- Email: [email protected]
pH and temperature optima for growth and sporulation in some common fungi from city waste
243
Mycoscience 36: 243-246, 1995
Short Communication
pH and temperature optima for growth and sporulation in some common fungi from city waste Sudhir Kumar M e h r a and Ajai K u m a r Jaitly Department of Plant Science, Faculty of Life Sciences, Rohilkhand University, Bareilly-243001, India Accepted for publication 28 April 1995
Relationships between the growth of certain fungi isolated from city waste and pH and temperature were examined by two methods. The tested isolates showed their maximum growth and sporulation at different pHs while temperature requirements were the same (28°C), except for Humicola grisea (43°C). Cladosporium herbarum and H. grisea showed double pH optima. The ranges of pH and temperature for sporulation were more limited than those for the vegetative growth. Although all the tested isolates showed wide tolerances to pH and temperature, the degree of tolerance varied with the isolates. A considerable change from the initial pH of the liquid medium was noted at the end of the experiment. Key Words
city waste; fungal growth; pH; temperature.
The environment in which organisms exist exerts a profound effect upon them and often dictates their behaviour. The microbial population of a habitat is influenced either singly or in combination by a number of biotic and abiotic factors. City waste provides a complex habitat supporting an abundant and extremely diverse population of microorganisms. There is considerable evidence that ecological conditions play an important role in microbial activity (Hudson, 1968; Rai et al., 1981; Jaitly, 1982, 1987), of which hydrogen ion concentration and temperature are very important in determining the growth, activity and composition of organisms (Alexander, 1961 ; Agnihotri, 1963, 1964; Mukerjee, 1966). Individual species may have lower or higher tolerance to these factors than the general fungal community (Jaitly, 1982, 1987). Hegarty and Curran (1985) observed that maximum wood decay by most of their isolates was in the temperature and pH ranges of 1 5 - 2 5 ° C and 5-8, respectively. Investigations have been conducted on pH and temperature optima of fungi from various habitats, but no report is available on these lines for the fungi from city waste. Furthermore, because of authors' interest in biodegradation and enzymatic hydrolysis of composting material, it was essential to find the optimal pH and temperature for the experimental fungal isolates. Effects of pH and temperature on the growth and sporulation of some common fungi from city waste were evaluated by a diameter measurement method because this has been shown to be of sufficient reliability (Brancato and Golding, 1953; Jaitly, 1982, 1987). Dry weight measurements were also taken (Jaitly, 1987). Czapek-Dox agar was prepared in citrate-phosphate buffer of different pH values ranging from 3.0 to 8.0 at in-
tervals of 0.5 unit. Sterilized medium was poured in sterilized Petri dishes (10 cm) and single point central inoculations were made with 2 mm discs of freshly grown mycelium of the test fungus. Three replicates of each isolate were grown at a constant temperature of 29__+ 1 °C. Plates were regularly examined at 24-h intervals for seven days. Two diameters were measured at right angles to each other as a measure of the average diameter for each colony. Isolates were examined for sporulation regularly. In the second method, 25 mt of Czapek-Dox medium of required pH values was poured in 100-ml Erlenmeyer flasks and autoclaved at 15 Ibs pressure for 15 min. The flasks were inoculated with 0.5 ml of homogenous spore suspension of the test fungus containing 107spores/ml. The flasks were incubated at 29___ 1°C. T w o replictes were taken for each. After seven days of incubation, the biomass obtained was filtered through preweighed Whatman filter paper No. 1 and dried for 24 h at 80°C. Final weight was taken to calculate the dry weight of fungal biomass. Test fungi were grown on Czapek-Dox medium at the optimum pH for their growth at different tempera, tures in the same way as stated for pH. The results of pH-growth relationship from colony diameter measurements were in agreement with those of the dry weight measurements. Tested isolates showed their maximum growth at different pHs (Table 1). After an initial lag phase, the increase in diameter was more or less linear for all the tested fungal isolates (Fig. 1 ) at pH near to their optima. The isolates grew optimally in the acidic to neutral pH range but also showed tolerance to alkaline pH. Slow growth was observed at pH 3.0 for all the isolates. Maximum growth for all the tested isolates
244
S.K. Mehra and A. K. Jaitly Table 1.
Effect of pH on growth of some fungi after seven days (dry weight measurement). Dry weight in mg*
Fungus 3.0
3.5
4.0
4.5
5.0
5.5
Altemaria hurnicola Oudemans
130
150
175
Aspergillus niger van Tieghem
120 +
180 +
240 4-F
Aspergillus terreus Thom
390 -iF
440 4+4-
520 II I I l
Cladosporiurn herbarum (Pers.) Link ex Gray
150
190
290
Curvularia lunata (Wakker) Boedijn
180 --
185 +
t I I t
230 lit 180 TII 535 Illl 330 4-4360 11t 445 I It 80 + 230 H-r-
360 IIlI 210 Itl 580 Iitti 450 ~ 340 I[I 490
290 I1t 230 II I 530 I I Ill 480 III 330 Ilt 380
pH**
Humicola grisea Traaen
320
330
300
30
45
75
190
200
210
--
Myrothecium roridum Tode ex Fr. Penicillium chrysogenurn Thorn
450
+
- -
4-+
44-
I[l
140 ~ 245 4~-
I
Iit
160 ++ 260 ill
6.0
6.5
240 200 4-~ -320 200 I I}t Iti 510 470 I t It III 580 560 t I t I t I ~11 I 325 210 -H-
-F+
360
380 4-F 90 + 380 I I[l
-H-
190 H-F 350 ++-t-t-
7.0
8.0
100 -180 ~+ 420 4-4390 ttt 260 .... 480 ÷-~ 50 .... 230 I II
110 100 35O 4-F 510 4-F 25O 35O 4-F 45 260 4-F
* Growth on Czapek-Dox medium. ** Citrate phosphate buffer. + , -- Degree of sporulation.
w a s obtained at pH b e t w e e n 4 . 0 and 6.5. Humicola grisea Traaen and Cladosporium herbarum (Pers.) Link ex Gray s h o w e d double pH o p t i m a , one in the acidic range (pH 5.0 and 6.0, respectively) and the other at neutral and alkaline pH i.e., p H 7 and p H 8 , respectively. M i c r o s c o p i c e x a m i n a t i o n of tested isolates revealed that the pH range for sporulation is n a r r o w e r than t h a t for the v e g e t a t i v e g r o w t h . Normally, excellent sporulation w a s o b t a i n e d b e t w e e n pH 4 . 0 and 5.5. All the t e s t e d isolates w e r e observed t o g r o w at pH 8 . 0 w i t h varying colony diameter, indicating t h a t t h e y have different abilities to tolerate e x t r e m e pH. A considerable change f r o m the initial pH of the liquid m e d i u m w a s noted (Table 3). T e m p e r a t u r e requirements w e r e same for all the test-
pH
80-
4.0 5.0
E
v
60-
6.0
t~ i,i FLIJ 7.0
40>Z
o._J o
8.0 209.0
I
f
[
I
48
96
144
192
TIME (]1) Fig.
Cardinal growth of Curvularia lunata on Czapek-Dox agar medium. In relation to pH. 1.
ed fungat isolates, e x c e p t for H. grisea (Table 2). A f t e r an initial lag phase, colony d i a m e t e r increased linearly w i t h t i m e for all the isolates (Fig. 2). Tested fungat isolates s h o w e d o p t i m u m g r o w t h at 2 8 ° C . H o w e v e r the mycetial g r o w t h of H. grisea w a s m a x i m u m at 4 3 ° C . M o s t of the fungi s h o w e d l o w e r g r o w t h at b e l o w 2 3 ° C and above 3 3 ° C . Sporulation w a s also m a x i m u m in the t e m p e r a t u r e range of 2 8 - 3 3 ° C . The t w o m e t h o d s used t o d e t e r m i n e g r o w t h in relation to pH or t e m p e r a t u r e w e r e s a t i s f a c t o r y and yielded similar results. A t e x t r e m e s of pH and t e m p e r a t u r e , the isolates g r e w only after a long incubation period. The existence of m i c r o o r g a n i s m s in the w a s t e materials is affected by a number of environmental factors such as pH, t e m perature, aeration, moisture, organic m a t t e r etc., of w h i c h pH and t e m p e r a t u r e are the m o s t i m p o r t a n t (Hegart y and Curran, 1 9 8 5 ; Pearce and Malajczuk, 1 9 9 0 ; B o w e r m a n and Goos, 1991). All the tested isolates s h o w e d their pH o p t i m a in the range of 4 . 0 t o 6.5. This m i g h t be due to the presence of high c o n c e n t r a t i o n s of organic m a t t e r w h i c h d e c o m p o s e s to f o r m different nitrogenous and carbonaceous substances, thus l o w e r ing t h e pH of w a s t e material. These studies are in agreem e n t w i t h those p e r f o r m e d by Jaitly (1987). A l e x a n d e r (1971) also stated t h a t the application of fertilizers containing nitrogenous substances leads to the f o r m a t i o n of acids by fungi on one hand and inhibition of the g r o w t h of bacteria and a c t i n o m y c e t e s on the other hand. Tested isolates s h o w e d some tolerance t o alkaline pH but their o p t i m u m g r o w t h lay in the l o w e r range. This is in a g r e e m e n t w i t h the results obtained by several w o r k e r s (Wolf et al., 1950; Brewer, 1960; Rai et at., 1970, Jaitly, 1987). In the present investigation, C. herbarum and H. grisea s h o w e d double pH o p t i m a . Biomodal curves have also been obtained by several o t h e r w o r k e r s (Jones and Irvine, 1972; Jaitly, 1987).
pH and temperature optima
Table 2.
245
Effect of temperature on growth of some fungi after seven days (dry weight measurement). Dry weight in mg*
Fungus Temperature °C
23
28
33
38
43
48
Altemaria hum/cola
200 44-
310 IIi
230 4-I-
220 +4-
190 +
140 --
Aspergillus niger
190 +4-
450 Jilt
330 ~
240 4-+4-
210 +4.
180 -FF
Aspergillus terreus
140 4-F
550 I ] 111
460 I I I I
370 II I
180 44-
070 4-
Cladosporium herbarum
240 -IF
420 I II I
380 I I I I
320 I] I
270 4-F
230
Curvularia tunata
250
540
490
430
320
180
I I F
I I ] q q
I I I I
Itl
4W
4-
Hum/cola grisea
150 --
250 ill
280 ~
320 444-}-
390 r Fli
310 lit
Myrothecium roridum
110 4-{-
270 Ill
190 II~
180 4-+
120 44
1O0 4-
Pen/c/Ilium chrysogenum
230 -FF
380 Itl
350 4-FF
310 +++
270 4-F
210 4-F
* + , --
Growth on Czapek-Dox medium at optimum pH. Degree of sporutation. Table 3.
Shift in the initial pH of basal medium. Final pH values after 7 days of growth
Initial pH
Altemaria humicola
Aspergillus niger
Aspergillus Cladosporium Curvularia terreus herbarum lunata
3.0 3.5
5,0 5.0
4.5 4.5
5.0 5.0
5.5 5.5
4.0 5,0
4.5 4,5
4.0 4.0
4.0 4.0
4.0 4,5
5,5 6.0
5.0 5.5
5.8 6.0
5.5 5.8
5.0 5.5
5.5 5.5
5.0 5.0
4.4 4.6
5.0
6,0
6.0
6.0
6.2
5.0
6,0
5.5
4.6
5,5
6.0
6.0
6.0
6.5
5,5
6.5
5.5
4.6
6.0 6,5
6.O 6.5
6.5 6.5
6.0 6.5
6.5 6.5
5.5 6.0
6,5 6.5
5.5 6.0
4.6 4,8
7,0
7.0
6.8
6,8
6.5
6.0
6.8
6.0
5.0
8.0
7.5
7.0
7.0
6,0
6.5
6.8
6.5
5.0
Fungi are k n o w n t o alter t h e pH o f m e d i u m in w h i c h t h e y g r e w , The p r e s e n t results also s h o w e d a c h a n g e in t h e initial pH of t h e basal m e d i u m . Generally t h e l o w e r pH v a l u e s b e c a m e higher and t h e h i g h e r b e c a m e l o w e r , w h i l e t h o s e in t h e v i c i n i t y o f pH 7 . 0 did n o t shift sign i f i c a n t l y t o either side (Table 3). In t h e p r e s e n t s t u d y t h e species g r e w a f t e r a long inc u b a t i o n period at t h e e x t r e m e t e m p e r a t u r e s . M a n y w o r kers (Tansey, 1 9 7 2 ; J a i t l y , 1 9 8 2 ) s u g g e s t e d t h a t a long lag phase can be e x p e c t e d at i n c u b a t i o n t e m p e r a t u r e s differing f r o m t h a t of t h e i n o c u l u m . Evans ( 1 9 7 1 ) c o n cluded t h a t a s h o r t i n c u b a t i o n period m a y n o t reflect t h e t r u e cardinal t e m p e r a t u r e s for g r o w t h . During t h e p r e s e n t i n v e s t i g a t i o n , all t h e t e s t e d isolates s h o w e d their m a x i m u m g r o w t h at 2 8 ° C , e x c e p t H, grisea, w h i c h s h o w e d o p t i m u m g r o w t h at 4 3 ° C , indicating t h a t t h e species is o f t h e r m o t o l e r a n t nature. The ranges o f pH and t e m p e r a t u r e for s p o r u l a t i o n w e r e l o w e r t h a n t h o s e for vegetative growth (Jatilty, 1982, 1987).
Hum/cola Myrothecium Pen/c/Ilium grisea roridum chrysogenum
80-
TEMP. ( °C ) 28
60-~ r~ uJ FL~J
<
33 38 43 23
40-
>-
z o
_J
0 c.)
48
20-
I
I
I
i
48
96
144
192
T I M E (h) Fig.
Cardinal growth of Curvularia lunata on Czapek-Dox agar medium. In relation to temperature. 2.
246
S.K. Mehra and A. K, Jaitly
Literature cited Agnihotri, V, P. 1963, Studies on Colletotrichum capsci III. The effect of pH on growth and amino acid composition, Mycopathol. Mycol. Appl, 24: 305-314. Agnihotri, V. P. 1964. Studies of Aspergilli XVI. Effect of pH, temperature and carbon and nitrogen interaction. Mycopathol, Mycol, Appl. 25: 3 0 5 - 3 1 4 . Alexander, M, 1961. "Introduction to soil microbiology," John Wiley, New York. 472 p. Alexander, M, 1971. Biochemical ecology of microorganisms. Ann. Rev. Microbiol. 25: 3 6 1 - 3 9 1 . Bowerman, L. and Goos, R. D. 1991. Physiological studies of t w o fungi isolated from Nymphaea odorata. Mycologia 83: 624-632. Brancato, F, P. and Golding, N. S. 1953, The diameter of the mold colony as a reliable measure of growth. Mycologia 45: 8 4 8 - 8 6 4 . Brewer, D. 1960. Studies on AscochytaplsiLib. Can. J, Bot, 38:705-717. Deploey, J, 1992. Some factors affecting germination of Rhizomucor miehei sporangiospores. Mycologia 84: 7 7 81. Evans, H. C, 1971. Thermophilous fungi of coal spoil tips. tl. Occurrence, distribution and temperature relationships. Trans. Br. Mycot. Soc. 57: 2 2 5 - 2 6 6 . Hegarty, B. M. and Curran, P. M. T, 1985, The biodeterioration of beech (Fagus sylvatica) by marine and non-marine fungi in response to temperature, pH, light and dark, Int. Biodeterioration 21 : 11 - 18, Hudson, H. J. 1968. The ecology of fungi on the plant remains
above the soil. New Phytol. 67: 837-874. Jaitly, A. K, 1982. Ecological studies of thermophitic fungi native to mangrove swamps. I. Temperature-growth relationships. Trans. Mycol. Soc. Japan 23: 6 5 - 7 1 , Jaitiy, A. K. 1987. pH optima of the fungi isolated from mangrove soils in India. Trans. Mycol. Soc. Japan 28: 157t63. Jones, E. B. G. and Irvine, J. 1972. The role of marine fungi in the biodeterioration of materials. In: "Biodeterioration of materials," (ed. by Waiters, A. H. and Hueck-Vender Plas, E, H.), pp. 4 2 2 - 4 3 1 . Applied Science, London. Mukerjee, K.G. 1966. Studies on the effect of hydrogen ion concentration on growth and sporulation of certain soil fungi. Mycopathol. Mycol. Appl. 28: 3 1 2 - 3 1 6 . Pearce, M, H. and Malajczuk, N. 1990. Factors effecting growth of Armillaria luteobubalina rhizomorphs in soil. Mycol, Res, 94: 3 8 - 4 8 . Rai, J.N.,Garg, K. L a n d J a i t l y , A.K. 1981. Saprophytic fungi from woods in mangrove swamps and their wood decaying capability. Trans. Mycol. Soc. Japan 22: 6 5 - 7 4 . Rai, J. N., Sharma, B. B. and Agarwal, S.C. 1970, Increased pH tolerance of some Aspergilli isolated from 'Usar' (alkaline) soils--A possible indication of ecological specialization. Sydowia 24: 3 3 6 - 3 4 3 . Tansey, M, R. 1972. Effect of temperature on growth rate and development of the thermophilic fungus--Chaetornium thermophile. Mycologia 64: 1290-1299. Wolf, F. T., Bogden, R. R. and MacLaren, J. N. 1950. The nutrition of Monosporium apiospermum. Mycologia 42: 2 3 3 241.
Mycoscience 36: 243-246, 1995
Short Communication
pH and temperature optima for growth and sporulation in some common fungi from city waste Sudhir Kumar M e h r a and Ajai K u m a r Jaitly Department of Plant Science, Faculty of Life Sciences, Rohilkhand University, Bareilly-243001, India Accepted for publication 28 April 1995
Relationships between the growth of certain fungi isolated from city waste and pH and temperature were examined by two methods. The tested isolates showed their maximum growth and sporulation at different pHs while temperature requirements were the same (28°C), except for Humicola grisea (43°C). Cladosporium herbarum and H. grisea showed double pH optima. The ranges of pH and temperature for sporulation were more limited than those for the vegetative growth. Although all the tested isolates showed wide tolerances to pH and temperature, the degree of tolerance varied with the isolates. A considerable change from the initial pH of the liquid medium was noted at the end of the experiment. Key Words
city waste; fungal growth; pH; temperature.
The environment in which organisms exist exerts a profound effect upon them and often dictates their behaviour. The microbial population of a habitat is influenced either singly or in combination by a number of biotic and abiotic factors. City waste provides a complex habitat supporting an abundant and extremely diverse population of microorganisms. There is considerable evidence that ecological conditions play an important role in microbial activity (Hudson, 1968; Rai et al., 1981; Jaitly, 1982, 1987), of which hydrogen ion concentration and temperature are very important in determining the growth, activity and composition of organisms (Alexander, 1961 ; Agnihotri, 1963, 1964; Mukerjee, 1966). Individual species may have lower or higher tolerance to these factors than the general fungal community (Jaitly, 1982, 1987). Hegarty and Curran (1985) observed that maximum wood decay by most of their isolates was in the temperature and pH ranges of 1 5 - 2 5 ° C and 5-8, respectively. Investigations have been conducted on pH and temperature optima of fungi from various habitats, but no report is available on these lines for the fungi from city waste. Furthermore, because of authors' interest in biodegradation and enzymatic hydrolysis of composting material, it was essential to find the optimal pH and temperature for the experimental fungal isolates. Effects of pH and temperature on the growth and sporulation of some common fungi from city waste were evaluated by a diameter measurement method because this has been shown to be of sufficient reliability (Brancato and Golding, 1953; Jaitly, 1982, 1987). Dry weight measurements were also taken (Jaitly, 1987). Czapek-Dox agar was prepared in citrate-phosphate buffer of different pH values ranging from 3.0 to 8.0 at in-
tervals of 0.5 unit. Sterilized medium was poured in sterilized Petri dishes (10 cm) and single point central inoculations were made with 2 mm discs of freshly grown mycelium of the test fungus. Three replicates of each isolate were grown at a constant temperature of 29__+ 1 °C. Plates were regularly examined at 24-h intervals for seven days. Two diameters were measured at right angles to each other as a measure of the average diameter for each colony. Isolates were examined for sporulation regularly. In the second method, 25 mt of Czapek-Dox medium of required pH values was poured in 100-ml Erlenmeyer flasks and autoclaved at 15 Ibs pressure for 15 min. The flasks were inoculated with 0.5 ml of homogenous spore suspension of the test fungus containing 107spores/ml. The flasks were incubated at 29___ 1°C. T w o replictes were taken for each. After seven days of incubation, the biomass obtained was filtered through preweighed Whatman filter paper No. 1 and dried for 24 h at 80°C. Final weight was taken to calculate the dry weight of fungal biomass. Test fungi were grown on Czapek-Dox medium at the optimum pH for their growth at different tempera, tures in the same way as stated for pH. The results of pH-growth relationship from colony diameter measurements were in agreement with those of the dry weight measurements. Tested isolates showed their maximum growth at different pHs (Table 1). After an initial lag phase, the increase in diameter was more or less linear for all the tested fungal isolates (Fig. 1 ) at pH near to their optima. The isolates grew optimally in the acidic to neutral pH range but also showed tolerance to alkaline pH. Slow growth was observed at pH 3.0 for all the isolates. Maximum growth for all the tested isolates
244
S.K. Mehra and A. K. Jaitly Table 1.
Effect of pH on growth of some fungi after seven days (dry weight measurement). Dry weight in mg*
Fungus 3.0
3.5
4.0
4.5
5.0
5.5
Altemaria hurnicola Oudemans
130
150
175
Aspergillus niger van Tieghem
120 +
180 +
240 4-F
Aspergillus terreus Thom
390 -iF
440 4+4-
520 II I I l
Cladosporiurn herbarum (Pers.) Link ex Gray
150
190
290
Curvularia lunata (Wakker) Boedijn
180 --
185 +
t I I t
230 lit 180 TII 535 Illl 330 4-4360 11t 445 I It 80 + 230 H-r-
360 IIlI 210 Itl 580 Iitti 450 ~ 340 I[I 490
290 I1t 230 II I 530 I I Ill 480 III 330 Ilt 380
pH**
Humicola grisea Traaen
320
330
300
30
45
75
190
200
210
--
Myrothecium roridum Tode ex Fr. Penicillium chrysogenurn Thorn
450
+
- -
4-+
44-
I[l
140 ~ 245 4~-
I
Iit
160 ++ 260 ill
6.0
6.5
240 200 4-~ -320 200 I I}t Iti 510 470 I t It III 580 560 t I t I t I ~11 I 325 210 -H-
-F+
360
380 4-F 90 + 380 I I[l
-H-
190 H-F 350 ++-t-t-
7.0
8.0
100 -180 ~+ 420 4-4390 ttt 260 .... 480 ÷-~ 50 .... 230 I II
110 100 35O 4-F 510 4-F 25O 35O 4-F 45 260 4-F
* Growth on Czapek-Dox medium. ** Citrate phosphate buffer. + , -- Degree of sporulation.
w a s obtained at pH b e t w e e n 4 . 0 and 6.5. Humicola grisea Traaen and Cladosporium herbarum (Pers.) Link ex Gray s h o w e d double pH o p t i m a , one in the acidic range (pH 5.0 and 6.0, respectively) and the other at neutral and alkaline pH i.e., p H 7 and p H 8 , respectively. M i c r o s c o p i c e x a m i n a t i o n of tested isolates revealed that the pH range for sporulation is n a r r o w e r than t h a t for the v e g e t a t i v e g r o w t h . Normally, excellent sporulation w a s o b t a i n e d b e t w e e n pH 4 . 0 and 5.5. All the t e s t e d isolates w e r e observed t o g r o w at pH 8 . 0 w i t h varying colony diameter, indicating t h a t t h e y have different abilities to tolerate e x t r e m e pH. A considerable change f r o m the initial pH of the liquid m e d i u m w a s noted (Table 3). T e m p e r a t u r e requirements w e r e same for all the test-
pH
80-
4.0 5.0
E
v
60-
6.0
t~ i,i FLIJ 7.0
40>Z
o._J o
8.0 209.0
I
f
[
I
48
96
144
192
TIME (]1) Fig.
Cardinal growth of Curvularia lunata on Czapek-Dox agar medium. In relation to pH. 1.
ed fungat isolates, e x c e p t for H. grisea (Table 2). A f t e r an initial lag phase, colony d i a m e t e r increased linearly w i t h t i m e for all the isolates (Fig. 2). Tested fungat isolates s h o w e d o p t i m u m g r o w t h at 2 8 ° C . H o w e v e r the mycetial g r o w t h of H. grisea w a s m a x i m u m at 4 3 ° C . M o s t of the fungi s h o w e d l o w e r g r o w t h at b e l o w 2 3 ° C and above 3 3 ° C . Sporulation w a s also m a x i m u m in the t e m p e r a t u r e range of 2 8 - 3 3 ° C . The t w o m e t h o d s used t o d e t e r m i n e g r o w t h in relation to pH or t e m p e r a t u r e w e r e s a t i s f a c t o r y and yielded similar results. A t e x t r e m e s of pH and t e m p e r a t u r e , the isolates g r e w only after a long incubation period. The existence of m i c r o o r g a n i s m s in the w a s t e materials is affected by a number of environmental factors such as pH, t e m perature, aeration, moisture, organic m a t t e r etc., of w h i c h pH and t e m p e r a t u r e are the m o s t i m p o r t a n t (Hegart y and Curran, 1 9 8 5 ; Pearce and Malajczuk, 1 9 9 0 ; B o w e r m a n and Goos, 1991). All the tested isolates s h o w e d their pH o p t i m a in the range of 4 . 0 t o 6.5. This m i g h t be due to the presence of high c o n c e n t r a t i o n s of organic m a t t e r w h i c h d e c o m p o s e s to f o r m different nitrogenous and carbonaceous substances, thus l o w e r ing t h e pH of w a s t e material. These studies are in agreem e n t w i t h those p e r f o r m e d by Jaitly (1987). A l e x a n d e r (1971) also stated t h a t the application of fertilizers containing nitrogenous substances leads to the f o r m a t i o n of acids by fungi on one hand and inhibition of the g r o w t h of bacteria and a c t i n o m y c e t e s on the other hand. Tested isolates s h o w e d some tolerance t o alkaline pH but their o p t i m u m g r o w t h lay in the l o w e r range. This is in a g r e e m e n t w i t h the results obtained by several w o r k e r s (Wolf et al., 1950; Brewer, 1960; Rai et at., 1970, Jaitly, 1987). In the present investigation, C. herbarum and H. grisea s h o w e d double pH o p t i m a . Biomodal curves have also been obtained by several o t h e r w o r k e r s (Jones and Irvine, 1972; Jaitly, 1987).
pH and temperature optima
Table 2.
245
Effect of temperature on growth of some fungi after seven days (dry weight measurement). Dry weight in mg*
Fungus Temperature °C
23
28
33
38
43
48
Altemaria hum/cola
200 44-
310 IIi
230 4-I-
220 +4-
190 +
140 --
Aspergillus niger
190 +4-
450 Jilt
330 ~
240 4-+4-
210 +4.
180 -FF
Aspergillus terreus
140 4-F
550 I ] 111
460 I I I I
370 II I
180 44-
070 4-
Cladosporium herbarum
240 -IF
420 I II I
380 I I I I
320 I] I
270 4-F
230
Curvularia tunata
250
540
490
430
320
180
I I F
I I ] q q
I I I I
Itl
4W
4-
Hum/cola grisea
150 --
250 ill
280 ~
320 444-}-
390 r Fli
310 lit
Myrothecium roridum
110 4-{-
270 Ill
190 II~
180 4-+
120 44
1O0 4-
Pen/c/Ilium chrysogenum
230 -FF
380 Itl
350 4-FF
310 +++
270 4-F
210 4-F
* + , --
Growth on Czapek-Dox medium at optimum pH. Degree of sporutation. Table 3.
Shift in the initial pH of basal medium. Final pH values after 7 days of growth
Initial pH
Altemaria humicola
Aspergillus niger
Aspergillus Cladosporium Curvularia terreus herbarum lunata
3.0 3.5
5,0 5.0
4.5 4.5
5.0 5.0
5.5 5.5
4.0 5,0
4.5 4,5
4.0 4.0
4.0 4.0
4.0 4,5
5,5 6.0
5.0 5.5
5.8 6.0
5.5 5.8
5.0 5.5
5.5 5.5
5.0 5.0
4.4 4.6
5.0
6,0
6.0
6.0
6.2
5.0
6,0
5.5
4.6
5,5
6.0
6.0
6.0
6.5
5,5
6.5
5.5
4.6
6.0 6,5
6.O 6.5
6.5 6.5
6.0 6.5
6.5 6.5
5.5 6.0
6,5 6.5
5.5 6.0
4.6 4,8
7,0
7.0
6.8
6,8
6.5
6.0
6.8
6.0
5.0
8.0
7.5
7.0
7.0
6,0
6.5
6.8
6.5
5.0
Fungi are k n o w n t o alter t h e pH o f m e d i u m in w h i c h t h e y g r e w , The p r e s e n t results also s h o w e d a c h a n g e in t h e initial pH of t h e basal m e d i u m . Generally t h e l o w e r pH v a l u e s b e c a m e higher and t h e h i g h e r b e c a m e l o w e r , w h i l e t h o s e in t h e v i c i n i t y o f pH 7 . 0 did n o t shift sign i f i c a n t l y t o either side (Table 3). In t h e p r e s e n t s t u d y t h e species g r e w a f t e r a long inc u b a t i o n period at t h e e x t r e m e t e m p e r a t u r e s . M a n y w o r kers (Tansey, 1 9 7 2 ; J a i t l y , 1 9 8 2 ) s u g g e s t e d t h a t a long lag phase can be e x p e c t e d at i n c u b a t i o n t e m p e r a t u r e s differing f r o m t h a t of t h e i n o c u l u m . Evans ( 1 9 7 1 ) c o n cluded t h a t a s h o r t i n c u b a t i o n period m a y n o t reflect t h e t r u e cardinal t e m p e r a t u r e s for g r o w t h . During t h e p r e s e n t i n v e s t i g a t i o n , all t h e t e s t e d isolates s h o w e d their m a x i m u m g r o w t h at 2 8 ° C , e x c e p t H, grisea, w h i c h s h o w e d o p t i m u m g r o w t h at 4 3 ° C , indicating t h a t t h e species is o f t h e r m o t o l e r a n t nature. The ranges o f pH and t e m p e r a t u r e for s p o r u l a t i o n w e r e l o w e r t h a n t h o s e for vegetative growth (Jatilty, 1982, 1987).
Hum/cola Myrothecium Pen/c/Ilium grisea roridum chrysogenum
80-
TEMP. ( °C ) 28
60-~ r~ uJ FL~J
<
33 38 43 23
40-
>-
z o
_J
0 c.)
48
20-
I
I
I
i
48
96
144
192
T I M E (h) Fig.
Cardinal growth of Curvularia lunata on Czapek-Dox agar medium. In relation to temperature. 2.
246
S.K. Mehra and A. K, Jaitly
Literature cited Agnihotri, V, P. 1963, Studies on Colletotrichum capsci III. The effect of pH on growth and amino acid composition, Mycopathol. Mycol. Appl, 24: 305-314. Agnihotri, V. P. 1964. Studies of Aspergilli XVI. Effect of pH, temperature and carbon and nitrogen interaction. Mycopathol, Mycol, Appl. 25: 3 0 5 - 3 1 4 . Alexander, M, 1961. "Introduction to soil microbiology," John Wiley, New York. 472 p. Alexander, M, 1971. Biochemical ecology of microorganisms. Ann. Rev. Microbiol. 25: 3 6 1 - 3 9 1 . Bowerman, L. and Goos, R. D. 1991. Physiological studies of t w o fungi isolated from Nymphaea odorata. Mycologia 83: 624-632. Brancato, F, P. and Golding, N. S. 1953, The diameter of the mold colony as a reliable measure of growth. Mycologia 45: 8 4 8 - 8 6 4 . Brewer, D. 1960. Studies on AscochytaplsiLib. Can. J, Bot, 38:705-717. Deploey, J, 1992. Some factors affecting germination of Rhizomucor miehei sporangiospores. Mycologia 84: 7 7 81. Evans, H. C, 1971. Thermophilous fungi of coal spoil tips. tl. Occurrence, distribution and temperature relationships. Trans. Br. Mycot. Soc. 57: 2 2 5 - 2 6 6 . Hegarty, B. M. and Curran, P. M. T, 1985, The biodeterioration of beech (Fagus sylvatica) by marine and non-marine fungi in response to temperature, pH, light and dark, Int. Biodeterioration 21 : 11 - 18, Hudson, H. J. 1968. The ecology of fungi on the plant remains
above the soil. New Phytol. 67: 837-874. Jaitly, A. K, 1982. Ecological studies of thermophitic fungi native to mangrove swamps. I. Temperature-growth relationships. Trans. Mycol. Soc. Japan 23: 6 5 - 7 1 , Jaitiy, A. K. 1987. pH optima of the fungi isolated from mangrove soils in India. Trans. Mycol. Soc. Japan 28: 157t63. Jones, E. B. G. and Irvine, J. 1972. The role of marine fungi in the biodeterioration of materials. In: "Biodeterioration of materials," (ed. by Waiters, A. H. and Hueck-Vender Plas, E, H.), pp. 4 2 2 - 4 3 1 . Applied Science, London. Mukerjee, K.G. 1966. Studies on the effect of hydrogen ion concentration on growth and sporulation of certain soil fungi. Mycopathol. Mycol. Appl. 28: 3 1 2 - 3 1 6 . Pearce, M, H. and Malajczuk, N. 1990. Factors effecting growth of Armillaria luteobubalina rhizomorphs in soil. Mycol, Res, 94: 3 8 - 4 8 . Rai, J.N.,Garg, K. L a n d J a i t l y , A.K. 1981. Saprophytic fungi from woods in mangrove swamps and their wood decaying capability. Trans. Mycol. Soc. Japan 22: 6 5 - 7 4 . Rai, J. N., Sharma, B. B. and Agarwal, S.C. 1970, Increased pH tolerance of some Aspergilli isolated from 'Usar' (alkaline) soils--A possible indication of ecological specialization. Sydowia 24: 3 3 6 - 3 4 3 . Tansey, M, R. 1972. Effect of temperature on growth rate and development of the thermophilic fungus--Chaetornium thermophile. Mycologia 64: 1290-1299. Wolf, F. T., Bogden, R. R. and MacLaren, J. N. 1950. The nutrition of Monosporium apiospermum. Mycologia 42: 2 3 3 241.