c
Sterile black 1 10.00 ± 2.70 cd
36.58 ± 5.68 c Fusarium oxysporum
9.67 ± 2.81 d 39.17 ± 6.26 abc
Control Carbofuran 7.25 ± 2.74 e
20.83 ± 3.46 d Ulocladium
sp 4.00 ± 1.41 f
18.67 ± 4.70 de Fusarium
sp 3 3.92 ± 1.44 f
17.00 ± 4.79 e Control Nematodes only
1.17 ± 1.12 g 10.58 ± 2.43 f
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05,
according to Duncan’s Multiple Range Test
38
4.1.4.4 Effect on height and stem diameter of RKN free plants
The data indicates that all the tested endophytic fungi isolates generally showed significant increase in plant height every week during the four week period of
observation in comparison to the control treatments, the plant heights varied depending on the different species used, the most effective endophytic fungi were
Chrysosporium sp, Fusarium chlamydosporum, isolate XP 9, Nigrospora sp, Torula
sp, Trichoderma hamatum, Fusarium oxysporum and Sterile black 2 while the least significant species were Ulocladium sp, and Fusarium sp 3. Table 8 and appendix 3
On the same note all endophytic fungi isolates used showed significant effects in increasing stem diameter every week during the four week period of observation in
comparison to the control treatments, the stem diameter varied depending on the different species of endophytic fungi, the most effective endophytic fungi isolate in
increasing the plant diameter were Trichoderma pseudokoningii, Nigrospora sp, Torula
sp, Sterile black 1, isolate XP 9, Fusarium oxysporum and Trichoderma hamatum
while the least significant endophytic fungi in increasing stem diameter were Ulocladium sp, and Fusarium sp3 Table 9.
39
Table 8. Effect of endophytic fungi on plant height of RKN free tomato plant Treatment
Mean Plant height cm 8 th Week
9 th Week 10 th Week
11 th Week Chrysosporium
sp 65.08 ± 2.81 a
69.92 ± 2.78 a 73.58 ± 2.64 a
77.33 ± 2.27 a Fusarium chlamydosporum
63.75 ± 3.84 ab 65.00 ± 3.02 bc
69.33 ± 2.96 b 73.25 ± 2.49 b
Isolate XP 9 63.00 ± 3.30 ab
69.00 ± 3.50 a 72.17 ± 2.95 ab
75.17 ± 3.16 ab Nigrospora
sp 62.92 ± 4.12 ab
69.50 ± 2.91 a 73.33 ± 2.27 a
77.17 ± 2.55 a Torula
sp 62.58 ± 4.25 ab
64.33 ± 2.81 c 66.42 ± 2.84 c
69.92 ± 2.58 c Trichoderma hamatum
62.25 ± 3.08 ab 64.25 ± 1.91 c
65.92 ± 2.23 c 68.83 ± 2.32 c
Fusarium oxysporum 61.33 ± 2.06 ab
60.83 ± 2.29 d 62.67 ± 1.61 d
65.75 ± 1.66 d Sterile black 2
61.17 ± 3.22 ab 63.08 ± 2.11 cd
65.17 ± 1.95 cd 69.75 ± 1.66 c
Sterile black 1 60.67 ± 3.63 b
62.83 ± 1.95 cd 65.00 ± 1.41 cd
68.83 ± 1.90 c Trichoderma pseudokoningii
60.25 ± 3.57 b 67.58 ± 3.15 ab
71.00 ± 2.13 ab 74.00 ± 2.37 b
Fusarium sp 3
41.92 ± 6.82 c 43.25 ± 6.33 e
46.00 ± 6.27 e 49.08 ± 5.80 e
Ulocladium sp
38.83 ± 8.04 c 39.33 ± 7.92 f
40.25 ± 7.63 f 41.17 ± 7.61 f
Control Without Endophytic fungi 24.08 ± 3.26 d
25.42 ± 2.75 g 26.58 ± 2.43 g
28.33 ± 2.61 g
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05, according to Duncan’s Multiple Range Test
39
40
Table 9. Effect of endophytic fungi on stem diameter of RKN free tomato plant Treatment
Mean stem diameter cm 8 th week
9 th week 10 th week
11 th week Trichoderma pseudokoningii
0.90 ± 0.13 a 0.90 ± 0.131 a
0.93 ± 0.13 a 0.95 ± 0.13 a
Nigrospora sp 0.84 ± 0.11 ab
0.87 ± 0.07 ab 0.90 ± 0.06 ab
0.93 ± 0.07 ab Torula sp
0.83 ± 0.09 ab 0.85 ± 0.08 abc
0.87 ± 0.08 abc 0.89 ± 0.08 abc
Sterile black 1 0.83 ± 0.08 ab
0.83 ± 0.08 abcd 0.85 ± 0.07 bcd 0.88 ± 0.08 abc
Isolate XP 9 0.83 ± 0.12 ab
0.83 ± 0.12 abcd 0.85 ± 0.12 bcd 0.87 ± 0.13 bcd
Fusarium oxysporum 0.83 ± 0.10 ab
0.83 ± 0.09 abcd 0.84 ± 0.07 bcd 0.86 ± 0.08 bcde
Trichoderma hamatum 0.82 ± 0.09 ab
0.82 ± 0.09 bcd 0.84 ± 0.08 bcd 0.86 ± 0.08 bcde
Chrysosporium sp 0.79 ± 0.06 b
0.80 ± 0.06 bcd 0.82 ± 0.06 cd 0.83 ± 0.06 cde
Fusarium chlamydosporum 0.78 ± 0.07 b
0.78 ± 0.07 cd 0.78 ± 0.07 d
0.79 ± 0.06 e Sterile black 2
0.76 ± 0.05 b 0.76 ± 0.05 d
0.78 ± 0.05 d 0.80 ± 0.05 de
Fusarium sp 3
0.57 ± 0.05 c 0.57 ± 0.05 e
0.57 ± 0.04 e 0.58 ± 0.05 f
Ulocladium sp 0.56 ± 0.04 c
0.56 ± 0.04 e 0.57 ± 0.05 e
0.58 ± 0.04 f Control Without endophytic fungi
0.48 ± 0.09 d 0.48 ± 0.09 f
0.48 ± 0.08 f 0.48 ± 0.08 g
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05, according to Duncan’s Multiple Range Test
40
41
4.1.4.5 Effect on plant fresh and dry weight of RKN free plants
The data indicated significant increase in plant fresh weight by endophytic fungi treatments, most effective treatments were Nigrospora sp, isolate XP 9 and
Trichoderma pseudokoningii, with significant mean plant fresh weight of 134.33
g, 131.33 g and 130.00 g respectively in that order, least mean plant fresh weights were observed in treatments with Fusarium sp3 and Ulocladium sp, that is 70.08 g
and 69.17 g respectively Table 10. The data also indicated significant increase in plant dry weight, most
effective treatments were Nigrospora sp, isolate XP 9 and Trichoderma pseudokoningii
with the highest mean dry weight of 43.91 g, 41.50 g. and 41.33 g, respectively in that order, least dry weights were observed in treatments with
Fusarium sp3 and Ulocladium sp, that is 18.67 cm and 15.67 g respectively
Table 10. Table 10. Effect of endophytic fungi on plant fresh and dry weight
Treatment Mean fresh weight Mean dry weight
Nigrospora sp
134.33 ± 14.75 a 43.91 ± 2.15 a
Isolate XP 9 131.33 ± 18.15 ab
41.50 ± 1.93 ab Trichoderma pseudokoningii
130.00 ± 19.98 ab 41.33 ± 3.70 ab
Fusarium oxysporum 121.33 ± 15.62 b
39.08 ± 3.62 bc Sterile black 1
107.25 ± 10.95 c 36.17 ± 4.76 c
Trichoderma hamatum 106.50 ± 15.05 c
37.92 ± 4.50 bc Torula
sp 104.08 ± 11.21 c
37.50 ± 4.54 c Chrysosporium
sp 103.83 ± 15.49 c
38.67 ± 3.60 bc Sterile black 2
102.17 ± 4.46 c 36.08 ± 4.54 c
Fusarium chlamydosporum 97.92 ± 17.05 c
38.17 ± 4.69 bc Fusarium
sp 3 70.08 ± 4.46 d
18.67 ± 4.70 d Ulocladium
sp 69.17 ± 4.28 d
15.67 ± 4.44 d Control Without endophytic fungi
27.58 ± 1.78 e 11.25 ± 2.83 e
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05,
according to Duncan’s Multiple Range Test
4.1.4.6 Effect on number of fruits and root length RKN free plants
The data on mean number of fruits indicated higher significant increase in number of fruits by Nigrospora sp, isolate XP 9, Trichoderma pseudokoningii,
Trichoderma hamatum, and Chrysosporium sp with mean number of fruits
42 of 12.42, 12.42, 12.25, 12.00 and 11.83 respectively in that order, least mean
number of fruits were observed in treatments with Ulocladium sp, and Fusarium sp 3 that is 4.00 and 3.92 respectively.
The data also indicates that all endophytic fungi isolates used were highly significant in increasing root length development in comparison to the control
treatments, whereby Nigrospora sp, isolate XP 9, Trichoderma pseudokoningii had the highest mean root length of 41.58 cm, 41.00 cm and 39.42 cm
respectively, least root length was observed in treatments with Ulocladium sp, and Fusarium
sp3 that is 17.83 cm and 16.25 cm respectively Table 11. Table 11. Effect of endophytic fungi on number of fruits and root length
Treatments No. of fruits
Root length cm Nigrospora
sp 12.42 ± 1.44 a
41.58 ± 1.96 a Isolate XP 9
12.42 ± 2.43 a 41.00 ± 1.93 ab
Trichoderma pseudokoningii 12.25 ± 1.66 a
39.42 ± 3.70 abc Trichoderma hamatum
12.00 ± 1.86 ab 36.92 ± 4.15 cde
Chrysosporium sp
11.83 ± 1.90 abc 37.50 ± 4.50 bc
Torula sp
10.42 ± 2.16 bcd 33.50 ± 4.58 e
Fusarium chlamydosporum 10.08 ± 2.64 cd
38.25 ± 3.63 abcd Sterile black 2
10.08 ± 2.58 cd 33.42 ± 4.54 e
Sterile black 1 10.00 ± 2.70 d
34.75 ± 5.68 de Fusarium oxysporum
8.92 ± 2.28 d 38.75 ± 6.26 abc
Ulocladium sp
4.00 ± 1.41 e 17.83 ± 4.70 f
Fusarium sp 3
3.92 ± 1.31 e 16.25 ± 4.79 f
Control Without endophytic fungi 1.42 ± 1.31 f 8.92 ± 2.43 g
Mean ± Standard deviation Means in the same column followed by the same letter are not significantly different at p0.05,
according to Duncan’s Multiple Range Test
4.1.5 Antibiosis In vitro Test
Culture filtrate of the twelve different isolates of endophytic fungi significantly caused mortality of RKN juveniles in vitro. Significant differences
were observed between the control treatments and the endophytic fungi culture filtrate at different concentrations Table 12. The percentage of dead RKN
juveniles was different between the independent treatments at different concentrations, indicating that the endophytic fungi effect on juvenile mortality
varied between repeat experiments with different concentrations.
43 The effect of endophytic fungi culture filtrate on RKN juvenile mortality
was also influenced by the increase in concentration and length of exposure of RKN juveniles to culture filtrate Table 13. The percentage of dead RKN
juveniles differed significantly at each exposure time. Dead RKN juveniles spread straight uncurved with elongated bodies and could not gain life when
transfered back to sterile distilled water supplied with aerator while live RKN juveniles retained the normal sigmoid shape and were moving when placed back
to the sterile distilled water supplied with the aerator. At 30 culture filtrate concentration mortality rate were observed to be low,
highest mortality rates at this concentration were observed in Nigrospora sp where the mean mortality rate were 34.25 after 6 hours, 39.25 after 12 hours,
40.50 after 18 hours and 43.00 after 24 hours, followed by isolate XP 9 where the mean mortality rates were 31.50 after 6 hours, 35.50 after 12 hours,
38 after 18 hours and 43 after 24 hours, while least mortality rates were observed in Ulocladium sp and Fusarium sp3. It was also evident that mortality
rate increased with increase in exposure time Table 12. At 50 culture filtrate concentration mortality rates were observed to be
moderately effective in some culture filtrate with the highest mortality rate at this concentration being observed in Trichoderma pseudokoningii, Nigrospora sp,
isolate XP 9, Trichoderma hamatum, Chrysosporium sp where the mean mortality rates were 75.25 after 6 hours, 80.25 after 12 hours, 85.25 after
18 hours and 88.75 after 24 hours for Trichoderma pseudokoningii, mean mortality rates were 74.25 after 6 hours, 86.50 after 12 hours, 90.00 after
18 hours and 96.00 after 24 hours for Nigrospora sp, mean mortality rates were 71.50 after 6 hours, 81.75 after 12 hours, 87.75 after 18 hours and 93.00
after 24 hours for isolate XP 9, mean mortality rates were 73.25 after 6 hours, 76.75 after 12 hours, 83.25 after 18 hours and 86.50 after 24 hours for
Trichoderma hamatum and mean mortality rates were 65.25 after 6 hours, then
69.50 after 12 hours, 74.00 after 18 hours and 79.25 after 24 hours for Chrysosporium
sp. Least mortality rates were observed in Ulocladium sp and Fusarium
sp3 Table 12.
44 Juvenile mortality at 90 culture filtrate concentration were observed to be
highly effective in all culture filtrate concentrations except in Ulocladium sp and Fusarium
sp3. Most effective mortality rate at 90 culture filtrate concentration were observed in Nigrospora sp, isolate XP 9, Trichoderma hamatum,
Trichoderma pseudokoningii, where the mean mortality rates were 100 from 6
hours of exposure period of the RKN juveniles to the culture filtrate at 90 concentration, on the other hand least mortality rates were observed in
Ulocladium sp and Fusarium sp3 Table 12. There were no mortality rates of the
RKN juveniles observed after 24 hours period in the two control treatments sterile distilled water and PDB indicating that the that the effect on mortality of
RKN juvenile were neither influenced by sterile distilled water nor PDB. Mortality rate was observed to be higher in the 90 culture filtrate
concentration, followed by the 50 culture filtrate concentration, but was observed to be low in the 30 culture filtrates concentration. Microwells
containing culture filtrates of Trichoderma pseudokoningii, Nigrospora sp and isolate XP 9 showed complete mortality rates by 100 of the RKN juvenile after
the first 6 hours at 90 culture filtrate concentration, this indicated that the culture filtrates were very effectivity within short period of exposure time. While
on the other hand RKN juvenile mortality rates were observed to be lower in microwells with culture filtrates of Ulocladium sp and Fusarium sp 3 Figure 6.
45
Figure 6. RKN juvenile percentage mortality rate in different culture filtrate concentrations 45
46 Table 12. Effectiveness of different culture filtrate on juvenile mortality rate in-vitro at different concentration and time exposure
Mortality Rate of RKN Juveniles Treatment
Conc 6 hrs
12 hrs 18 hours
24 hours Nigrospora
sp 90
100.00 ± 0.00 a 100.00 ± 0.00 a
100.00 ± 0.00 a 100.00 ± 0.00 a
50 74.25 ± 9.32 ab
86.50 ± 6.14 a 90.00 ± 5.23 a
96.00 ± 2.83 a 30
34.25 ± 5.19 a 39.25 ± 4.03 a
40.50 ± 3.42 a 43.00 ± 1.83 ab
Isolate XP 9 90
100.00 ± 0.00 a 100.00 ± 0.00 a
100.00 ± 0.00 a 100.00 ± 0.00 a
50 71.50 ± 7.05 ab
81.75 ± 2.89 ab 87.75 ± 6.29 ab
93.00 ± 6.63 ab 30
31.50 ± 3.32 ab 35.50 ± 2.89 ab
38.00 ± 4.55 ab 43.50 ± 6.35 a
Trichoderma hamatum 90
100.00 ± 0.00 a 100.00 ± 0.00 a
100.00 ± 0.00 a 100.00 ± 0.00 a
50 73.25 ± 9.85 ab
76.75 ± 9.32 bcd 83.25 ± 10.28 abc 86.50 ± 6.46 bcd
30 27.50 ± 4.65 bc
32.25 ± 3.69 bc 36.75 ± 4.11 ab
42.25 ± 3.40 ab Trichoderma pseudokoningii
90 100.00 ± 0.00 a
100.00 ± 0.00 a 100.00 ± 0.00 a
100.00 ± 0.00 a 50
75.25 ± 7.18 a 80.25 ± 5.91 abc
85.25 ± 7.50 ab 88.75 ± 6.60 abc
30 25.75 ± 5.62 c
30.50 ± 3.70 c 35.00 ± 3.56 b
37.50 ± 4.20 b Chrysosporium
sp 90
83.00 ± 3.46 b 85.25 ± 4.35 b
88.75 ± 4.50 b 94.25 ± 3.86 b
50 65.25 ± 5.56 abc
69.50 ± 6.56 de 74.00 ± 3.56 cd
79.25 ± 3.30 d 30
13.50 ± 3.11 d 17.25 ± 4.92 d
21.75 ± 6.50 c 25.25 ± 7.68 c
Fusarium chlamydosporum 90
76.50 ± 5.74 bc 82.25 ± 4.92 bc
83.25 ± 5.06 bc 87.25 ± 3.95 c
50 58.25 ± 9.88 c
66.25 ± 6.75 e 73.00 ± 11.02 cd
83.25 ± 5.56 cd 30
10.75 ± 2.99 def 12.00 ± 1.41 e
14.00 ± 0.82 d 16.25 ± 2.22 d
Mean ± Standard deviation Means in the same column and with the same concentration followed by the same letter are not significantly different at p0.05, according to Duncan’s
Multiple Range Test.
46
47
Table 12. Effectiveness of different culture filtrate on juvenile mortality rate in-vitro at different concentration and time exposure Mortality Rate of RKN Juveniles
Treatment Conc
6 hrs 12 hrs
18 hours 24 hours
Fusarium oxysporum 90
71.50 ± 10.72 c 85.75 ± 8.99 b
90.25 ± 6.24 b 94.75 ± 4.35 b
50 57.50 ± 9.40 c
67.00 ± 5.24 e 70.75 ± 8.99 d
81.50 ± 7.59 cd 30
9.50 ± 1.29 def 11.50 ± 1.29 e
13.50 ± 1.29 d 15.75 ± 1.71 d
Sterile black 1 90
76.75 ± 11.70 bc 77.25 ± 12.15 bc
90.50 ± 3.11 b 94.00 ± 0.00 b
50 63.25 ± 8.77 bc
72.50 ± 10.08 cde 78.50 ± 8.10 cd
83.00 ± 5.35 cd 30
11.25 ± 1.89 de 11.50 ± 1.73 e
13.25 ± 1.71 d 14.50 ± 1.29 d
Sterile black 2 90
70.75 ± 11.12 c 74.00 ± 10.86 c
81.00 ± 4.62 c 84.25 ± 3.78 c
50 26.50 ± 4.93 d
30.25 ± 3.95 f 35.75 ± 5.72 e
39.00 ± 5.72 e 30
8.00 ± 4.24 ef 10.25 ± 4.27 e
11.50 ± 0.58 d 12.25 ± 1.26 d
Torula sp 90
58.25 ± 9.88 d 73.00 ± 9.87 c
78.00 ± 4.35 c 85.25 ± 4.35 c
50 27.50 ± 4.65 d
32.25 ± 3.69 f 36.75 ± 4.11 e
42.25 ± 3.40 e 30
5.50 ± 1.73 f 7.75 ± 2.22 e
10.50 ± 2.38 d 11.50 ± 2.52 d
Ulocladium sp
90 26.50 ± 4.93 e
30.25 ± 3.95 d 35.75 ± 2.99 d
38.00 ± 4.55 d 50
7.50 ± 2.08 e 10.25 ± 2.36 g
11.75 ± 1.50 f 12.75 ± 2.06 f
30 0.00 ± 0.00 g
0.00 ± 0.00 f 1.00 ± 0.82 e
1.75 ± 0.50 e Fusarium sp
3 90
27.50 ± 4.66 e 32.25 ± 3.69 d
36.75 ± 4.11 d 42.25 ± 3.40 d
50 10.25 ± 1.29 e
12.50 ± 1.29 g 14.50 ± 1.73 f
18.25 ± 2.21 f 30
0.00 ± 0.00 g 0.00 ± 0.00 f
1.00 ± 1.41 e 3.00 ± 0.82 e
Control Sterile distilled Water Control PDB
Mean ± Standard deviation Means in the same column and with the same concentration followed by the same letter are not significantly different at p0.05, according to
Duncan’s Multiple Range Test.
47
48
4.2 Discussion 4.2.1 Exploration of Endophytic Fungi
Endophytic fungi were abundant in roots of healthy tomato plants 50 than in nematode infected plants 15, this was similar to research conducted by
Petrini 1991 indicating that species composition of endophytic communities in healthy plants is higher than species composition of endophytic fungi in infected
plants. Caroll 1988 and Clay 1988 also determined that the association between endophytic fungi and host plants increases the plant resistance to most
pathogens, other publications by Niere 2001 also showed that endophyte treated plants had lower total number of nematode densities.
Most potential endophytic fungi were obtained from lowland than in highland areas, this may be directly related to the climatic factors and correlates to
altitude, these findings can be related to research outcome of Sieber 1988 who demonstrated that frequency of colonization by endophytic fungi decreases with
increasing altitude in Norway spruce Picea abies L. Karst needles and by low temperature in annual shoots of scot pine Pinus silvesris L. and Norway spruce
Barklund Unestam 1988. Diversity index analysis indicated higher diversity of endophytic fungi in
healthy tomato plants 0.926 than in nematode infected plants 0.645 this was according to Simpson’s index Simpson 1949. It would seem that many
processes affect the endophytic fungi diversity patterns of healthy and infected tomato plants. Fungal diversity seems to be related most strongly to habitat and
host plants, although the abundance of resources might also be important in certain groups. Publications by Sieber-Canavesi et al. 1988 and Petrini 1987
stated that species composition of endophyte communities varies in relation to their collection sites and host plants. On the other hand the similarity index of
endophytic fungi between healthy and infected plants was low 0.154 indicating that the endophytic fungi obtained from healthy and nematode infected plants
were different, this was according to Sorensen index Sorensen 1948 modified by Bray Curtis 1957.
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