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Distribution patterns of protein mineralizing and ammonifying bacterial populations in fish-farming ponds under different management systems
Aquaculture, 44 (1985) 57-65 Elsevier Science Publishers B.V., Amsterdam - Printed in The Netherlands
57
DISTRIBUTION PATTERNS OF PROTEIN MINERALIZING AND AMMONIFYING BACTERIAL POPULATIONS IN FISH-FARMING PONDS UNDER DIFFERENT MANAGEMENT SYSTEMS
B.B. JANA and S.K. ROY Deportment (Accepted
of Zoology,
University of Kalyani, Kalyani 741235,
West Bengal (India)
28 August 1984)
ABSTRACT Jana, B.B. and Roy, S.K., 1985. Distribution patterns of protein mineralizing and ammonifying bacterial populations in fish-farming ponds under different management systems. Aquaculture, 44: 57-65. Enumeration of protein mineralizing and ammonifying bacterial populations in water and sediment samples of six fish-farming ponds (polyculture, monoculture and traditional systems) over a period of two and a half years revealed significant differences between the culture systems with maximal and lowest counts in the monoculture and traditional systems respectively. This distribution pattern was related to the degree of organic loading in the system. Both groups of bacteria were most numerous during winter but were greatly reduced in number during the summer. The factors responsible for their spatial and seasonal distributions are discussed.
INTRODUCTION
Recent research on intensive fish culture has shown that ammonia plays a major role as a limiting factor for fish growth. Both autochthonous and allochthonous organic matter, and excretion of ammonium and amino acids by zooplankton (Brezonik, 1972), as weli as direct autolysis after cell death (Krause, 1964), are chief sources of ammonia in fish ponds. A number of micro-organisms (Escherichia coli, Proteus vulgaris, Bacillus subtilis, Aeriobacter cloacai, Pseudomonas sp. and Flavobacterium sp.) are capable of hydrolysing proteins into simpler compounds such as peptides, urea, amino acids etc. which, in turn, are metabolized by ammonifying bacteria to liberate ammonia or ammonium sulphate (Morris and Koffron, 1967; Grant and Patel, 1969; Little et al., 1969; Kruger, 1978). Even some bacteria capable of carrying out denitrification (Achromobacter sp. and Micrococcus sp.) can actively take part in proteolysis (Alexander, 1971). Despite a number of studies on proteolytic bacteria and ammonification rates in a variety of temperate waters (Rheinheimer, 1959, 1965; Jones et al., 1982), such information is extremely meagre for tropical waters
0044-8486/85/$03.30
o 1985 Elsevier Science Publishers B.V.
58
especially in intensive fish-farming ponds where the flux of organic nitrogen is often high due to the application of heavy doses of organic manures conducive to the massive development of both protein mineralizing and ammonifying bacteria. In the present investigation, the population density of these two groups of bacteria was determined in relation to the farming management system and water quality. MATERIALS
AND METHODS
Six fish ponds being farmed under polyculture, monoculture and traditional culture systems were used in this study. These ponds were distributed in two farms, Kalyani and Naihati, which are separated by a distance of 12 km. The water area and mean depth of the fish ponds ranged from 0.04 to 6.4 ha and 1.0 to 3.6 m respectively. The polyculture fish ponds were stocked at a density of 6000 ha-‘, with three species of Indian major carps (Lubeo rohitu, C&la cutlu and Cirrhinus mrigala) and three exotic carps (Hypoph thalmich thys molitrix, Ctenopharyngodon idella and Cyprinus carpio) in a ratio of 6:2:3:4:2:3
l POLYCULTURE-1 &MONOCULTURE-l .TRADITIONAL-1
0 POLYCULTURE-2 nMONOCULTURE -2 q TRADITIONAL-2
KALYANI
‘;_ 8E m J3 0 g” z5 2 0 4-
3 m r
a
32lI, I I I I III JSNJMMJSNJMMJSN 1978
I I
I
I
I
I
I
I
I
1979
I,
I
I
I,
1,
1,
1
1980
Fig. 1. Seasonal changes of protein mineralizing bacteria occurring in water samples of six fish-farming ponds.
59
respectively. Cirrhinus mrigala were introduced into the monoculture ponds maintaining the same stocking rate. The traditional fish ponds had been stocked largely with three species of Indian major carps. All the fish ponds, with the exception of those managed in the traditional way, were fertilized at monthly intervals with the application of 50 kg ammonium sulphate or 10.5 kg N ha-’ and 50 kg single superphosphate or 8 kg P ha-‘, and manured with 2000 kg ha-’ cattle dung (wet). Samples of water and surface sediments were collected from these six ponds twice a month at regular intervals for nearly two and a half years from July 1978 to October 1980, Enumeration of the protein mineralizing and ammonifying bacteria in the present study is based on the idea that the first decomposes protein to simple compounds such as peptides, urea, amino acids etc. while the latter convert these simple compounds to ammonia. Milk protein and peptone were used as substrates for the growth of protein mineralizing and ammonifying bacteria respectively. The numbers of the two groups of bacteria are, thus, not additive. The protein mineralizing bacteria (PMB) were grown on milk agar medium as described by APHA (1976). A series of dilutions of water samples and
ll-
. POLYCULTURE1 . MONOCULTURE-l , TRADlTlONAL-1
1978
KALYANI
o POLYCULTURE-2 n o TRADITIONAL-2
1979
NAIHATI
1980
Fig. 2. Seasonal changes of protein mineralizing bacteria occurring in sediment samples of six fish-farming ponds.
60
soil extract (10-1-10-6) were prepared; each dilution was then plated in quadruplicate. The abundance of ammonifying bacteria (AB) was determined by the MPN (most probable number) technique using the peptonewater medium of Meiklejohn (1965) at an incubation temperature of 35°C for 3 days. The data were statistically evaluated using analysis of variance (split plot model) to determine the effect of farm site, cultural practices and season, and their interactions, if any, on the distribution pattern of the monthly mean bacterial density. In this model, individual ponds were considered as the whole plot treatment and season as the sub-plot. The locality of the farm was considered as a replicate. The physico-chemical parameters of the water and bottom sediments were monitored at a fixed hour of the day (10 a.m.) at bimonthy intervals during the period of the study. RESULTS
Spatial distribution It can be seen from Figs. 1 to 4 that the density of PMB and AB in the Naihati ponds was significantly higher than in those at Kalyani. The highest . POLYCULTURE-1
1.9
0 POLYCULTURE-2 A MONOCULTURE-2 q TRADITIONAL-2
1.8
NAIMATI
1.7
lllllIllllI11111Illlllllllr11
JSNJMMJSNJMMJS 1979
Fig. 3. Seasonal changes of ammonifying farming ponds.
1979
bacterial populations
7980
in waters of six fish-
61 .PoLYCULTURE-1 AMONOCULTURE-1 n TRADITIONAL-1
oPOLYCULTURE-2 nMONOCULTURE-2 q TRADITIONAL-2
a,22 E 2 2.0 ”
18
L 16 2 : 14
m p 12 ,x 10 : 08 E E 06 a 04
JSNJMMJSNJMMJS 1978
Fig. 4. Seasonal farming ponds.
changes
1979
of ammonifying
bacterial
populations
1980
in sediments
of six fish-
and lowest counts of PMB (F2.? > 117.2, P Q 0.01) as well as AB (Fz,! > 40.59, P < 0.05) were observed in the monoculture and traditional systems respectively. The density observed in the polyculture system was intermediate between those of the other two systems. Comparing the population size of PMB and AB in the surface sediments and overlying water, significantly higher populations can be observed in the former. This phenomenon might be due to the effect of a greater accumulation of important nutrients in the sediments serving as a principal site of sedimentation and decomposition of organic matter. Kuznetsov (1968), Stewart et al. (1977), Sugiyama and Kawai (1979), and Shilo and Rimon (1982) also confirmed similar results in other waters. Seasonal distribution The PMB (F27,54 > 59.91, P < 0.001) and AB (F,,,,, > 8.15, P < 0.05) populations were extremely variable in number at different times of the year (Figs. l-4). They were most numerous in winter but greatly reduced in summer.
Surface water
10.1 0.09 1.0 1.8 1.7 30.1 0.21
11.1 0.15 0.97 1.67 1.66 30.7 0.20
7.3 15.2 2.078 2.9 35.8 0.027 0.058
Sediments PH PD.-P (mg I-‘) Organic C (X) CaCO, (%) C/N Available N (96) Total N (%)
*KaIyani. **Naihati.
10.7
11.2
1.4 17.5 2.2 3.2 31.1 0.037 0.071
0.20
10.0 0.09 1.41 2.64 2.02 28.0 0.20
9.6 0.1 1.5 2.8 2.1 28.2
10.5
7.1
6.8
8.7
8.1 10.4
106.0
107.0
96.0
95.0
9.5 91.0
10.2 90.0
5.5 94.0
5.1 98.0
8.3
_
28.6 8.3
8.1
28.3 8.1
-
Bottom water
Monoculture-l*
Surface water
Bottom water
PoIyculture-1*
6.7 12.4 1.3 4.9 35.5 0.0149 0.039
0.16
10.4 0.06 0.97 0.80 1.40 24.8
11.8
5.9
63.0
5.5 62.0
28.3 8.3
Surface water
0.44
0.16
7.3 12.7 1.6 4.5 26.9 0.028 0.061
8.3 0.15 1.11 1.56 1.57 26.1
8.1
8.1
182.0
10.6 167.0
28.8 8.2
Surface water
8.3 0.08 1.6 2.50 1.95 27.3
7.9 0.08 1.3 1.7 1.6 0.08
7.1 17.2 1.9 5.6 27.6 0.033 0.069
0.69
6.9
1.7
0.44
8.2
179.0
14.5 184.0
28.6 8.1
Surface water
0.70
7.6 0.09 1.8 2.7 2.0 0.09
6.6
8.5
177.0
15.9 182.0
8.1
_
Bottom water
Monoculture-2**
8.4
179.0
10.7 164.0
8.3
-
Bottom water
Polyculture--2**
7.4 6.9 0.9 2.9 39.7 0.011 0.248
0.65
5.5 0.05 0.61 0.76 1.19 25.3
7.3
4.6
200.0
13.6 211.0
28.7 8.2
Surface water
0.65
5.2 0.06 0.8 0.8 1.2 0.65
7.1
4.9
195.0
15.8 207.0
8.2
_
Bottom water
Traditional-2**
sediments in various fish-farming ponds (each mean value
9.9 0.07 0.6 0.8 1.4 22.3
11.5
6.3
62.0
6.3 59.0
8.3
_
Bottom water
Traditional--l*
parameters or water and bottom
Water Temperature (“C) PH Free CO, (mg IV’) HCO; (mg 1.‘) Total hardness (mg I-‘) Dissolved organic carbon (mg 1-l) Dissolved organic matter (mg I-‘) D&solved 0, (mg I-‘) PO,-P (mg IV’) NH,-N (mg 1-l) NO,-N (mg I-‘) NO,-N (mg I-‘) so, (mg IV’) Conductivity (rmho cm-‘)
Parameter
Monthly mean vaIues of physlco-chemical represents 28 months’ data)
TABLE I
%
63
Physicochemical
analysis
Physicochemical analysis of the water and bottom sediments of the ponds were made at bimonthly intervals for the 28-month period of this study; the overall monthly mean values are given in Table I. The seasonal variation of all of these physicochemical parameters will be reported elsewhere. DISCUSSION
Since the density of AB was much higher than that of the PMB in these fish ponds regardless of the culture system, it implies that selection of ammonifiers was perhaps favoured by a wide variety of organic nitrogenrich substrates originating from the cattle dung and fish meal applied. Some ammonifying micro-organisms are reported to be substrate-specific, utilizing only peptone and not simple amino acids, or using urea but not uric acid; in contrast, other species are known to use a wide variety of organic nitrogen sources (Kormondy, 1978; Sepers, 1981). The relatively larger populations of PMB and AB in all Naihati farm ponds were largely responsible for regeneration of greater amounts of ammonia in the pond water from Naihati than in that from Kalyani. It is clearly evident from this study that the management system of a fish farm plays a significant role in the distribution pattern of PMB and AB. Rearing of a single fish species in a monoculture system perhaps caused an ecological imbalance with surplus nitrogen which was conducive to the massive development of microbial populations (Jana and Roy, 1983; Jana and Patel, 1984) including the PMB and AB. Since both PMB and AB act upon the indiffusible protein fraction of organic matter and initiate the deamination process, their relationships with different species of nitrogen were not unexpected. Stewart et al. (1977) observed a close relationship between the number of ammonifying bacteria in sediments and the levels of interstitial NH: present. Jana and Barat (1984) obtained results which were similar to those found in this study; they observed a good correlation between the amount of dissolved oxygen in the water and the density of ammonifiers in the fish ponds. Furthermore, the seasonal changes of ammonification rates have been found to be directly dependent upon the population dynamics of PMB and AB in these ponds (Roy, 1984). Although the optimal temperature for ammonification has been reported to be 30-35°C (Rheinheimer, 1980), the winter temperature of around 20°C in the present investigation was perhaps conducive to the massive development of PMB and AB in these waters having a high flux of organic nitrogenous substances. These findings corroborate the observations of Rheinheimer (1959, 1965) who reported that the most intense ammonification occurred with a greater abundance of proteolytic bacteria in the winter
64
compared with the summer. However, this relationship was restricted to sewage-loaded waters and did not occur in clear rivers (Rheinheimer, 1980). Niewolak (1965), on the other hand, reported maximal values of ammonification in summer and early autumn with a decline in winter. ACKNOWLEDGEMENTS
This research was supported by the research grant 11 (8)-76-ASR (I) from the Indian Council of Agricultural Research, New Delhi (to BBJ). We are thankful to the Department of State Fisheries, Govt. of West Bengal, for their cooperation which enabled us to collect data from Kalyani fish farm.
REFERENCES Alexander, M., 1971. Microbial Ecology. John Wiley and Sons Inc., New York, 511 PP. American Public Health Association, 1976. Standard Methods for the Examination of Water and Waste Water. Fourteenth edition. A.P.H.A., New York, 1193 pp. Brezonik, P.L., 1972. Nitrogen: Sources and transformation in natural waters. In: H.E. Allen and J.R. Kramer (Editors), Nutrients in Natural Waters. Wiley-Interscience Publications, New York, pp. l-50. Grant, D.J.W. and Patel, J.C., 1969. The non-oxidative decarboxylation of p-hydroxybenzoic acid, genetisic acid, protocatecuic acid and gallic acid by Klebsiella aerogenes (Aembacter aemgenes). Antonie van Leeuwenhoek. J. Microbial. Serol., 35: 325-343. Jana, B.B. and Barat, S., 1984. Ammonification as affected by oxygen level of water. Limnologica, 16: 67-70. Jana, B.B. and Patel, G.N., 1984. Measurements of some groups of microbialpopulations with special reference to denitrification in water and sediments of fish-farming ponds under mono- and polyculture systems. Int. Rev. Gesamten Hydrobiol., 69: 231-240. Jana, B.B. and Roy, SK., 1983. Estimates of microbial population involved in N cycle and their activity in water and sediments of fish ponds under mono- and polyculture systems. Int. Rev. Gesamten Hydrobiol., 68: 581-590. Jones, J.G., Simon, B.M. and Horsley, R.W., 1982. Microbiological sources of ammonia in freshwater lake sediments. J. Gen. Microbial., 128: 2823-2831. Kormondy, E.D., 1978. Concepts of Ecology. Prentice Hall of India Private Limited, New Delhi, 238 pp. Krause, H.R., 1964. Zur Chemie und Biochemie der Zersetzung von susswasser Organismen unter besonderer Berucksichtigung des Abbaues der organischen Phosphorkomponenten. Verh. Int. Verh. Limnol., 15: 549-561. Kruger, D., 1978. The influence of silver carp (Hypophthalmichthys molitrix) on eutrophication of the environment of carp ponds. 2. Active microflora changes in nitrogen conversion. Rocz. Nauk. Roln. Ser. H Rybactwo, 99: 33-54. Kuznetsov, S.I., 1968. Recent Studies on the Role of Microorganisms in the Cycling of Substances in Lakes. Verlag Wissenschaften, 301 pp. Little, J.E., Robert, E.S. and Gerald, R.C., 1969. Measurement of proteolysis in natural waters. Appl. Environ. Microbial., 37: 900-908. Meiklejohn, J., 1965. Microbiological studies on large termite mounds. Rhod. Zambia Malawi J. Agric. Res., 3: 67-79. Morris, D.R. and Koffron, K.L., 1967. Urea production and putrescine biosynthesis by Escherichia coli. J. Bacterial., 94: 1516-1519.
65 Niewolak, S., 1965. Badanie intensywnosciniektorych przemian azotu w jeziorach ilawskich w latach 1962-1963. Zesz. Nauk Wyszsz. Szk. Roln. Olsztynie, 20: 425, 6984. Rheinheimer, G., 1959. Mikrobiologische Untersuchungen iiber den Stickstoffhaushalt der Elbe. Arch. Mikrobiol., 34: 358-373. Rheinheimer, G., 1965. Mikrobiologische Untersuchungen in der Elbe zwischen Schnackenburg und Cuxhaven. Arch. Hydrobiol., 29 (Suppl.): 181-251. Rheinheimer, G., 1980. Aquatic Microbiology. Second edition. John Wiley and Sons, New York, 235 pp. Roy, S.K., 1984. Spatial and seasonal distributions of microbial populations involved in N cycle and their activity in fish farming ponds under traditional, monoculture and polyculture systems. Ph.D. Thesis, Kalyani University, Kalyani, West Bengal, India, 294 pp. 1981. Diversity of ammonifying bacteria. Hydrobiologia, 83: 153Sepers, A.B.J., 160. Shilo, M. and Rimon, A., 1982. Factors which affect the intensification of fish breeding in Israel. 2. Ammonia transformation in intensive fish ponds. Bamidgeh, 34: 101-114. Stewart, W.D.P., Sinada, F., Christofi, N. and Daft, M.J., 1977. Primary production and microbial activity in Scottish fresh water habitats. In: F.A. Skinner and J.M. Shewan (Editors), Aquatic Microbiology. Academic Press, London, pp. 31-54. Sugiyama, M. and Kawai, A., 1979. Microbiological studies on the nitrogen cycle in aquatic environment. VI. Metabolic rate of ammonium nitrogen in a gold fish culturing pond. Bull. Jpn. Sot. Sci. Fish., 45: 785-789.
57
DISTRIBUTION PATTERNS OF PROTEIN MINERALIZING AND AMMONIFYING BACTERIAL POPULATIONS IN FISH-FARMING PONDS UNDER DIFFERENT MANAGEMENT SYSTEMS
B.B. JANA and S.K. ROY Deportment (Accepted
of Zoology,
University of Kalyani, Kalyani 741235,
West Bengal (India)
28 August 1984)
ABSTRACT Jana, B.B. and Roy, S.K., 1985. Distribution patterns of protein mineralizing and ammonifying bacterial populations in fish-farming ponds under different management systems. Aquaculture, 44: 57-65. Enumeration of protein mineralizing and ammonifying bacterial populations in water and sediment samples of six fish-farming ponds (polyculture, monoculture and traditional systems) over a period of two and a half years revealed significant differences between the culture systems with maximal and lowest counts in the monoculture and traditional systems respectively. This distribution pattern was related to the degree of organic loading in the system. Both groups of bacteria were most numerous during winter but were greatly reduced in number during the summer. The factors responsible for their spatial and seasonal distributions are discussed.
INTRODUCTION
Recent research on intensive fish culture has shown that ammonia plays a major role as a limiting factor for fish growth. Both autochthonous and allochthonous organic matter, and excretion of ammonium and amino acids by zooplankton (Brezonik, 1972), as weli as direct autolysis after cell death (Krause, 1964), are chief sources of ammonia in fish ponds. A number of micro-organisms (Escherichia coli, Proteus vulgaris, Bacillus subtilis, Aeriobacter cloacai, Pseudomonas sp. and Flavobacterium sp.) are capable of hydrolysing proteins into simpler compounds such as peptides, urea, amino acids etc. which, in turn, are metabolized by ammonifying bacteria to liberate ammonia or ammonium sulphate (Morris and Koffron, 1967; Grant and Patel, 1969; Little et al., 1969; Kruger, 1978). Even some bacteria capable of carrying out denitrification (Achromobacter sp. and Micrococcus sp.) can actively take part in proteolysis (Alexander, 1971). Despite a number of studies on proteolytic bacteria and ammonification rates in a variety of temperate waters (Rheinheimer, 1959, 1965; Jones et al., 1982), such information is extremely meagre for tropical waters
0044-8486/85/$03.30
o 1985 Elsevier Science Publishers B.V.
58
especially in intensive fish-farming ponds where the flux of organic nitrogen is often high due to the application of heavy doses of organic manures conducive to the massive development of both protein mineralizing and ammonifying bacteria. In the present investigation, the population density of these two groups of bacteria was determined in relation to the farming management system and water quality. MATERIALS
AND METHODS
Six fish ponds being farmed under polyculture, monoculture and traditional culture systems were used in this study. These ponds were distributed in two farms, Kalyani and Naihati, which are separated by a distance of 12 km. The water area and mean depth of the fish ponds ranged from 0.04 to 6.4 ha and 1.0 to 3.6 m respectively. The polyculture fish ponds were stocked at a density of 6000 ha-‘, with three species of Indian major carps (Lubeo rohitu, C&la cutlu and Cirrhinus mrigala) and three exotic carps (Hypoph thalmich thys molitrix, Ctenopharyngodon idella and Cyprinus carpio) in a ratio of 6:2:3:4:2:3
l POLYCULTURE-1 &MONOCULTURE-l .TRADITIONAL-1
0 POLYCULTURE-2 nMONOCULTURE -2 q TRADITIONAL-2
KALYANI
‘;_ 8E m J3 0 g” z5 2 0 4-
3 m r
a
32lI, I I I I III JSNJMMJSNJMMJSN 1978
I I
I
I
I
I
I
I
I
1979
I,
I
I
I,
1,
1,
1
1980
Fig. 1. Seasonal changes of protein mineralizing bacteria occurring in water samples of six fish-farming ponds.
59
respectively. Cirrhinus mrigala were introduced into the monoculture ponds maintaining the same stocking rate. The traditional fish ponds had been stocked largely with three species of Indian major carps. All the fish ponds, with the exception of those managed in the traditional way, were fertilized at monthly intervals with the application of 50 kg ammonium sulphate or 10.5 kg N ha-’ and 50 kg single superphosphate or 8 kg P ha-‘, and manured with 2000 kg ha-’ cattle dung (wet). Samples of water and surface sediments were collected from these six ponds twice a month at regular intervals for nearly two and a half years from July 1978 to October 1980, Enumeration of the protein mineralizing and ammonifying bacteria in the present study is based on the idea that the first decomposes protein to simple compounds such as peptides, urea, amino acids etc. while the latter convert these simple compounds to ammonia. Milk protein and peptone were used as substrates for the growth of protein mineralizing and ammonifying bacteria respectively. The numbers of the two groups of bacteria are, thus, not additive. The protein mineralizing bacteria (PMB) were grown on milk agar medium as described by APHA (1976). A series of dilutions of water samples and
ll-
. POLYCULTURE1 . MONOCULTURE-l , TRADlTlONAL-1
1978
KALYANI
o POLYCULTURE-2 n o TRADITIONAL-2
1979
NAIHATI
1980
Fig. 2. Seasonal changes of protein mineralizing bacteria occurring in sediment samples of six fish-farming ponds.
60
soil extract (10-1-10-6) were prepared; each dilution was then plated in quadruplicate. The abundance of ammonifying bacteria (AB) was determined by the MPN (most probable number) technique using the peptonewater medium of Meiklejohn (1965) at an incubation temperature of 35°C for 3 days. The data were statistically evaluated using analysis of variance (split plot model) to determine the effect of farm site, cultural practices and season, and their interactions, if any, on the distribution pattern of the monthly mean bacterial density. In this model, individual ponds were considered as the whole plot treatment and season as the sub-plot. The locality of the farm was considered as a replicate. The physico-chemical parameters of the water and bottom sediments were monitored at a fixed hour of the day (10 a.m.) at bimonthy intervals during the period of the study. RESULTS
Spatial distribution It can be seen from Figs. 1 to 4 that the density of PMB and AB in the Naihati ponds was significantly higher than in those at Kalyani. The highest . POLYCULTURE-1
1.9
0 POLYCULTURE-2 A MONOCULTURE-2 q TRADITIONAL-2
1.8
NAIMATI
1.7
lllllIllllI11111Illlllllllr11
JSNJMMJSNJMMJS 1979
Fig. 3. Seasonal changes of ammonifying farming ponds.
1979
bacterial populations
7980
in waters of six fish-
61 .PoLYCULTURE-1 AMONOCULTURE-1 n TRADITIONAL-1
oPOLYCULTURE-2 nMONOCULTURE-2 q TRADITIONAL-2
a,22 E 2 2.0 ”
18
L 16 2 : 14
m p 12 ,x 10 : 08 E E 06 a 04
JSNJMMJSNJMMJS 1978
Fig. 4. Seasonal farming ponds.
changes
1979
of ammonifying
bacterial
populations
1980
in sediments
of six fish-
and lowest counts of PMB (F2.? > 117.2, P Q 0.01) as well as AB (Fz,! > 40.59, P < 0.05) were observed in the monoculture and traditional systems respectively. The density observed in the polyculture system was intermediate between those of the other two systems. Comparing the population size of PMB and AB in the surface sediments and overlying water, significantly higher populations can be observed in the former. This phenomenon might be due to the effect of a greater accumulation of important nutrients in the sediments serving as a principal site of sedimentation and decomposition of organic matter. Kuznetsov (1968), Stewart et al. (1977), Sugiyama and Kawai (1979), and Shilo and Rimon (1982) also confirmed similar results in other waters. Seasonal distribution The PMB (F27,54 > 59.91, P < 0.001) and AB (F,,,,, > 8.15, P < 0.05) populations were extremely variable in number at different times of the year (Figs. l-4). They were most numerous in winter but greatly reduced in summer.
Surface water
10.1 0.09 1.0 1.8 1.7 30.1 0.21
11.1 0.15 0.97 1.67 1.66 30.7 0.20
7.3 15.2 2.078 2.9 35.8 0.027 0.058
Sediments PH PD.-P (mg I-‘) Organic C (X) CaCO, (%) C/N Available N (96) Total N (%)
*KaIyani. **Naihati.
10.7
11.2
1.4 17.5 2.2 3.2 31.1 0.037 0.071
0.20
10.0 0.09 1.41 2.64 2.02 28.0 0.20
9.6 0.1 1.5 2.8 2.1 28.2
10.5
7.1
6.8
8.7
8.1 10.4
106.0
107.0
96.0
95.0
9.5 91.0
10.2 90.0
5.5 94.0
5.1 98.0
8.3
_
28.6 8.3
8.1
28.3 8.1
-
Bottom water
Monoculture-l*
Surface water
Bottom water
PoIyculture-1*
6.7 12.4 1.3 4.9 35.5 0.0149 0.039
0.16
10.4 0.06 0.97 0.80 1.40 24.8
11.8
5.9
63.0
5.5 62.0
28.3 8.3
Surface water
0.44
0.16
7.3 12.7 1.6 4.5 26.9 0.028 0.061
8.3 0.15 1.11 1.56 1.57 26.1
8.1
8.1
182.0
10.6 167.0
28.8 8.2
Surface water
8.3 0.08 1.6 2.50 1.95 27.3
7.9 0.08 1.3 1.7 1.6 0.08
7.1 17.2 1.9 5.6 27.6 0.033 0.069
0.69
6.9
1.7
0.44
8.2
179.0
14.5 184.0
28.6 8.1
Surface water
0.70
7.6 0.09 1.8 2.7 2.0 0.09
6.6
8.5
177.0
15.9 182.0
8.1
_
Bottom water
Monoculture-2**
8.4
179.0
10.7 164.0
8.3
-
Bottom water
Polyculture--2**
7.4 6.9 0.9 2.9 39.7 0.011 0.248
0.65
5.5 0.05 0.61 0.76 1.19 25.3
7.3
4.6
200.0
13.6 211.0
28.7 8.2
Surface water
0.65
5.2 0.06 0.8 0.8 1.2 0.65
7.1
4.9
195.0
15.8 207.0
8.2
_
Bottom water
Traditional-2**
sediments in various fish-farming ponds (each mean value
9.9 0.07 0.6 0.8 1.4 22.3
11.5
6.3
62.0
6.3 59.0
8.3
_
Bottom water
Traditional--l*
parameters or water and bottom
Water Temperature (“C) PH Free CO, (mg IV’) HCO; (mg 1.‘) Total hardness (mg I-‘) Dissolved organic carbon (mg 1-l) Dissolved organic matter (mg I-‘) D&solved 0, (mg I-‘) PO,-P (mg IV’) NH,-N (mg 1-l) NO,-N (mg I-‘) NO,-N (mg I-‘) so, (mg IV’) Conductivity (rmho cm-‘)
Parameter
Monthly mean vaIues of physlco-chemical represents 28 months’ data)
TABLE I
%
63
Physicochemical
analysis
Physicochemical analysis of the water and bottom sediments of the ponds were made at bimonthly intervals for the 28-month period of this study; the overall monthly mean values are given in Table I. The seasonal variation of all of these physicochemical parameters will be reported elsewhere. DISCUSSION
Since the density of AB was much higher than that of the PMB in these fish ponds regardless of the culture system, it implies that selection of ammonifiers was perhaps favoured by a wide variety of organic nitrogenrich substrates originating from the cattle dung and fish meal applied. Some ammonifying micro-organisms are reported to be substrate-specific, utilizing only peptone and not simple amino acids, or using urea but not uric acid; in contrast, other species are known to use a wide variety of organic nitrogen sources (Kormondy, 1978; Sepers, 1981). The relatively larger populations of PMB and AB in all Naihati farm ponds were largely responsible for regeneration of greater amounts of ammonia in the pond water from Naihati than in that from Kalyani. It is clearly evident from this study that the management system of a fish farm plays a significant role in the distribution pattern of PMB and AB. Rearing of a single fish species in a monoculture system perhaps caused an ecological imbalance with surplus nitrogen which was conducive to the massive development of microbial populations (Jana and Roy, 1983; Jana and Patel, 1984) including the PMB and AB. Since both PMB and AB act upon the indiffusible protein fraction of organic matter and initiate the deamination process, their relationships with different species of nitrogen were not unexpected. Stewart et al. (1977) observed a close relationship between the number of ammonifying bacteria in sediments and the levels of interstitial NH: present. Jana and Barat (1984) obtained results which were similar to those found in this study; they observed a good correlation between the amount of dissolved oxygen in the water and the density of ammonifiers in the fish ponds. Furthermore, the seasonal changes of ammonification rates have been found to be directly dependent upon the population dynamics of PMB and AB in these ponds (Roy, 1984). Although the optimal temperature for ammonification has been reported to be 30-35°C (Rheinheimer, 1980), the winter temperature of around 20°C in the present investigation was perhaps conducive to the massive development of PMB and AB in these waters having a high flux of organic nitrogenous substances. These findings corroborate the observations of Rheinheimer (1959, 1965) who reported that the most intense ammonification occurred with a greater abundance of proteolytic bacteria in the winter
64
compared with the summer. However, this relationship was restricted to sewage-loaded waters and did not occur in clear rivers (Rheinheimer, 1980). Niewolak (1965), on the other hand, reported maximal values of ammonification in summer and early autumn with a decline in winter. ACKNOWLEDGEMENTS
This research was supported by the research grant 11 (8)-76-ASR (I) from the Indian Council of Agricultural Research, New Delhi (to BBJ). We are thankful to the Department of State Fisheries, Govt. of West Bengal, for their cooperation which enabled us to collect data from Kalyani fish farm.
REFERENCES Alexander, M., 1971. Microbial Ecology. John Wiley and Sons Inc., New York, 511 PP. American Public Health Association, 1976. Standard Methods for the Examination of Water and Waste Water. Fourteenth edition. A.P.H.A., New York, 1193 pp. Brezonik, P.L., 1972. Nitrogen: Sources and transformation in natural waters. In: H.E. Allen and J.R. Kramer (Editors), Nutrients in Natural Waters. Wiley-Interscience Publications, New York, pp. l-50. Grant, D.J.W. and Patel, J.C., 1969. The non-oxidative decarboxylation of p-hydroxybenzoic acid, genetisic acid, protocatecuic acid and gallic acid by Klebsiella aerogenes (Aembacter aemgenes). Antonie van Leeuwenhoek. J. Microbial. Serol., 35: 325-343. Jana, B.B. and Barat, S., 1984. Ammonification as affected by oxygen level of water. Limnologica, 16: 67-70. Jana, B.B. and Patel, G.N., 1984. Measurements of some groups of microbialpopulations with special reference to denitrification in water and sediments of fish-farming ponds under mono- and polyculture systems. Int. Rev. Gesamten Hydrobiol., 69: 231-240. Jana, B.B. and Roy, SK., 1983. Estimates of microbial population involved in N cycle and their activity in water and sediments of fish ponds under mono- and polyculture systems. Int. Rev. Gesamten Hydrobiol., 68: 581-590. Jones, J.G., Simon, B.M. and Horsley, R.W., 1982. Microbiological sources of ammonia in freshwater lake sediments. J. Gen. Microbial., 128: 2823-2831. Kormondy, E.D., 1978. Concepts of Ecology. Prentice Hall of India Private Limited, New Delhi, 238 pp. Krause, H.R., 1964. Zur Chemie und Biochemie der Zersetzung von susswasser Organismen unter besonderer Berucksichtigung des Abbaues der organischen Phosphorkomponenten. Verh. Int. Verh. Limnol., 15: 549-561. Kruger, D., 1978. The influence of silver carp (Hypophthalmichthys molitrix) on eutrophication of the environment of carp ponds. 2. Active microflora changes in nitrogen conversion. Rocz. Nauk. Roln. Ser. H Rybactwo, 99: 33-54. Kuznetsov, S.I., 1968. Recent Studies on the Role of Microorganisms in the Cycling of Substances in Lakes. Verlag Wissenschaften, 301 pp. Little, J.E., Robert, E.S. and Gerald, R.C., 1969. Measurement of proteolysis in natural waters. Appl. Environ. Microbial., 37: 900-908. Meiklejohn, J., 1965. Microbiological studies on large termite mounds. Rhod. Zambia Malawi J. Agric. Res., 3: 67-79. Morris, D.R. and Koffron, K.L., 1967. Urea production and putrescine biosynthesis by Escherichia coli. J. Bacterial., 94: 1516-1519.
65 Niewolak, S., 1965. Badanie intensywnosciniektorych przemian azotu w jeziorach ilawskich w latach 1962-1963. Zesz. Nauk Wyszsz. Szk. Roln. Olsztynie, 20: 425, 6984. Rheinheimer, G., 1959. Mikrobiologische Untersuchungen iiber den Stickstoffhaushalt der Elbe. Arch. Mikrobiol., 34: 358-373. Rheinheimer, G., 1965. Mikrobiologische Untersuchungen in der Elbe zwischen Schnackenburg und Cuxhaven. Arch. Hydrobiol., 29 (Suppl.): 181-251. Rheinheimer, G., 1980. Aquatic Microbiology. Second edition. John Wiley and Sons, New York, 235 pp. Roy, S.K., 1984. Spatial and seasonal distributions of microbial populations involved in N cycle and their activity in fish farming ponds under traditional, monoculture and polyculture systems. Ph.D. Thesis, Kalyani University, Kalyani, West Bengal, India, 294 pp. 1981. Diversity of ammonifying bacteria. Hydrobiologia, 83: 153Sepers, A.B.J., 160. Shilo, M. and Rimon, A., 1982. Factors which affect the intensification of fish breeding in Israel. 2. Ammonia transformation in intensive fish ponds. Bamidgeh, 34: 101-114. Stewart, W.D.P., Sinada, F., Christofi, N. and Daft, M.J., 1977. Primary production and microbial activity in Scottish fresh water habitats. In: F.A. Skinner and J.M. Shewan (Editors), Aquatic Microbiology. Academic Press, London, pp. 31-54. Sugiyama, M. and Kawai, A., 1979. Microbiological studies on the nitrogen cycle in aquatic environment. VI. Metabolic rate of ammonium nitrogen in a gold fish culturing pond. Bull. Jpn. Sot. Sci. Fish., 45: 785-789.