Hydrobiologia 295 : 263-273, 1995 .Y S. Wong & N. F Y Tam (eds), Asia-Pacific Symposium on Mangrove Ecosystems .

263©1995. Kluwer Academic Publishers . Printed in Belgium

Effect of synthetic wastewater on young Kandelia candel plants growingunder greenhouse conditions

G. Z. Chen', S. Y Mao', N. F Y Tame, Y S . Wong3*, S. H. Lil & C. Y Lan''Research Institute of Environmental Science, Zhongshan University, People's Republic of China2Department of Biology and Chemistry, City Polytechnic of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong3Research CentreBiology Department, Hong Kong University of Science and Technology, Clear Water Bay,Kowloon, Hong Kong(*author for correspondence)

Key words: Mangrove, Kandelia candel, seedling, growth, wastewater, pollution

Abstract

A greenhouse experiment was performed to evaluate the effects of synthetic wastewater in three different strengths,NW, MW and CW, on the growth of the one-year old Kandelia candel (L .) Druce plants. NW had the characteristicsand strength similar to natural municipal wastewater while MW and CW contained five and ten times of the nutrientsand heavy metals in NW, respectively. Artificial seawater was used as the control . During one year wastewatertreatment experiment, Kandelia were found to withstand wastewater of high strength and toxic symptoms were notdetected in all plants . Synthetic wastewater with strength similar to the natural municipal sewage (NW) stimulatedplant growth. The plants treated with NW had significantly higher aerial and root biomass, taller stem than thosefound in the CW, MW and the control . Maximum growth, in terms of both stem height and total biomass, of allplants occurred in summer months, from June to September . With respect to the physiological and biochemicalactivities, CW and MW treated plants had significantly lower levels of chlorophyll a, total chlorophyll and catalaseactivity than those found in NW and control groups . In contrast, the proline content of plants treated with wastewaterwas similar to that of the control . These results suggest that normal wastewater (NW), attributed to its nutrients andtrace elements, enhanced plant growth . The medium (MW) and concentrated wastewater (CW) supported similaramount of plant growth as the control but the physiological and biochemical parameters indicate that these treatedplants might have been exposed to some kind of stress, probably due to the excess heavy metals present in MWand CW.

Introduction

Many investigations have shown that wetland (bothnatural and constructed) has a potential for treatingwastewater (De Jong, 1976 ; Gambrell et al., 1987 ;Gersberg et al., 1986 ; Wathugala et al., 1987). In awetland system, higher aquatic plant is the most obvi-ous biological component and is of great significance inremoving pollutants from sewage . The aquatic macro-phytes such as Phragmites australis, Typha capensisand T latifolia have been successfully employed topurify wastewater, through systems of reed ponds (DeJong, 1976; Rogers et al., 1991), sand or gravel fil-tration (Conley et al., 1991 ; Gersberg et al., 1986 ;

Wathugala et al., 1987), and natural marsh (Boyt et al.,1977; Reed etal., 1988). However, the ability of plantsto remove nutrients and pollutants from wastewaterhas not yet been fully examined . The major mech-anism for wastewater purification includes the plantuptake and the microbial transformation by bacteriaassociated with the plant rhizosphere (Gersberg et al.,1986; Reed et al., 1988 ; Rogers et al., 1991). Dif-ferent plant species with varied growth rates and pri-mary productivity levels would have different abilitiesto remove nutrients and pollutants from dischargedsewage. Mangrove plants, which are intertidal andwhich dominate the estuarine shores within tropicaland subtropical regions, are well known for their high

264

standing biomass and productivity (Lugo, 1980) . Theplants are perennial, consist of extensive root systemand specially adapted to the harsh environment of shift-ing aerobic and anaerobic, as well as alternating wet-ting and drying conditions (Lugo & Snedaker, 1974) .These features enable the mangrove communities towithstand and retain wastewater-borne nutrients andpollutants (Nedwell, 1974) . Clough et al. (1983) esti-mated that mangrove plants, by incorporation into theplant tissues, can annually immobilize very significantamounts of N and P with values around 150 to 250 kgN ha 1 and 10 to 20 kg P ha-1 , respectively .

On the other hand, sewage effluent will affect theplant communities of wetlands such as Eleocharissphacelala (sedge) and Typha orientalis (bulrush), pri-marily through changing the hydrological characteris-tics, the nutrient status and the pollutant levels (Cookeet al ., 1990; Kadlec, 1987). Mangrove plants, well-known to be resistant to environmental stresses such ashigh salinity and waterlogging, are also able to toleratehigh concentrations of nutrients (Clough et al., 1983 ;Nedwell, 1974) and heavy metals (Lin & Chen, 1990) .It has been suggested that the input of inorganic nutri-ents and trace elements from sewage discharges arenot likely to be deleterious to mangroves, and indeed,may well be beneficial to their growth, especially inmangrove communities which are limiting in nutrients(Boto, 1992 ; Clough & Attiwill, 1982; Onuf et al .,1977). Boto and Wellington (1983) reported that addi-tion of N and P to the mangrove communities resultedin 30% increase in mangrove growth coupled with ahigher nutrient content in plant tissues . However, it hasalso been reported that sewage is an important biot-ic factor contributing to the gradual disappearance ofmangrove vegetation near Bombay (Navalkar, 1951) .The effects of sewage addition to mangrove plants maydiffer significantly among different mangals as theyvary widely in their physical, chemical and biologicalproperties (Clough et al., 1983) . Moreover, the addi-tion of sewage may cause sub-lethal damages in plantssuch as changes in photosynthetic rates, chlorophyllconcentrations and enzymatic activities (Culic, 1984 ;Peng, 1990), which can be used as an early warningto wastewater pollution problems. The present studyis therefore aimed (1) to examine the possible effectsof sewage discharge on the growth, and the physio-logical and biochemical responses of young Kandeliacandel plants growing under greenhouse conditions ;and (2) to compare the impact of different strengths ofwastewater on mangrove plants .

Materials and methods

Experimental set-ups

Twelve polyethylene tanks (0 .7 x 0.5 x 0.4 m3) werefilled to a depth of 30 cm with 100 kg of soil collect-ed from a mangrove swamp in Futian National NatureReserve, Shenzhen Special Economic Zone, the Peo-ple's Republic of China . The mangrove soil was pre-viously air-dried, ground and passed through a 2-mmmesh sieve before placed in the tide tank. One-year oldKandelia candel plants were transplanted from Futianmangrove swamp to the tanks . Each tank was floodedwith artificial seawater (prepared by dissolving com-mercially available salts in deionized water and thesalinity was 1 .2-1 .6%) twice a day (at 10 am to 2 pmand 10 pm to 2 am) to simulate high tide . This gaveabout 16 hours exposure every day which simulatedthe natural tidal regime of Shenzhen area . During hightide period (about 8 hours per day), the base of theplants (about 2 cm from the soil surface) were sub-merged in the artificial seawater . Fifteen plants weregrown in each tank in a greenhouse for about 9 monthsto acclimatize to the greenhouse environment prior totreatment with wastewater. At the beginning of thewastewater treatment, the young plants reached anaverage height of 17 .7± 1 .6 cm, stem diameter of6.1± 0.5 mm, leaf number of 10± 2, and the aver-age leaf, stem, hypocotyl, root and total biomass were1 .74, 1 .50, 6 .06, 1 .92 and 11 .22 g dry weight, respec-tively. The twelve tide tanks planted with Kandeliawere divided into four groups (each group was trip-licated) . Three groups were irrigated with syntheticwastewater of three different strengths, namely natu-ral wastewater (NW), medium wastewater (MW) andconcentrated wastewater (CW), twice a week with thehydraulic loading of 1 .75 1 per tank per irrigation dur-ing the exposure period (i.e . at 5 pm of the day) . Thereason for adding wastewater during the exposure peri-od was to increase the amount of wastewater infiltratedinto the soil and to enhance the treatment efficiency .The remaining group (SW) was irrigated with artificialseawater and was used as a control . NW representedthe synthetic wastewater having its composition sim-ilar to the natural municipal sewage (Table 1), MWand CW had 5 and 10 times the amount of nutrientsand heavy metals of the NW, respectively . In betweentwo wastewater irrigation, all the tanks were flood-ed with seawater according to the tidal regime . Thisexperiment lasted for one year under the greenhouseconditions .

Table 1 . Characteristics of artificial wastewater with composition similar to the natural municipalwastewater (NW).

Determination ofphysiological/biochemicalresponses and growth of Kandelia plants

Plant growthPlant growth, in terms of stem height (H) and diam-eter at the base of the stem (D), was measured everymonth. The biomass values of different plant parts wereobtained by a non-destructive allometric technique(Snedaker & Snedaker, 1984) and was calculated everymonth . Twenty-one young Kandelia candel plants(about two-years old) were collected from Futian man-grove swamp. Stem height and diameter at the base ofthe stems were determined . The young plants weredivided into stem, leaves, hypocotyl, and root com-ponents, oven-dried (105 °C) and their weights wererecorded (Table 2). The least-square regression equa-tions using diameter and height as independent vari-ables were calculated according to : log biomass = a +blog (D2H) (Snedaker & Snedaker, 1984) . The allo-metric relations were highly significant (Table 3) andwere used for estimating biomass of various plant partsduring the experiment .

Physiological/biochemical measurementsAt the end of one-year treatment with wastewater, fiveto ten mature leaves (the third pair of the leaves fromthe apical part of the stem) of every plant from eachtank were collected, mixed and analyzed for vari-ous parameters . The content of chlorophyll a and b(extracted by acetone and measured at wavelength of663 and 645 nm ; Zhu et al., 1990), concentration offree proline (extracted by glacial acetic acid, triketohy-drindene and benzene, followed by measuring at wave-length of 515 nm; Zhu, 1983), and catalase activity(with H202 as the substrate and titrated with KMn04 ;

Huang & Chen, 1990) were determined . The photo-synthetic rate of five mature leaves from every plantof each treatment were measured by a portable pho-tosynthetic system (LI-6200, LI-COR Inc .) and thedata were calculated based on the light flux density of175.0 p mol m -2 s-1 , CO2 concentration of 380 mg1-1 and air temperature at 293.15 °K.

Statistical analyses

The mean and standard deviation of the triplicates ofall measured parameters in each group were calcu-lated. The results collected except the plant growthdata were treated with a parametric one-way analysisof variance (ANOVA) to test the significant differencebetween four treatments (three types of wastewater andthe control) . The plant growth, in terms of monthlyincrement in stem height and biomass, was also evalu-ated by two-way ANOVA, with treatment and time asthe main effects. The least significant difference (lsd)values at 5% probability level was calculated if theresults of the ANOVA indicated significant differences .All statistical analyses were performed by means of anIBM-compatible computing package 'SPSS' .

Results and discussion

Plant growth during one-year treatment withwastewater

Stem height and diameterThe monthly accumulation of stem height indicatesthat irrigation with wastewater of strength similar tothe natural municipal sewage (NW) led to best plant

265

Nutrient Compoundused

Concentration(mg l-1 )

Nutrient Compoundused

Concentration(mg 1-")

COD glucose 500.0 Cue+ CuC12 .2H20 2 .0NHa -N NH4C1 40.0 Zn2+ ZnSO4.7H20 5 .0N03 -N NaNO3 1 .0 Cd2+ CdCl2 .2H2O 0.1Organic N urea 10.0 Ni2+ NiC12 .6H20 1 .0POI-P KH2PO4 10.0 Pb2+ Pb(N03)2 1 .0K+ KCI 50.0 Crb+ K2Cr2O7 0.5Fe3+ FeCI3.6H20 30.0 Mn2+ MnCI2 .4H2O 5 .0

266

Table 2. Stem height (H), diameter at the base of the stem (D) and the biomass (ovendried weight, W) of 21 young Kandelia candel plants for regression analyses .

Plant No. D (cm) H (cm) Stem

1

0.678

6.8

0.72

0.538

15 .0

0.83

0.478

13.0

0.84

0.656

19.4

1.65

0.548

12.3

0.96

0.872

36.3

5.47

0.564

20.3

1.78

0.720

26.0

2.49

0.724

22.3

1.910

0.623

16.8

1.211

0.432

10.2

0.512

0.825

21.4

2.713

0.401

3.8

0.314

0.729

22.2

2.615

0.353

7.0

0.216

0.708

17.8

1 .617

0.822

30.4

3.818

0.551

8.2

2.119

0.583

16.9

1.220

0.669

14.5

1 .321

0.788

23.2

4.4

Table 3 . The regression equations showing the relationships between biomass(W), stem diameter (D) and stem height (H) of Kandelia plants .

Biomass

Regression equation

Correlation(dry weight, g)

Coefficient (r)

Stem log WS = -0.47 + 0.79 log (D 2 H) 0.93**Leaf log WL = -0.26 + 0 .96 log (D2 H) 0.70Aerial (stem + leaf) log WA = -0.05 + 0 .63 log (D2H) 0.93**Hypocotyl log WH = 0 .66 + 0.15 log (D2H) 0.62Root log WR = -0.22 + 0 .61 log (D2H) 0.82*Total biomass

logWT = 0 .78 + 0 .33 log (D2H)

0.89**

The regression analyses were calculated based on data from 21 plants shown inTable 2 ; * and ** indicated the r values were significant at probability levels of0 .05 and 0 .01, respectively, the r values without any asterisk meant statisticallynot significant (probability greater than 0 .05) .

growth, followed by CW, and MW treatment shared triplicated treatment group were very obvious . Onlysimilar height increment as the control (Fig . 1). The plants from tanks treated with NW had significantlygrowth of individual plants within the same treatment higher amount of stem growth than the other treatedgroup tended to differ profoundly . Variations among groups. At the end of the one year wastewater treat-15 individuals in the same tide tank and among the

ment experiment, the stem height of the plants from

Biomass (g)Leaf Hypocotyl Root Total

1 .1 5.3 1 .3 8 .40 .5 6.2 0 .4 7 .91 .7 5 .4 1 .2 9 .11 .4 5 .9 1 .5 10 .41 .6 4 .4 0 .9 7 .85 .5 8 .6 5 .7 25 .20 .9 6.6 2 .3 11 .53 .1 5 .5 3 .7 14 .71 .8 6.3 2 .2 12 .21 .0 5 .7 2 .6 10.51 .0 5 .8 0 .5 7.82.9 8 .4 2 .9 16.90.5 5 .5 1 .0 7.31 .5 6 .8 2 .8 13 .70.5 4 .8 0 .6 6.12.1 6 .8 3 .5 14 .01 .4 8 .9 3 .9 18 .00.8 3 .8 1 .2 7 .91 .2 3 .6 1 .7 7 .71 .4 5 .3 2.1 10.10 .6 6 .7 4.2 15 .9

NW tanks increased from an initial 17 .74 cm to thefinal 28 .14 cm, with an average of 10 cm total increasein stem height, whereas the total stem increment wasaround 7 cm in MW and the control (Table 4) . Theplants treated with concentrated wastewater (CW) hadintermediate level of growth . Similarly, the averagestem diameter of Kandelia followed the descendingorder of NW>CW>MW, control (Table 5). Theseresults suggest that addition of wastewater stimulat-ed stem growth as the mangrove soil is often deficientin nutrients and mangrove plants always response pos-itively to sewage discharge (Boto & Wellington, 1983 ;Clough et al., 1983). Tam et al. (1993) reported thatthe amount of bioavailable P and N in the native soil ofthe Futian mangrove fell within the range recorded inother mangrove ecosystems and could be consideredas nutrient deficiency . There was no inhibition in stemgrowth when the plants were irrigated with wastewa-ter containing very high content of nutrients and heavymetals such as MW and CW, implying that the man-grove plants, even at a relatively young growing stage,possessed resistance to wastewater pollution .

The monthly increment in stem height shows that,as expected, more active growth occurred during thesummer months, from July to October (some evenextended to November), with maximum growth inAugust in both treated and control plants (Fig . 2) . Verylittle growth occurred during the winter season. Thisgrowth pattern reflects that the ambient temperature isone of the most important factors affecting the growthof mangrove plants . Indeed, the distribution of man-gals is closely related to temperature as they can onlybe found in tropical and subtropical regions, at lati-tudes between 25 °N and 25 °S (Lugo & Snedaker,1974). Wastewater addition did not alter the growthpattern measured in terms of the monthly increase instem height.

Plant biomassFigure 3 shows that the pattern of the cumulativeincrease in total biomass of plants during the one yearstudy which was similar to that found in the cumu-lative stem height. The plants treated with NW hadthe highest total biomass, followed by CW, and MWand the control were similar. The differences betweenthe wastewater treated groups and the control werevery small during the initial 5 months (from Marchto July 1992), but the plants irrigated with NW hadmuch more growth than the other three groups fromAugust onwards (Fig . 3) . At the end of the study, total

267

biomass production in NW group was 66% more thanthat of the control . Clough et al. (1983) concluded thataddition of wastewater significantly increased the con-centrations of available nutrient such as NH4 -N andP04- -P of the mangrove soil, and enhanced both theplant productivity and the nutrient content of the planttissues . Therefore it is not surprised to find that dis-charge of NW resulted increases in both stem heightand biomass production . Similar research work exam-ined the effects of nutrient enrichment of Rhizophoramangle in Florida also indicated that the plants in fer-tilized sites had higher growth rates, greater additionsof leaves, reproductive parts and new lateral branches,and larger increases to existing stems (Boto & Welling-ton, 1983 ; Onuf et al., 1977). On the other hand,wastewater containing very high concentrations of N,P and various heavy metals such as CW did not causeany reduction in biomass . This could be related to thefact that the mangrove soils were capable to immobilizeexcess N, P and heavy metals and made them less avail-able to plant uptake . Tam & Wong (1992 & 1993) foundthat most of the P and heavy metals added to mangrovesoil from discharged sewage were bound to the man-grove soils either by adsorption on ion-exchange sites,incorporation into lattice structures or by precipita-tion as insoluble sulphide . Only a very small portionof these elements were bio-available . Similar findingswere also reported by Boyt et al. (1977) and Cookeet al. (1990). This explained why the growth of theplants was not inhibited by irrigation of CW. Further-more, the mangrove environment is highly variableowing to a combination of periodic fluctuations andextremes in physico-chemical parameters . This vari-ability also enables the mangrove flora and fauna tobecome highly adaptable to adverse conditions . It hasbeen reported that the growth of the seedlings of Avi-cennia alba, Rhizophora muoronata and R. manglewere not inhibited by Zn, Pb, Cd and Hg (Chen & Lin,1988) .

With respect to monthly increment, Fig . 4 revealsthat the maximum biomass production occurred fromJune to August, a bit earlier than the maximum increas-es in stem height (Fig . 2). As the biomass was calculat-ed according to the regression equation based on stemheight and diameter, this meant that the plants startedto have rapid growth by first expanding their diameterfollowed by an increase in stem height . The averagemonthly increases in biomass and stem height (over12 months of the experimental period) were shownin Table 4 . The root biomass contributed to quite asignificant portion of the total biomass, and the aerial

268

12

-0

Fig . 1 . Effects of wastewater in different strengths on cumulative increases in stem height during one year study . (0 : control; 0: NW; V :MW; T : CW) .

Table 4. Monthly increments in stem height, aerial (stem + leaves), root and totalbiomass of Kandelia candel plants treated with various kinds of wastewater .

The mean and standard deviation values (in brackets) of 12 months data on 45 individualplants of each treatment were calculated. The means followed by different superscriptswithin each column indicated that they were significantly different at a probability levelof 0 .05 according to ANOVA test .

to root biomass ratio was around 1 .55:1 . This reflects

Physiological/biochemical responsesthat mangrove plants had very extensive root systemin order to adapt to the stress environment .

Chlorophyll contentIn all wastewater treated plants, the chlorophyll a and bcontent was significantly lower than that of the control,

TreatmentHeight (cm)Monthly Increment

Biomass (g dry wt. per plant)Monthly Increment

Mean Minimum Maximum Aerial Root Total

Control 0.4886 0 .208 1 .142 0.158° 0.119a 0.249°(0.063) (0 .037) (0 .020) (0.049)

NW 0.8676 0.183 2 .350 0.208 6 0.1806 0.3856(0.368) (0 .038) (0 .013) (0.027)

MW 0.552a 0.117 1 .625 0.152° 0.121° 0.246°(0 .135) (0 .016) (0 .010) (0.038)

CW 0.717° 0.150 2 .183 0.180° 0.159° 0.305a(0 .293) (0 .053) (0 .053) (0.088)

1 .8

1 .5Uv

10

0 .0Apr May June

Table 5. Changes of stem diameter (cm) under differentwastewater treatments during one year study .

July Aug Sept Oct Nov Dec

Months

Jan Feb Mar

269

Fig. 2 . Monthly increments in stem height of plants treated with various kinds of wastewater and the control . (∎ : control ; 0: NW; ®: MW ;® : CW) .

with a descending order of: control>NW>CW>MW in Chl b than Chl a . The ratio of Chl a to Chl b was the(Table 6) . The Chl a and b contents of plants treated highest in plants treated with CW (Table 6), and thewith CW were 65% and 50% of the control, respec- ratio increased with the strengths of wastewater. Thetively. Wastewater treatment caused a bigger decline

effect of wastewater discharge on chlorophyll content

Months Control NW MW CW

March 92 0 .610 0.610 0.610 0.610April 0.603 0.621 0.616 0.626May 0.610 0 .628 0 .614 0 .620June 0.673 0 .663 0 .700 0 .684July 0.712 0,717 0 .733 0 .712August 0.730 0,736 0 .725 0 .707September 0.733 0.736 0 .711 0 .729October 0 .732 0.748 0.716 0.724November 0.741 0 .781 0.726 0.754December 0 .749 0.788 0.727 0.763January 93 0 .751 0.793 0.725 0 .761Feb 0 .752 0 .801 0.742 0.772March 0 .753 0.822 0.747 0.774

270

Mar Apr May June

MonthsFig . 3 . Effects of wastewater in different strengths on cumulative increases in total biomass during one year study . (0: control; 0: NW ; V :MW; V : CW) .

was similar to other studies on stress physiology ofmangrove plants . It has been found that the environ-mental stress not only caused a drop in chlorophyllcontent (Culic, 1984 ; Lin et al., 1984), it also changedthe ratio of chlorophyll a and b (Wang, 1990) . Lin et al.(1984) found that under high soil salinity (1% salini-ty), the synthesis and accumulation of chlorophyll inKandelia candel were reduced and the ratio of Chl a toChl b raised. The reduced chlorophyll level of plantstreated with wastewater suggest that the plant vigourand their physiological activity might have been affect-ed although the growth of these plants was not reducedby wastewater treatment .

Photosynthetic ratesDespite the lower chlorophyll content in plants treatedwith wastewater, the photosynthetic rates measured inthe treated groups, NW and CW, were significantlyhigher than that of the control (Table 7) . This suggeststhe nutrient input from wastewater treatment enhancedthe photosynthesis, in particular, the photosyntheticrate per unit chlorophyll molecule . Lugo et al. (1976)also concluded that gross photosynthesis appears to

July Aug Sept Oct Nov Dec Jan Feb Mar

be sensitive to terrestrial nutrient input and that thedevelopment of mangrove biomass is dependent onthe quantity of nutrients .

Proline content and catalase activityAll treated and control plants had similar level of freeproline but those treated with MW and CW had sig-nificantly lower catalase activity than NW and thecontrol (Table 7). Catalase is an enzyme commonlyfound in plant tissue and its activity is closely relat-ed to the degree of pollution . It has been found thehigher the pollutant concentration, the more the leaveswere stressed, the lower the activity of catalase (Peng,1990). This suggests that the young plants treated withMW and CW might have been subjected to some kindof stress . The high heavy metal concentrations of thesewastewater treatment might have created the stressedenvironment which required further investigation . Thepresent results also suggest that catalase activity mightbe a more sensitive parameter in indicating cellulardamages caused by wastewater treatment than free pro-line level, another parameter commonly used to relatethe stress resistance of many plants (Peng, 1990) .

m

1 .2 -

1 .0

m

0 0 .8

MA

19

0 .4

9300 .2

0 .0

Conclusion

The present greenhouse experiment demonstrates thatthe young plants of Kandelia candel, the most dom-inant mangrove species in Hong Kong and South-east China, could tolerate the pollution impact fromsewage effluents. Treatment with wastewater of vari-ous strengths did not reduce any plant growth, in termsof stem height, diameter, biomass and photosynthetic

iii

Apr May June July Aug Sept Oct Nov Dec Jan Feb Mar

MonthsFig. 4. Monthly increments in total biomass of plants treated with various kinds of wastewater and the control (∎ : control ; 11 : NW; ®: MW ;® : CW).

Table 6. Chlorophyll content (mg g t dry weight) of K. candel plants at the end ofone-year wastewater treatment experiment .

The mean and standard deviation values (in brackets) of three parallel groups (with 15plants in each group) were calculated . The mean values followed by different superscriptswithin each column indicated that they were significantly different at a probability levelof 0 .05 according to ANOVA test .

∎

i

i

271

rates. Addition of wastewater with nutrient concentra-tion similar to the natural municipal sewage stimulatedplant growth. This could be attributed to the extra nutri-ents and trace elements available in NW Although theKandelia plants appeared to be tolerant to wastewaterof high strength (even up to 10 times of the concentra-tions ofthe natural municipal sewage) and maintainedtheir growth under severe water pollution situation,irrigation with concentrated wastewater (MW or CW)

Treatment Chi a Chl b Total Chi a: Chl b

Control 1 .164° (0 .015) 0.744° (0 .039) 1 .9084 (0 .054) 1 .565° (0 .065)NW 1 .106 b (0.130) 0.643b (0.050) 1 .749b (0.177) 1 .719b (0.069)MW 0.543` (0.046) 0.286` (0.079) 0.829` (0.112) 1 .899' (0.445)CW 0.7564 (0 .090) 0.3704 (0 .027) 1 .1264 (0 .113) 2.045c (0.107)

272

Table 7. Physiological and biochemical properties of K . candel plants afterreceiving wastewater of different strengths for one year .

Treatment Proline content

Catalase activity

Photosynthetic rate(µg g-') (mg g-1 min- 1 ) (µcool CO2 M-2 S-1)

Control 144 .65a (12 .65) 0.334" (0 .029) 3 .458" (0 .209)NW 134.58" (11 .84) 0 .348" (0.008) 4.724' (0 .876)MW 149.81" (20 .82) 0 .301' (0 .016) 3.591" (0 .157)CW

154.71" (21,22)

0.289' (0 .007)

4.384' (0 .782)

All data were expressed on fresh weight basis ; the mean and standard deviationvalues (in brackets) of three parallel groups (with 15 plants in each group) werecalculated . The mean values followed by different superscripts within each col-umn indicated that they were significantly different at a probability level of 0 .05according to ANOVA test.

did create a slightly stressed environment. The phys-iological and biochemical indicators such as contentof chlorophyll a and b and catalase activity decreasedsignificantly when the plants were exposed to MW andCW. As the present study only lasted for one year,further investigation on the longterm effect of wastew-ater treatment on growth and physiological activitiesof mangrove plants should be carried out to furtherunderstand the resistance mechanisms of mangrovesto high levels of nutrients and heavy metals .

Acknowledgments

The authors would like to thank the technicians ofFutian Nature Reserve, Shenzhen Special EconomicZone and Zhongshan University, Guangzhou, the PRCfor their assistance in field sampling and laboratoryanalyses . We would also like to express our gratitudeto the Biotechnology Research Institute and the HongKong Research Grant Council for their financial sup-port .

References

Boto, K . G ., 1992 . Nutrients and mangroves . In : Connel D . W. &D. W. Hawker (eds .) Pollution in Tropical Aquatic Systems . CRCPress, Inc . Ann Arbor, London : 129-145 .

Boto, K . G . & J . T. Wellington, 1983 . Nitrogen and phosphorusnutritional status of a northern Australian mangrove forest . Mar.Ecol. Press Ser. 11 : 63-69 .

Boyt, F. L ., S . E . Baylay & J . Zoltek, 1977 . Removal of nutrientsfrom treated municipal wastewater by wetland vegetation. J. Wat.Pollut . Cont . Fed . 49 : 789-799 .

Chen, R . H . & P. Lin, 1988 . Influence of mercury and salinity onthe growth of seedlings of three mangrove species . UniversitatisAmoiensis Acta Scientiarum Naturalium 27 : 110-115 .

Clough, B . F. & P. M . Attiwill, 1982 . Primary productivity of man-groves. In : B . F . Clough (ed .) Mangrove Ecosystems in Australia .Miami, FL : 213-222 .

Clough, B . F., K . G. Boto & P. M . Attiwill, 1983 . Mangrove andsewage : are-evaluation. In : H . J . Teas (ed .), Biology and Ecologyof Mangroves, Vol. 8 . Dr W. Junk Publishers, Lancaster: 151-161 .

Conley, L . M., R . I. Dick & L . W. Lion, 1991 . An assessment of theroot zone method of wastewater treatment. Res . J . Wat . Pollut.Cont. Fed . 63 : 239-247 .

Cooke, J . G ., A . B . Cooper & N . M . U. Clunie, 1990 . Changes in thewater, soil, and vegetation of a wetland after receiving a sewageeffluent . New Zealand J . Ecol . 14 : 37-47 .

Culic, P., 1984 . The effects of 2,4-D on the growth of Rhizophorastylosa Griff. seedlings . In : H . J . Teas (ed .), Physiology andManagement of Mangroves, Vol . 9 . Dr . W . Junk Publishers, TheHague: 57-63 .

De Jong, J ., 1976 . The purification of wastewater with the aid of rushor reed ponds . In : Jr. Towbier & R, W. Pierson (eds .), BiologicalControl of Water Pollution . University of Pennsylvania Press,Philadelphia : 133-139.

Gambrell, R . P., R . A . Khalid & W. H . Jr. Patrick, 1987 . Capacity of aswamp forest to assimilate the TOC loading from a sugar refinerywastewater stream . J . Wat. Pollut. Cont. Fed . 59 : 897-904 .

Gersberg, R . M., B . V. Elkins, S . R . Lyon & C . R. Goldman, 1986 .Role of aquatic plants in wastewater treatment by artificial wet-lands . Wat . Res . 20 : 363-368 .

Huang, X . L . & R . Z. Chen, 1990 . An Experimental Handbook ofSeed Physiology. Agriculture Press, Beijing : 122-124 .

Kadlec, J . A., 1987 . Nutrient dynamics in wetlands . In : Reddy K. R .& W. H . Smith (eds), Aquatic Plants for Water Treatment andResource Recovery, Magnolia, Publishing Inc ., Orlando, Florida:393-419 .

Lin, P. & R . Chen, 1990. Role of mangrove in mercury cyclingand removal in the Jiulong estuary . Acta Oceanologica Sinica 9:622-624 .

Lin, P, D . H . Chen & W. J . Li, 1984 . A preliminary study of the inter-relationship between the physiological characteristics of certainenzymes in the leaves of mangrove plants and soil salinity in tidalswamps . Universitatis Amoiensis Acta Scientiarum 27 : 110-115 .

Lugo, A. E ., 1980. Mangrove ecosystems : successional or steadystate? Biotropica 12 : 65-72.

Lugo, A. E . & S . C. Snedaker, 1974. The ecology of mangroves .Annu. Rev. Ecol . Syst. 5: 39-64 .

Lugo, A . E ., M. Sell & S . C . Snedaker, 1976 . Mangrove ecosystemanalysis . In : B . C. Patten (ed .), Systems Analysis and Simulationin Ecology . Academic Press, New York : 113-145 .

Navalkar, B . S ., 1951 . Succession of the mangrove vegetation ofBombay and Salsette Islands . J . Bombay Nat. Hist. Soc . 50 : 157-161 .

Nedwell, D . B ., 1974 . Sewage treatment and discharge into tropicalcoastal waters . Ecology 5 : 187-190 .

Onuf, C . P., J. M . Teal & I . Valiela, 1977 . Interactions of nutrients,plant growth and herbivory in a mangrove ecosystem . Ecology58:514-526 .

Peng, S . Q ., 1990 . The Stress Physiological Fundamentals of Plants .University of Northeast Forestry Press, Haerbin, 106 pp .

Reed, S . C ., E . J. Middlebrooks & R . W. Crites, 1988 . NaturalSystems for Waste Management and Treatment. McGraw-HillBook Co., New York, N .Y

Rogers, K . H ., P. F. Breen & A. J . Chick, 1991 . Nitrogen removal inexperimental wetland treatment systems : evidence for the role ofaquatic plants . J . Wat. Pollut . Cont. Fed . 63 : 934-941 .

Snedaker, S . C . & J . G . Snedaker, 1984 . The Mangrove Ecosystem :

273

Research Methods . United Nations Educational, Scientific andCultural Organization Published, Richard Clay (The ChaucerPress) Ltd., U .K .

Tam, N . F. Y & Y S . Wong, 1992. Nutrient and heavy metal retentionin mangrove sediment receiving wastewater. Proc . Int. SpecialistConf. on Wetland Systems in Water Pollution Control . 30 Nov .-3 Dec. 1992, Sydney, Australia: 45 .1-45 .9 .

Tam, N. F. Y. & Y. S . Wong, 1993 . Retention of nutrients and heavymetals in mangrove sediment receiving wastewater of differentstrengths . Envir. Technol . 14 : 719-729 .

Tam, N . F. Y, S . H . Li, C. Y. Lan, G . Z . Chen, M . S . Li & Y S . Wong,1993 . Nutrients and heavy metal contamination of plants and sed-iments in Futian mangrove forest . Presented at the Asia-PacificSymposium in Mangrove Ecology, January 1993, Hong Kong.

Wang, H. X ., 1990 . Fundamentals of Pollution Ecology . YunnanUniversity Press, Kunming: 91-94 .

Wathugala, A . G ., T. Suzuki & Y Kurihara, 1987 . Removal of nitro-gen, phosphorus and COD from wastewater using sand filtrationsystem with Phragmites australis . Wat. Res . 21 : 1217-1224 .

Zhu, G. L ., 1983 . Determination of free proline in plants . PlantPhysiol . Commun . 1 : 35-37 .

Zhu, G . L ., H . W. Zhong & A . Q . Zhang, 1990 . Plant PhysiologicalExperiments . Beijing University Press, Beijing : 122-124.