In vitro Studies on Physiology of Fungi Isolated from Stem pieces of Cuscuta in Pakistan, KH Wüstner

Tags: mycelial growth, fungi, stem cuttings, temperatures, C. Iunata, pH levels, darkness, Cuscuta, culture media, petri plates, fg, gm, Plant Diseases, Braunschweig, Germany, culture collections, Technical information, Islamabad, Pakistan, J. Biol, developing countries, universal medium, Agar Agar, Potato Dextrose Agar, fungal organisms, Pakistan Agricultural Research Council
Content: Pakistan Journal of Biological Sciences, 2 (3): 993-995, 1999
RESEARCH ARTICLE
In vitro Studies on Physiology of Fungi Isolated from Stem pieces of Cuscuta in Pakistan
Iqbal M.Z., A.S. Shakir and S.T. Sahi Department of Plant Pathology, University of Agriculture, Faisalabad-38040, Pakistan
Abstract In vitro studies on physiology of four fungi namely Alternaria alternata, Colltotrichum gloeosporioides, Curvularia lunata and Fusarium pallidoroseum isolated from Cuscuta stem cuttings were conducted. All the fungi gave the best mycelial growth on universal medium. Out of these fungi A. alternata and F. pallidoroseum gave the best growth at 25EC and C. gloeosporioides gave the best mycelia! growth at 20EC while C. gloeosporioides required pH 6 for better mycelia! growth. 24 hours continuous light favoured the growth of A. alternata and C. glpeosporioides while C. lunata and F. pallidoroseum gave better results under 12 hours light and 12 hours darkness.
Introduction Species of Cuscuta are widely distributed throughout the Pakistan which parasitize a variety of hosts, belonging to different botanical families. Citrus plantation specially khatti (Citrus limon) and hedges (Clerodendrum inerme) are severely infested by Cuscuta (Hafiz, 1986). In Pakistan, no attention has been paid to control this severe phanerogamia parasite which is urgent need of time. Malik (1992) found that universal medium was proved to be the best medium for many fungi. Tariq et al. (1993) conducted an experiment on physiological studies of Botrytis gladiolorum and concluded that this fungus grew best on PEA, at 25EC, pH 7 and in continuous darkness. Ehsan-ul-Haq et al. (1998) found universal medium to be the best for mycelial growth of Botrytis cinerea, Curvularia lunata, Fusarium oxysporum f. sp. gladioli and Stemphylium botryosum 25EC temperature was found better for all the fungi except Curvularia lunata which grew best at 30EC. A pH of 6 favoured the growth of B. cinerea and S. botryosum gave good mycelial growth under 24 hours continuous light while mycelial growth of other two fungi was observed under 12 hours alternate cycle of light and darkness. Recently four fungi namely Alternaria alternata, Colletotrichum gloeosporioides, Curvularia lunata and Fusarium pallidoroseum have been reported from Cuscuta stem (Shakir et al., 1999) but their biology is yet uninvestigated. A very little work has been done on biology of fungi associated with Cuscuta, therefore it was planned to study the effect of different media, temperature, light regimes and pH levels on growth of fungi associated with Cuscuta stem. Materials and Methods Pure cultures of Alternaria alternata, Colletotrichum gloeosporioides, Curvularia lunata and Fusarium pallidorseum isolated from stem cuttings of Cuscuta, were collected from culture collection, Department of Plant Pathology, University of Agriculture, Faisalabad (Pakistan) to carried out physiological studies. Mycelial growth of
fungus was observed on universal medium (Malt-extract = 30 gm, soya peptone = 3 gm, Agar = 15 gm, distilled water = 1000 ml) Potato Dextrose Agar (Potato starch = 20 gm, Dextrose = 20 gm, Agar Agar = 20 gm, Distilled water = 1000 ml, corn meal agar (Corn meal = 17 gm, Agar Agar = 5 gm, Distilled water = 1000 ml) and on water Agar with Cuscuta stem were boiled in 500 ml of water for 30 minutes. Agar was dissolved in another 500 ml of water and both the solutions were mixed to make the volume of water one liter. The above mentioned four media were prepared separately and autoclaved for 15 minutes at 15 lbs pressure and 121EC temperature. Chloramphenicol was used as an antibacterial at the 0.05 g/liter of medium. About 20 ml of each of the sterilized medium was poured aseptically into each petri plate (9 cm). On solidification four plates of each medium were inoculated with 5 mm culture discs of each fungus in the centre, cut with a sterilized cork borer (5 mm) aseptically from fresh cultures of fungi namely A. aftemata, C. gloeosporioides, C. lunata and F. pallidoroseum. Inoculated petri plates were incubated at 25EC. Experiment was run in quadruplicate following factorial arrangement. To determine the most suitable temperature for the growth of above mentioned fungi, autoclaved universal medium was used, as it proved to be the best among different media tested for fungal growth. Petri plates thus inoculated with 5 mm discs of actively growing cultures of four fungi were incubated at 15, 20, 25, 30 and 35EC. Data regarding colony diameter of each fungus were recorded on eighth day of inoculation. Mycelial growth of the fungi was also studied on pH levels of 5, 6, 7, 8, 9 and 10 pH, which were adjusted by the addition of N/10 HCI or (1N) NaOH solution. Medium for respective pH was autoclaved and adjusted again for its pH and poured aseptically in petri plates. Each petri plate was inoculated with fresh culture discs (5 mm) of different fungi in the centre and then placed at 25EC except Curvularia lunata which required 30EC. Data were recorded on eighth day of inoculations. To study the effect of duration of light on mycelial growth of fungi, autoclaved universal medium was poured
993
lqbal et al.: In vitro, physiology, fungi, Cuscuta, Pakistan
aseptically in petri plate (9 cm) and inoculated with fresh culture discs (5 mm) of different fungi and placed at 25EC except C. lunata which required 30EC. Following light regimes were analysed 1. 24 hours light 2. 16 hours light + 8 hours darkness 3. 12 hours light + 12 hours darkness 4. 8 hours light + 16 hours darkness 5. 24 hours darkness Source of light was fluorescent tube at the distance of 41 cm from petri plates.
Results and Discussion Effect of different culture media on mycelial growth: Universal medium proved to be the best medium for mycelial growth of A. alternata, C. lunata, C. gloeosporioides and F. paffidoroseium (Table 1). Malik (1992) also found the same medium to be the best for many fungi. Effect of different temperatures on mycelial growth: Impact of temperature on the growth of different fungi was different (Table 2). The study revealed that 25EC temperature was the best for A. alternata and F. pallidoroseum and 20EC was the best temperature for
Table 1: Effect of different culture media on mycelial growth (mm) of different fungi isolated from stem cuttings of Cuscuta
Name of fungi
---------------------------------------------------------------------------------------------------------------------------------------------------------
Media
F. pallidoroseum
A. alternata
C. gloeosporioides
C. Iunata
1
75.63 a
58.94 of
65.19 cd
73.25 ab
2
69.63 bc
50.50 gh
60.88 de
66.44 c
3
55.31 fg
40.38 I
49.75 h
48.44 gh
4
60.88 de
39.75 I
46.94 h
41.19 I
Table 2: Effect of different temperatures on mycelial growth (mm) of different fungi isolated from stem cuttings of Cuscuta
Name of fungi
---------------------------------------------------------------------------------------------------------------------------------------------------------
Temperature F. pallidoroseum
A. alternata
C. gloeosporioides
C. Iunata
15
42.13 g*
45.00 f
62.38 d
45.25 f
20
60.94 d
56.19 e
80.19 a
61.13 d
25
74.69 b
71.06 c
72.19 bc
70.69 c
30
62.31 d
71.44c
61.56 d
80.81 a
Any two figures sharing similar letter (s) do not differ significantly at p = 0.05
Table 3: Effect of different pH levels on mycelial growth (mm) of different fungi isolated from stem cuttings of Cuscuta
Name of fungi
---------------------------------------------------------------------------------------------------------------------------------------------------------
pH levels
F. pallidoroseum
A. alternata
C. gloeosporioides
C. Iunata
5
64.81 efg
63.00 g
63.44 fg
64.94 efg
6
76.94 c
74.94 cd
81.31 b
73.88 d
7
84.88 a
83.00 ab
77.25 c
85.31 a
8
66.88 e
73.31 d
73.63 d
67.00 e
9
55.19 h
63.50 fg
66.00 of
55.75 h
10
42.38 j
45.69 I
44.19 ij
36.19 k
Any two figures sharing similar letter (s) do not differ significantly at p = 0.05
Table 4: Effect of different light regimes on mycelial growth (mm) of different fungi isolated from stem cuttings of Cuscuta
Name of fungi
Light
---------------------------------------------------------------------------------------------------------------------------------------------------------
intensities
F. pallidoroseum
A. alternata
C. gloeosporioides
C. Iunata
1
74.38 c*
79.13 b
82.44 a
66.2564
2
71.44 d
73.19 cd
73.63 cd
63.13 g
3
84.88 a
78.13 b
67.56 e
84.00 a
4
71.81 cd
66.25 ef
63.63 fg
57.44 h
5
65.25 efg
63.44 fg
53.88 I
55.38 h
1. 24 hours light, 2. 16 hours light + 8 hours darkness, 3. 12 hours light + 12 hours darkness, 4. 8 hours light + 16 hours
darkness, 5. 24 hours darkness
Pak. J. Biol. Sci., 2 (3): 993-995, 1999
994
lqbal et al.: In vitro, physiology, fungi, Cuscuta, Pakistan
C. gloeosporioides while C. lunata gave different response as compared to other fungi and preferred 30EC. McClellan and Marshall (1950) also concluded in his study that about 30EC was the best temperature for C. lunata. Effect of pH levels on mycelial growth: pH 7 suitable for A. alternata, C. lunata and F. pallidoroseum, while pH 6 was proved to be the best for growth of C. gloeosporioides (Table 3). Effect of different light regimes on mycelial growth: C. lunata and F. pallidoroseum gave good growth under 12 hours light and 12 hours darkness while A. alternata and C. gloeosporioides gave better mycelial growth under continuous light (Table 4). Our these results verifies the results of Tariq et al. (1993) and Ehsan-ul-Haq et al. (1998) who obtain the same results. References Ehsan-ul-Haq, A., S. Shakir and S.T. Sahi, 1998. In vitro studies on physiology and chemical control of fungi isolated from Corms and leaves of gladiolus in Pakistan. Pak. J. Biol. Sci., 1: 97-100.
Hafiz, A., 1986. Plant Diseases. Pakistan Agricultural Research Council, Islamabad, Pakistan, Pages: 390. Malik, K.A., 1992. technical information for culture collections curators in developing countries. UNESCO/WFCC Education Committee. Braunschweig, Germany, pp: 40. McClellan, W.D. and B.H. Marshall, 1950. Effect of temperature on the development of some diseases of Gladiolus, Narcissus and Lilium. Phytopathology, 40: 872-872. Shakir, M.Z. Iqbal and S.T. Sahi, 1999. First report on association of some fungal organisms with dodder (Cuscuta) blight from Pakistan, Pak. J. Biol. Sci., 2: 991-992. Tariq, M., J.H. Mirza and A.S. Shakir, 1993. Physiological studies of Botrytis gladiolorum and its in vitro sensitivity to fungicides. Pak. J. Phytopathol., 5: 89-92.
Pak. J. Biol. Sci., 2 (3): 993-995, 1999
995

KH Wüstner

File: in-vitro-studies-on-physiology-of-fungi-isolated-from-stem-pieces.pdf
Title: In vitro Studies on Physiology of Fungi Isolated from Stem pieces of Cuscuta in Pakistan
Author: KH Wüstner
Author: Iqbal M.Z., A.S. Shakir and S.T. Sahi
Subject: Pakistan Journal of Biological Sciences
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