dilution were plated in to R2A agar (Difco laboratories, USA) supplemented with .... column (Wynne and Rhee, 1988). association between phyto- and bacterio-.
Indian J. Fish., 53(2) : 167-173, Apr.-Jun., 2006
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Water alkaline phosphatase activity and phosphorus availability during summer in inland water bodies S.K. MANNA, A.B. SOM AND S. SAMANTA Central Inland Fisheries Research Institute (ICAR), Barrackpore, Kolkata - 700 120, India
ABSTRACT Phosphorus is a limiting nutrient in most freshwater environments. Alkaline phosphatase enzyme plays an important role in phosphorus cycling and may be decisive in primary and secondary production. In the present investigation total water alkaline phosphatase activity was high during summer in all the water bodies. Relative contributions of algal, bacterial and dissolved fractions to total water phosphatase activity were 11-50, 3-8 and 41-83% respectively. Unlike cell free and bacterial phosphatases, phytoplankton specific enzyme activity was positively correlated to orthophosphate concentration. Algal specific enzyme activity was low suggesting P-uptake to be a more important process than P regeneration in hyper-eutrophic sewage fed ponds.
Introduction Phosphorus (P) may be a limiting nutrient in most lacustrine and certain marine environments. Orthophosphate (o-P) is a convenient form of phosphorus, which is immediately available to planktonic community. However, during summer season concentration of available soluble reactive phosphorus (SRP, o-P) becomes very low in water column due to rapid uptake (Ramm and Scheps, 1997) and efficiency of utilization of dissolved organically bound phosphates (DOP) may be decisive in primary and secondary production (Currie and Kalff, 1984). Algae and bacteria express phosphatase enzymes mostly at their cell periphery (i.e., they are ectoenzymes) that hydrolyse the ester bond between phosphate and organic
moiety releasing o-P for cellular assimilation. Role of alkaline phosphatase (ALP) in P regeneration has been widely studied in aquatic environments and ALP activity has been used as an indicator of potential organic phosphorus mineralization (Newman and Reddy, 1992; Hantke et al., 1996; Spijkerman and Coesel, 1998; Berman et al., 1990; Manna et al., 2004). It has also been noted that ALP is expressed at higher levels under phosphorus deficiency states and high ALP activity has been considered as an indicator of phytoplankton community P deficiency (Pettersson, 1980; Gage and Gorham, 1985; Jansson et al., 1988; Rose and Axler, 1998). However, this has been debated by other workers arguing that ALP is an indicator of productive system
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(Heath and Cooke, 1975; Huber and Hamel, 1985; Pick, 1987; Jamet et al., 1997). The present work was aimed at defining the relationship between ALP activity with phosphorus availability and phytoplankton load in different aquatic environments and to know if water ALP was primarily of algal or bacterial origin during summer season.
Materials and methods The study was conducted during MayJuly, 2000 in five inland water bodies located in West Bengal, India (Table 1), detailed descriptions of which are available elsewhere (Manna et al., 2004). Briefly, water samples were collected from euphotic zone, at about 0.4-0.6 m depth and immediately filtered through 100 µm mesh size net to remove zooplankton. Sampling was done in 3 occasions at 15-18 days intervals. Chlorophyll-a (chl-a) was estimated spectrophotometrically following filtration and acetone extraction, and inorganic phosphate was measured by stannous chloride method (APHA, 1995).
For estimation of aerobic plate count of bacteria samples were serially diluted in normal saline solution and 1ml of each dilution were plated in to R2A agar (Difco laboratories, USA) supplemented with cycloheximide (25 mg/litre) following pour plate method; colonies were counted after 1 month of incubation at 30 oC. Phosphatase activity in particulate fraction of water was measured by filter method described by Berman et al. (1990) using Tris buffer (0.05 M, pH8.6, containing 1mM MgCl 2 ) and pnitrophenyl phosphate (pNPP, 10 mM) as substrate. About 25-100 ml water was passed through membrane filters of different porosity using vacuum at less than 100 mm Hg. Enzyme activity in size fraction 1.20-100 µm and 0.22-1.20 µm were accounted for algal and bacterial phosphatase respectively. Dissolved ALP was measured in 0.22 µm filtrate by test tube method (Berman et al., 1990).
Results Some of the limnological characters of the water bodies are given in Table 1.
TABLE 1: Description of water bodies Area (ha)
Av. depth (m)
Pond A
0.4
1.9
Av. temp. (oC) 30
Beel
30
1.4
30.2
Sewage pond2
0.56
0.8
30.6
Sewage pond4
0.56
0.8
30.6
River Ganga
Total 4.2 (at length sampling 2525 site) km.
Allochthonus input
Remarks
Nil to negligible. vegetation. Moderate, storm water
Some littoral
Profuse, regular, treated city sewage. Profuse, regular, treated city
sewage. 29.4 Carried from upstream. Some city sewages.
Closed, partly macrophyte infested. Samples taken from macrophyte-free areas. Bloom round the year. Fish culture ponds. Bloom round the year. Fish culture ponds. Study area is a tidal zone. Samples taken in low tide.
Water alkaline phosphatase activity in summer Chl-a concentration ranged between 6.27 µg l -1 and 372.33 µg l -1. High Chl-a concentrations were recorded in sewage fed ponds receiving organic rich allochthonus input. The river water o-P concentration was comparatively higher than few other water bodies: sharp increase in concentration was recorded in second and third sampling due to headwater contribution. In sewage-fed fisheries, water flows from first pond to the second and so on in a series. Thus nutrient (including P) loading are supposed to be higher in pond 2 than in pond 4. Strikingly only traces of inorganic phosphate were recorded in pond 2 than the 4 (Fig. 1). Water o-P was negatively related with Chl-a concentration (n = 15, P
