August 2012. Promoters Prof. Dr. Eng. Peter Goethals Prof ... · Ecological study of a large scale treatment pond system in Ho Chi Minh City (Vietnam) PREFACE This work is the reflection

Ecological study of a large scale treatment pond system in Ho Chi Minh City (Vietnam)

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“The author and promoters give permission to put this thesis to disposal for consultation and to

copy parts of it for personal use. Any other use falls under the limitations of copyright, in

particular the obligation to explicitly mention the source when citing parts out of this thesis”.

August 2012.

Promoters

Prof. Dr. Eng. Peter Goethals Prof. Dr. Ir. Ingmar Nopens

Author

Khiet Bui


PREFACE

This work is the reflection of a year of research, but also the support of a number of people

without whom it never would have come to this I would them this therefore like to thank. In the

first place, my supervisors, Professor Peter Goethals and Professor Ingmar Nopens, I want thank

you for the opportunities they offered me to carry out this research and to successful conclusion.

Thank you for the tireless input of ideas that this work for me so have made interesting.

I would like to express sincere gratitude and profound to Dr. Dao Thanh Son and Ngo T.T.

Huyen, who were enthusiastic and helpful during time to implement the project. I also give many

thanks to staff members of Binh Hung Hoa WWTP for their cooperations.

I want to thank to all my friends who are living and studying in Belgium and other countries for

their encouragement and sincere susggestions. For those who lives in OBSG, you know that you

are in my heart also.

I would also like to thank some other important people in my life, my parents, my brothers and

my sisters for their confidence in me; especially, YOU, my endless moral support during my

living and working in Ghent.

Finally my sincerely thanks go to all people who helped me in various ways but whose names

are not listed here


LIST OF TABLES ...................................................................................................................... i

LIST OF FIGURES ................................................................................................................... ii

LIST OF ABBREVIATIONS.................................................................................................... iii

ABSTRACT ............................................................................................................................. iv

I. Introduction .........................................................................................................................1

II. Literature review .................................................................................................................2

1 WASTE STABILIZATION PONDS ................................................................................2

1.2 Why treat wastewater ....................................................................................................2

1.3 Waste stabilization ponds ..............................................................................................3

1.4 Processes of waste stabilization ponds ...........................................................................5

1.5 Physical and chemical factors ........................................................................................8

2 PLANKTON AND WASTE STABILIZATION PONDS ............................................... 10

2.1 Introduction ................................................................................................................. 10

2.2 Plankton in waste stabilization ponds ........................................................................... 11

3 TOXICITY .................................................................................................................... 13

3.1 Biology of Daphnia magna .......................................................................................... 13

3.2 Daphnia and toxicity studies ........................................................................................ 14

III. Material and methods ..................................................................................................... 15

1 THE WASTE STABILIZATION POND ........................................................................ 15

1.1 Study area .................................................................................................................... 15

1.2 Description of the treatment process ............................................................................ 15

2 SAMPLING FOR MONITORING ON ENVIRONMENTAL PARAMETERS AND

PLANKTON ......................................................................................................................... 17

2.1 Physical and chemical parameters ................................................................................ 17

2.2 Sampling for phytoplankton and zooplankton .............................................................. 17

2.3 Phytoplankton and zooplankton identification and counting ......................................... 18

2.4 Diversity index ............................................................................................................ 18

2.5 Brachionus:Trichocerca index...................................................................................... 19

2.6 Evenness ..................................................................................................................... 19

3 TOXICITY STUDY ....................................................................................................... 19

4 STATISTICAL ANALYSIS........................................................................................... 20

IV. RESULTS ...................................................................................................................... 21

1 Water quality .................................................................................................................. 21


1 Physical parameter.......................................................................................................... 21

2 Plankton communities ................................................................................................... 21

2.1 Phytoplankton.............................................................................................................. 21

2.2 Zooplankton ................................................................................................................ 24

2.3 Correlation between phytoplankton, zooplankton and environmental factors................ 28

3 Toxicity test.................................................................................................................... 30

3.1 Survival rate ................................................................................................................ 30

3.2 Maturation ................................................................................................................... 31

3.3 Reproduction ............................................................................................................... 31

3.4 Fecundity and malformation ........................................................................................ 32

V. Discussion ............................................................................................................................ 34

1 Water quality .................................................................................................................. 34

1 Physical and chemical parameters ................................................................................... 34

2 Plankton communities ................................................................................................... 35

2.1 Phytoplankton.............................................................................................................. 35

2.1.2 Diversity indices ....................................................................................................... 36

2.2 Zooplankton ................................................................................................................ 37

2.3 Correlation between phytoplankton, zooplankton and environmental factors................ 39

3 Toxicity test................................................................................................................... 40

3.1 Survival rate ................................................................................................................ 40

3.2 Maturation ................................................................................................................... 40

3.3 Reproduction ............................................................................................................... 41

3.4 Fecundity and malformation ........................................................................................ 42

VI. CONCLUSIONS AND PERSPECTIVES ........................................................................... 43

6.1 Conclusions ................................................................................................................. 43

6.2 Perspectives ................................................................................................................. 43

VII. REFERENCES .................................................................................................................. 44


i

LIST OF TABLES

Table 1. Major constituents of typical domestic wastewater

Table 2. Comparison of factors of importance in wastewater treatment in developed and developing

countries

Table 3. Algae found in wastewater stabilization pond

Table 4. General ecology of WSP’s zooplankton

Table 5. Criteria of pollution by Shannon – Wiener diversity index

Table 6. Criteria of diversity by Simpson index

Table 7. Wastewater toxicity classification (Persoone et al., 2003)

Table 8. Pond performance as an average of one month’s result.

Table 9. List of phytoplankton taxa identified in BHH wastewater treatment plant

Table 10. Structure of phytoplankton

Table 11. Diversity indices and eveness of phytoplankton in three maturation ponds

Table 12. Structure of zooplankton

Table 13. A list of zooplankton recorded in three maturation ponds during the study period

Table 14. Diversity indices and eveness of zooplankton in three maturation ponds

Table 15. Results of Brachionus – Trichocerca quotient in three maturation ponds

Table 16. Spearman’s correlation between main groups of phytoplankton and zooplankton and water pa-rameters

Table 17. The number of individual Daphnia magna mother miscarriage and birth defects


ii

LIST OF FIGURES

Figure 1. Typical layout of a waste stabilization pond system: A, anaerobic pond; F, falcutative pond,

M1-Mn: maturation pond.

Figure 2. Major reactions in a waste stabilization ponds (from Shammas et al., 2009)

Figure 3. General plan of the wastewater treatment plan

Figure 4. Diagram of Binh Hung Hoa wastewater treatment plan procedure

Figure 5. Phytoplankton succession in taxonomic orders

Figure 6. Microscopic pictures of a) Merismopedia tenuissima and b) Cyclotella sp.

Figure 7. Two zooplankton species firstly recorded in Vietnam. a) Asplanchna amphora and b) Lecane robusta

Figure 8. Zooplankton succession in taxonomic orders

Figure 9. a) Brachionus angularis, b) B. caudatus, c) B. calyciflorus, d) B. urceous, e) Filinia longiseta and f) Testudinella elliptica

Figure 10. Survival rate of D. magna: a) influent, b) effluent

Figure 11. Maturation age of the Daphnia magna (days ± standard deviation).. n = 15: number of

experimental animals.

Figure 12. Number of D. magna offsprings born. a) influent, b) effluent


iii

LIST OF ABBREVIATIONS

Abbreviation Explanation

BOD5 5-day Biochemical Oxygen Demand

COD Chemical Oxygen Demand

DO Dissolved Oxygen

EC Electrical Conductivity

HCMC Hochiminh City

TCVN Vietnamese standard

VFA Volatile Fatty Acid

WSP Waste Stabilization Pond


iv

ABSTRACT

An evaluation of opperation of wastewater treatment pond system in Binh Hung Hoa, HCMC,

Vietnam was conducted based on the composition and density of phytoplankton and

zooplankton. Environmental parameters (temperature, pH, EC, DO, BOD5 and NH3-N) were also

evaluated based upon Vietnamese standard TCVN 5945:2005 (Discharge standards for industrial

wastewater allowed to be discharged into the water bodies other than those water bodies using

for sources of domestic water supply). Despite the fact that all parameters of the effluent satified

TCVN 5945:2005, the results from phytoplankton and zooplankton showed the moderate

pollution of the effluent. A toxiciy test was also conducted to gain more insight about the

efficiency of the system. The results of chronic test showed significant differences between the

influent and effluent. The influent had adverse effects on survival rate, days of maturation,

number of offsprings and fecundity of Daphnia magna. However, the effluent affected more on

the number of offsprings. In addition, the influent was classified in Class III (toxic).


1

I. INTRODUCTION

Hochiminh city is one of the largest cities in Vietnam, with population more than 7 millions

(2009); however, infrastructure and utility service have not kept pace with the development.

Therefore, low-income area have developed. Among of these, Tan Hoa – Lo Gom canal basin is

one of the most concentrated population communities. The area is located in south – west of the

inner city, with total area is 2.498 ha, including districs of Tan Binh, 11, 6, 8 and Binh Chanh.

The population is around 750.000; there are many industrial enterprises registered in the area and

thousands of house-based factories, while about 470.000 people drains directly to the canal.

Together, these create a big challenge for managers of the city. In this context, in June 1997,

there was an agreement signed between Belgian and Vietnamese Governments to solve the

problems in the area, namely “Tan Hoa Lo Gom Canal Sanitation and Urban Upgrading Project”.

The waste water treatment plant in Binh Hung Hoa, a ward in HCMC is part of the project. The

treatment plant used aerated lagoon and stabilization pond technology to treat wastewater from

nearby Den canal. Den canal has an area of 785 ha and population in the area is around 120.000

(1999). The canal is also received untreated wastewater from surrounding industrial activites,

creating black colour and bad oudour in the water with physico-chemical parameters as follow:

SS 250mg/L, BOD5 200mg/L, COD 300mg/L, NH3-N 25mg/L, and pH 6.5 - 7 (Smet et al.,

2006).

The objectives of this study are i) to characterize the phytoplankton and zooplankton community

present in three maturation ponds. Through a quantitative evaluation, the distribution of algae

and zooplankton is determined, and the efficiency of the pond system is also evaluated by using

phytoplankton and zooplankton; ii) to examine the toxicity effects of influent and effluent on

Daphnia magna since there are a lot of small-scale industrial facilities in the surrounding area .

Through the experiment, we can gain more insight about the efficiency of the system.


2

II. LITERATURE REVIEW

1 WASTE STABILIZATION PONDS

1.1 Introduction

Domestic wastewater is water that is used for the purpose of eating, living, bathing, cleaning the

house ... of residential areas, public facilities, service facilities ... and wastewater is formed in the

process of human activities. Characteristics of waste water is a large organic matter content,

which contains many microorganisms, including pathogenic microorganisms. Also in the waste

water there are many decomposing-organic matter bacteria, which are essential for metabolisms

of contaminants in the water. Table 1 shows the major constituents in domestic wastewater.

Table 1. Major constituents of typical domestic wastewater

Constituents Concentration (mg/l)

Strong Medium Weak

Total solids 1200 700 350

Dissolved solids (TDS) 850 500 250

Suspended solids 350 200 100

Nitrogen (as N) 85 40 20

Phosphorus (as P) 20 10 6

Chloride 100 50 30

Alkalinity 200 100 50

Grease 150 100 50

BOD5 300 200 100

Sources: Pescod (1992)

Such wastewater of urban, residential and service establishments, public buildings is in large

quantities, high levels of contaminants, many bacteria are one of the main pollution sources for

the aquatic environment.

1.2 Why treat wastewater

Due to industrialization, urbanization and the rapidly increasing in population, water resource is

under pressure of pollution. Water pollution in urban areas is best seen in Hanoi and Ho Chi

Minh cities. In these two cities, for a long time, there was no centralized wastewater treatment

system, and wastewater was directly discharged into the receiving water (rivers, lakes, canals). In


3

addition, many manufacturing facilities, most hospitals and large medical facilities do not have

wastewater treatment system. Today, pollution levels in the canals, rivers and lakes in large cities

are very heavy. If water pollution is not solved soon, it will directly affect people's health and

deterioration of the environment.

There are many methods commonly used in treatment of domestic wastewater: chemical

methods, chemical and physical methods and biological methods. However, due to the fact that

there are different concerns of factors of importance in wastewater treatment between developed

and developing countries (Table 2) and many developing countries have a warm and hot climate.

Therefore, the choice of wastewater treatment should concern all these issues.

Table 2. Comparison of factors of importance in wastewater treatment in developed and developing

countries

Factor Developed countries Developing countries

Efficiency C***** ****

Reliability C***** C*****

Sludge production *** C*****

Land requirement C***** **

Environmental impact **** **

Operational cost *** C*****

Construction cost ** C*****

Sustainability *** C*****

Simplicity * C*****

Note: C, critical; *****, extremely important → *, no impact

Source: Von Sperling (1996)

1.3 Waste stabilization ponds

Biological methods have gain much more attention due to their low capital and operating cost,

true destruction of organics, oxidation of wide organic compounds, removal of reduced organic

compounds, operational flexibility and reduction of aquatic toxicity (Schultz, 2005). Among of

the biological processes, the stabilization pond is one of the simplest forms. The advantages of

stabilization pond are that they are simple, low-cost, highly efficient and robust (Mara, 2003).

However, it also contains disadvantages: the ponds must be lined or constructed in clay soil to

prevent leakage; there may be chances of overflow when heavy rainfall occurs; odors may be a


4

problem if there are extended periods of overcast windless days; a lot of space is needed;

potential danger for people, especially children. When comparing with other treatment methods,

waste stabilization ponds is the most important method for wastewater treatment in developing

countries since land space is available and temperature is suitable for the process (Mara, 2003).

Examples can be found in France (Racault & Boutin, 2005), New Zealand (Archer & Mara,

2003), but also in other tropical regions like in Tanzania (Mbwele et al., 2003), Cameroon

(Noumsi et al., 2005), Venezuela (Botero et al., 1997) and Ghana (Hodgson, 2000). However, a

big concern needs to be taken is that waste stabilization ponds usually serve communities which

have populations of 10.000 or fewer (Federal Water Quality Administration, 1970).

According to Mara (2003), stabilization ponds can be divided into three general classes:

anaerobic, falcutative and maturation ponds. Figure 1 shows a typical layout of a waste

stabilization pond system.

Figure 1. Typical layout of a waste stabilization pond system: A, anaerobic pond; F, falcutative pond,

M1-Mn: maturation pond.

From: Mara (2003)

1.3.1 Anaerobic ponds

These are usually the first type of ponds of the system, with depth of 2-5 m. Organic load is very

high ( > 100 g BOD/m3.day, equivalent to 3000 kg/ha.day) and can reduce BOD up to 80%.

However, regular desludging is required due to gradual accumulation of digested solids.

1.3.2 Falcutative ponds

There are two types of falcutative ponds: primary falcutative ponds which receive water from

premilinary treatment and secondary falcutative ponds which receive water from anaerobic


5

ponds. The depth for these ponds is around 1 m, and are designed mainly for BOD removal on a

basis of low BOD loading (100-400 kg/ha.day) to allow a healthy growth of algal popultion.

1.3.3 Maturation ponds

The main function of maturation ponds is to reduce the number of pathogens in the effluent of

falcutative ponds. Depths of the ponds are usually 1 m and these ponds contain a great diversity

of algal genera. BOD, suspended solids and nutrients are removed very slowly.

In addition there is also another type of pond that is categorized in stabilization ponds (MAF,

2005), called aerated lagoon. This is a unit in which oxygen supply replies almost on mechanical

aeration devices and an activated sludge unit operated without sludge return (Mara, 2003). The

aerated lagoon is equivalent to falcutative pond except that the former is deeper and can receive

organic loading from medium to high.

1.4 Processes of waste stabilization ponds

Waste stabilization pond is one of the simplest forms of biological treatment processes. It serves

many purposes, including: i) storage wastewater, ii) settling and removal of solid particles, iii)

equalization, iv) aeration, v) biological treatment, and vi) evaporation. Despite of the simplicity

of stabilization ponds, many processes – chemically and biologically occur in a pond. Figure 2

demonstrates the major reactions that take place in a pond, and they can be listed as follow:

a) sedimentation

b) aerobic decomposition

c) anaerobic fermentation

d) bacterial – algal symbiosis

e) oxygen transfer across the water surface

f) Sulfur bacteria actions

g) evaporation, and

h) seepage


6

Figure 2. Major reactions in a waste stabilization ponds (from Shammas et al., 2009)

1.4.1 Sedimentation

Suspended solids from the influent will be precipitated leading to an enhancement of chemically

and biological flocculation in the pond. From 80 to 90% of suspended solids can be precipitated

depending on temperature, flowrate and depth of the pond. Synthesized bacteria cells and algal

cells are involved in sedimentation process. Moreover, not only BOD is removed, pathogens is

also removed in sedimentation (Amahmid et al., 2002).

1.4.2 Aerobic decomposition

This is one of two major steps in decomposition of organic matter in wastewater. As long as DO

is about the level of 0.1 – 0.2 mg/l, the aerobic decomposition will take place in the pond. At

first, the carbonaceous matter is oxidized by the aerobic bacteria with the formation of carbon

dioxide and other inorganic substances. These inorganic substances are used by algae in their

photosynthesis reactions, and the product of photosynthesis, oxygen, is then used by aerobic

microorganisms.

1.4.3 Anaerobic fermentation

The processes take place in an anaerobic zone of the pond which is under the presence of

fermentative bacterial genera, such as Pseudomonas, Escherichia and Aerobacter. Organic

matter is hydrolyzed into amino acids, long chain fatty acids, and mono and disaccharides. Then,


7

these compounds are going through acidogenesis in which carbonic acids and alcohol are

generated as a result.

The final stage of anaerobic fermentation is methanogenesis and can be illustrated by the

following reactions:

CH3COOH → CH4 + CO2

4H2 + CO2 → CH4 + 2H2O

Gases that are generated during methanogenesis may escape the pond causing bad odor.

1.4.4 Bacterial – algal symbiosis

As mentioned abve, bacteria and algae have a symbiotic relationship when the turbidity of the

pond is low and there is a lot of sunlight. In the upper zone, there are abundant of algae, and

these organisms can oversaturated the zone with molecular oxygen through photosynthesis. The

plentiful of oxygen will support the aerobic oxidation of organic matter by aerobic bacteria. In

return, the aerobic process releases the organic forms of nitrogen and phosphorus for the growth

of algae. In the other hand, the photosynthesis of algae also require large amount of CO2. The

sources of CO2 can be i) as an end product of bacterial oxidation and fermentation, ii) from the

atmosphere and iii) from the inorganic carbon species, such as in the CO2-HCO3--CO3

2- system.

1.4.5 Oxygen transfer across the water surface

Oxygen transfer is followed the following equation: 𝑑𝑐

𝑑𝑡 = Ka(cs – c)

where, c is the DO in the water, cs is the saturation concentration of DO in liquid, and Ka is the

reaeration coefficient.

In daylight, when photosynthesis is at its maximum, oversaturation of oxygen in the pond may

occur leading to escaping of oxygen from the pond. In constrast, when oxygen is below

saturation level, then atmospheric reaeration takes place.

1.4.6 Sulfur bacteria actions

In Figure 2, during aerobic processes, sulfur compounds in wastewater are converted to sulfate,

on the contrary, anaerobic bacteria reduced sulfate to sulfide. The main group of bacteria

reducing sulfide is Desulfovibrio sp. Sulfides can be undergone the following reactions:


8

CO2 + 2H2S→(CH2O) + H2O + 2S

3CO2 + 2S + 5H2O→(CH2O) + 4H+ + 2SO2−

The group of bacteria using CO2 as hydrogen acceptor is the photosynthesis bacteria, mainly

Chlorobacteriaceae and Thirohodaceae.

1.4.7 Evaporation

The loss of water in the pond is only significant in warm and dry climate regions, and depending

on temperature, wind speed, surface area of the pond, vapor pressure of the water, barometric

pressure and salt concentration of water. Ro hwer (1931)

E = 0.497(1 − 1.32 × 10−2

Pa)(1 + 0.268W)(V − v)

where E is evaporation, Pa is barometric pressure in inches of mercury, W is wind speed in

miles/h, and V and v are vapor pressures in inches of mercury at the water temperature and

dewpoint temperature of the atmosphere, respectively.

1.4.8 Seepage

Leaking of wastewater is a problem for the quality of groundwater and the process is depended

on wastewater qualiy and soil characteristics. An infiltration of 122 cm/d was recorded (Davis,

1972). Normally, natural sealing of the pond takes place; otherwise, artifical sealing of the pond

has to be done by using clay, bentoinite or asphalt.

1.5 Physical and chemical factors

1.5.1 Light

For a waste stabilization pond system, light has an important role: it is the main factor for

photosynthesis, algal growth is depent on light intensity; therefore, it affects the dissolved

oxygen and pH of the system. Moreover, light also kills pathogens.

Steele (1962) descriped the functional relationship between light and algal growth by the

following equation: p = apmIet-aI

where, p is the photosynthesis rate; pm is the maximum photosynthesis rate; I is light

intensity; and a is a constant.


9

1.5.2 Dissolved oxygen

Except in anaerobic pond and the bottom of facultative pond, oxygen is needed for aerobic

oxidation of organic matters. In a waste stabilization pond system, oxygen is mostly supplied by

i) mechanical aeration and ii) photosynthesis. Photosynthesis is the most important source of

oxygen in the ponds and can be illustrated by the following equation (Kirk, 1994):

CO2 + 2H2O → CH2O + H2O + O2

Otherwise, algae also comsume oxygen through respiration (Reynolds and Irish, 1997), and

oxygen uptake and carbon dioxide generation as a result. Dissolved oxygen is also a control

factor of odour by consuming H2S gas emitted from anaerobic decomposion process.

H2S + 2O2 𝑏𝑎𝑐𝑡𝑒𝑟𝑖𝑎 H2SO4

1.5.3 pH

pH is play an important role in pathogen removal, nutrient removal and odour control. When the

pH reaches around 9, the destruction of bacterial pathogens occurs (Pearson et al, 1987).

Depending on pH, the sulphide ions may exist in different forms: H2S, HS- and S

2-; only H2S has

a bad odour and exists at pH below 7.5; therefore, pH has an important role in odour control.

H2S ↔ HS- ↔ S

2-

Like other water bodies, pH is controlled by the carbonate bicarbonate buffering system:

CO2 + H2O ↔ H2CO3 ↔ H+ + HCO3

- ↔ H

+ + CO3

2-

As we can see in the equation above, pH changes can lead to changes of CO2, removal of CO2.

The causing of pH changes may be by increasing or decreasing in acids and the addition. The

main source of acids is volatile fatty acids (VFAs) which are formed when organic matter is

broken down through anaerobic processes in the incoming water. Though, increasing loading

rate tends to decrease the pH of waste stabilization ponds, and odour is affected by promoting

H2S formation and because many VFAs are odourous.

1.5.4 Temperature

Temperature has two important roles in waste stabilization ponds: affecting the rate of biological

processes and hydraulic properties of the water when WSP stratify and destratify (Llorens et al.,

1992).


10

As in all biological treatment systems, the effect of temperature on the rate of the process can be

expressed by the modified Arrhenius equation: kT = k20θ(T−20)

where, kT is reaction rate coefficient at temperature T, k20 is reaction rate coefficient at

20oC, and θ is temperature coefficient.

In depth stabilization ponds (8 m in depth), the influence of thermal stratification is a concern

since stratification affected the effectiveness of the system as seen in Llorens et al. (1992). In

addition, temperature also affects the mortality of coliforms and other bacteria.

1.5.5 Salinity

The salinity of a particular wastewater is mainly due to the influent. During the treatment of

wastewater in stabilization ponds, salinity is not decreased; however, evaporation increases the

salinity which affected population of larger organisms (Dillaha and Zolan, 1983).

2 PLANKTON AND WASTE STABILIZATION PONDS

2.1 Introduction

Plankton refers to any form of small biota (from microns to milimeter) living in water and

drifting at the mercy of current (Suthers and Rissik, 2009). However, for assessing water quality,

phytoplankton and zooplankton are the two groups of interest.

Phytoplankton live near the water surface where there is sufficient sunlight and nutrients

(nitrogens and phosphorus) to support photosynthesis. These groups of plankton include diatoms,

cyanobacteria, dinoflagellates and coccolithophores, and are considered as primary producers in

all types of water bodies. The numbers of phytoplankton in the water column reflect the

influence of environmental factors (Webber et al., 2005).

Zooplankton is another group of interest, including protozoas (body size from ten to hundred

micrometer), rotifers (40µm-2,5mm) and crustaceans (100µm-1cm). They feed on some groups

of phytoplankton and are seen as useful biological indicators for water quality, nutrition and

pollution (Jeppesen et al., 2011).

Currently, monitoring plankton in water bodies is increasingly concern due to these reasons:

some phytoplankton can produce toxins which can be concentrated in filter-feeding

animals like oysters, mussels.


11

surplus nitrients in the environment can be passed through trophic cacasde

plankton is a food source of early stages of mussels, oysters, prawn and fish

some species of phytoplankton and zooplankton can be indicators for environmental

quality

2.2 Plankton in waste stabilization ponds

Depending on the type of stabilization ponds, incoming water characteristics, time of the day and

period of the year, the plankton population in the ponds varies significantly (Table 3).

Phytoplankton population can vary differently among ponds and maxium counts can be as 15

million/ml. In most ponds, green algae dominate throughout the year since they can adapt better

to environmetal changes (Shammas et al., 2009). Through a study of algal population in

stabilization ponds, Raschke (1970) has confirmed the following algal is present in stabilization

ponds system:

Table 3. Algae found in wastewater stabilization pond

Division and order Species

Chlorophyta

Volvocales

(Green algae)

Chlaniydomonas supp.

Chlamydomonas celerrirna Pasch.

Chiarnydomonas tremulans Skuja

Chiorogonjum acus Nayal

Chiorogonjum fusiforme Matw.

Eudurina sp.

Ulotrichales

Chlorococcales

(Green flagellated algae)

Stigeolonium sp.

Ankistrodemus convolutes Corda

Chlorella ellipsoidea Gerneck

Chlorella vulgaris Beyernick

Coelastrum sp.

Cruciogenia irregularis Willie

Kirchneriella sp.

Micractinium pusillum Fresenius

Oocystis sp.

Scenedesmus sp.

Euglenophyta Euglena spp.


12

Euglenales

Chrysophyta

Pennales

(Brown or yellow-Green diatoms)

Euglena pisciformis Klebs

Gomphonema parvulum Küts.

Hontzschia sp.

Navicula accomoda Hust.

Navicula cuspidate var. ambigua (Ehr.) Cleve

Navicula excelsa Krasske

Navicula gregaria Donk.

Navicula kriegeri Krasske

Navicula lanceolata (Ag.) Küts.

Nitzschia spp. Nitzschia accomodata Hust.

Nitzschia amphibia Grun.

Nitzschia communia Rabh.

Nitzschia diserta Hust.

Nitzschia fonticola Grun.

Nitzschia lateens Hust.

Nitzschia palea (Kütz.) W. Smith

Nitzschia thermalis Kütz.

Cyanophyta

Oscillatoriales

(Blue-green algae)

Oscillatoria amoena (Kütz.) Gomont

Oscillatoria okeni; Ag. Ex Gomont

Oscillatoria tenuis var. Natans Gomont

Oscillatoria terebriformis Ag.

Source: Raschke (1970)

For a stabilization pond system, four essential groups of zooplankton exist: protozoa, rotifers,

copepods and cladocerans (Pietresanta and Bondon, 1994). Table 4 shows the general ecology of

WSP’s zooplankton.

Table 4. General ecology of WSP’s zooplankton

Characteristic Group

Protozoa Rotifers Copepods Cladocerans

Size 20 - 50 μm 40 - 80 μm 0,5 - 3,5 mm 0,2 - 3 mm

Food Bacteria

Bacteria, protozoa,

algae

Bacteria,

protozoa, algae,

(rotifers and

cladocers for

Bacteria, protozoa,

algae;

Particulate organic

matters


13

high species)

Mode of

reproduction

Sexed

Asexual

Asexual in favorable

conditions; sexed in

adverse

Sexed

Asexual in favorable

conditions; sexed in

adverse conditions

Population Not sexually

differentiated

Ovoviviparous

females

Males in adverse

conditions

Eggs of resistance

(adverse conditions)

Oviparous

females

Males

Eggs of

resistance

(adverse

conditions)

Ovoviviparous females

Males in adverse

conditions

Eggs of resistance

(adverse conditions)

Dynamics None Seasonal Seasonal Seasonal

Localization All ponds

(except in

winter the last

basins)

1st pond; Weak

density in

oligotrophic water,

Strong density in

eutrophic water

Rare presence in

WSP

Especially in the last

basins

Predominent

kind

Not announced Brachionus Cyclopoids Daphniids 75 to 95% of

the biomass of

zooplankton in

eutrophic basins (in

certain cases Moiniids)

Source: Pietresanta and Bondon, 1994

When there is abundant of phytoplankton, protozoa and rotifers can be found significantly.

According to Shammas et al. (2009), the role of these organisms is to control the prey population

and therefore the treatment process.

3 TOXICITY

3.1 Biology of Daphnia magna

Daphnia is a member of Cladocera, whose bodies are enclosed by uncalcified shell, and distri-

bute throughout the water bodies in the world. They are filter-feeders, body sizes range from 1


14

mm to 5 mm. Daphnia feed on small, suspended particles, green algae are among the best food;

therefore, in laboratory, they are feeded with Scenedesmus or Chlamydomonas.

There are many reports about the existence of species and sub-species of Daphnia genus in

wastewater treatment ponds (Dinges, 1973; Dor et al., 1987; Hathaway and Stefan, 1995) and

some of them shown sensitivity to toxic chemicals, organic matter, etc… (Shiny et al., 2005;

Cripp and Kumar, 2003). Moreover, Daphnia species are also used for assessing water quality in

other water bodies (Zeng et al., 2012; Storey et al., 2011); therefore, using Daphnia for assessing

water quality is an essential tool.

Daphnia can reproduce in two ways: through hatching of resting eggs or through asexual mode

of reproduction. The former needs adults of male and female, in this case, only two eggs are

produced and encapsulated in a structure called and ephippium. The ephippia are released into

the water may either sink to the bottom or float to the surface. The latter of reproduction does not

need an adult male, a female produces parthenogenetic eggs after every adult molt.

3.2 Daphnia and toxicity studies

Daphnia magna is an important organism in toxicity studies because of their sensitivity to toxic

compounds in water environment. So far, there are many studies of chronic toxicity

test in Daphnia magna is done in the world. Hassold and Backaus (2009) studied the chronic

effects of five major compounds from demethyclase (an fungicide), including piperazine

triforine, pyrimidine feranimol, pyridine pyrifenox, imidazole prochloraz and triazole

triadimefon on D. magna; the results shown that these compounds reduced fertility of D. magna.

In addition, the authors found that among these five compounds, prochloraz was the most toxic

compounds.

Dao (2010) studied the effect of cyanobacteria on D. magna’s fertility. The results showed that

different concentrations of microcystin effected on the maturation, reproduction and survival of

D. magna. In particular, the results also noted the phenomenon of pregnancy loss in D. magna.

Mahassen and Sami (2011) when studying the chronic effects of waste water

in the different processing stages in the village of El-Mofti, Egypt on D. magna also had the

results that waste water from septic tanks had effects on the survival, growth and reproduction of

D. magna. Also, the toxicity of wastewater reduced after being processed through each stage of

the treatment.


15

III. MATERIAL AND METHODS

1 THE WASTE STABILIZATION POND

1.1 Study area

Binh Hung Hoa wastewater treatment station receives 60-80% of wastewater from Den canal.

Wastewater flows directly to Den canal mainly from domestic wastewater of around 120.000 in-

habitants and a small part from industrial facilities in the area. Figure 3 shows the general plan of

Binh Hung Hoa wastewater treatment station.

Figure 3. General plan of the wastewater treatment plan

1.2 Description of the treatment process

Operating procedures of Binh Hung Hoa wastewater treatment systems are constant and serial

(Figure 4).


16

Figure 4. Diagram of Binh Hung Hoa wastewater treatment plan procedure

1.2.1 Pumping station

Wastewater from Den canal is pumped into the treatment plan through a station which has three

screw pumps (2 operate and 1 standby). Each pump has a capacity of 180 l/s.

1.2.2 Grit chamber

Wastewater from the pumping station flows into two grit chambers to remove settable

components, such as sand, grit, etc… then treated flow moves to splitting chamber and being

distributed to the pond system.

1.2.3 Aerated lagoons

There are two aerated lagoons in the system, each lagoon has eight aerators which are operated

in serial mode. Maximum water depth is 3.2 m. Surface area of each lagoon is 1.5 ha. Retention

time is 4 days. Under strong activity of aerobic microorganisms, organic matter in wastewater is

decomposed.

1.2.4 Sedimentation ponds

There are two sedimentation ponds which are used to settle slugde generated from aerated

lagoons. Maximum water depth is 3 m. Surface area of each pond is 0.94 ha. Retention time is

2.5 days.


17

1.2.5 Maturation ponds

There are six maturation ponds which are operated in parallel lines. Each line contains three

ponds: maturation pond 1, maturation pond 2 and maturation pond 3 with surface area of 2.4 ha,

2.5 ha and 3.3 ha, respectively. Maximum water depth is 2.5 m. Retention time is 8 days.

1.2.6 Sludge drying bed

Sludge that is settled in sedimentation ponds will be pumped periodically into a sludge drying

bed which has a surface area of 7000 m2 and 1 m depth. The sludge will be pumped out twice a

year in dry season and is dried in ten weeks.

2 SAMPLING FOR MONITORING ON ENVIRONMENTAL PARAMETERS AND

PLANKTON

2.1 Physical and chemical parameters

For environmental parameters, a routine monitoring program has been established to determine

the quality of effluent (Smet et al, 2006). Effluent samples from stabilization pond system were

collected 2 times per week on every Monday afternoon and Thursday morning for a period of

study to measure temperature, EC, pH and DO, BOD5 and NH3-N. For temperature, EC, pH and

DO, these parameters were measured directly on site. The samples were brought to the

laboratory and analyzed for BOD5 and NH3-N (these parameters as per the standard procedures

given in Environment Canada (1974) and Standard Methods for the Examination of Water and

Wastewater (APHA, 2005).

2.2 Sampling for phytoplankton and zooplankton

Samples of phytoplankton and zooplankton were taken at water surface during the month of

August (Fig. 3). Diffferent locations in the ponds were also sampled at different times (morning

and afternoon). Qualitative samples of phytoplankton were taken with a conical net of 25 µm

mesh size and quantitative ones were sampled at surface water and fixed with neutral Lugol

iodine solution (Sournia, 1978) in the field. For zooplankton, qualitative samples were taken with

a conical net of 40 µm mesh size and quantitative ones were sampled at surface water and fixed

with 4% formalin for enumeration.


18

2.3 Phytoplankton and zooplankton identification and counting

Phytoplankton was observed at 400 - 800x magnification (Olympus BX51 microscope).

Identification was based on morphology following the system of Komárek and Anagnostidis

(1989, 1999, 2005) for cyanobacteria, Krammer and Lange-Bertalot (1997a, b; 2004a, b) for

diatoms, and other taxonomy books for green algae, golden algae, dinoflagellates and euglenoids

(West and West, 1904; Smith, 1924; Prescott, 1951; Gojdics, 1953; Bourrelly, 1957; Blomqvist

and Olsin, 1981; Yamagishi and Akiyama, 1994a, 1994b, 1995). For counting, the technique is

based on the technique of Utermöhl (1958).

For zooplankton, the samples were also observed under Olympus BX51 microscope.

Zooplankton was identified based on morphology following the system of Shirota (1966), Smith

(2001), Thorp và Covich (2001), Voigt (1956). In quantitative analysis, zooplankton samples

were counted in Petri dishes and Sedgewick-Rafter counting chamber (APHA, 2005).

The numbers of plankton, both phytoplankton and zooplankton, present were calculated as

Shanthala et al. (2009):

Number of plankton ml-1

= Number of organisms counted

Number of replicates

2.4 Diversity index

Two diversity indices were used to explain the diversity of phytoplankton in stabilization ponds,

Shannon – Wiener index and Simpson index. The Shannon – Wiener index is calculated based

on the proportional abundances pi of each species:

H = - 𝑝𝑖 ln(𝑝𝑖)𝑠𝑖=1 where, pi = ni/N where ni = number of individuals of the i

th species and N

= total number of individuals, and S is the total number of species seen in this sample.

To determine the level of pollution in terms of species diversity index, biologists proposed

different scale of pollution (Table 5) (Afli et al., 2009)

Table 5. Criteria of pollution by Shannon – Wiener diversity index

> 4

3 – 4

2 – 3

1 – 2

< 1

Unpolluted

Slightly polluted

Meanly polluted

Heavily polluted

Extremely polluted


19

The Simpson index is another most widely used measure which is calculated from species

proportions:

D = 𝑝𝑖2𝑆

𝑖=1

The Simpson index value represented the level of diversity. According to Shanthala (2009)

diversity criteria using Simpson index diveristy is divided into 2 levels (Table 6).

Table 6. Criteria of diversity by Simpson index

0 – 0.5

0.5 – 1

Lowest possible diversity

Highest possible equal number of different species

2.5 Brachionus:Trichocerca index

The Brachionus:Trichocerca ratio (QB/T) was used to determine the trophic level of the pond

system. As suggested by Sládeček (1983), if the ratio = 1, the lake is oligotrophic, if it is between

1 and 2, it is mesotrophic, and if it is > 2, the lake is eutrophic.

2.6 Evenness

Evenness is an important feature of all ecological communities. The evenness was calculated

based on Peilou (1966):

J = 𝐻

𝑙𝑜𝑔2𝑆

where, H is Shannon-Wiener index, S is total species

3 TOXICITY STUDY

In this study, we test the chronic effects of wastewater on Daphnia magna. For this type of

experiment, samples were taken at the pumping station and at the end of the treatment station

(effluent before discharge to Den canal). Fifteen individuals of Daphnia magna (less than 1 day

old) (originated from Ecotixicology Lab, Institute for Environment and Resources) were

randomly selected and cultured individually for the experiment. Each one was placed in 50 ml

beaker containing 20 ml solution. D. magna were exposed to different concentrations of

wastewater (0, 10, 50 and 100%) and fed with Scenesdemus sp. (1 mg C/l). Solutions were newly

refreshed every 2 day.


20

For life history study, the survival, maturation, and fecundity of Daphnia were observed daily

for two months. Death of the animal was defined as the stop of heartbeat. Maturation of Daphnia

was defined as time point of first egg occurrence in the brood chamber. The time to first

reproduction was at first offspring release from brood chamber during molting. Fecundity of

animals was recorded as the number of clutches and number of offspring per clutch produced by

every mother Daphnia during exposure time. In case of release of decomposed eggs, embryos

or neonates, the offspring number of that clutch was assumed as zero. The experiment was lasted

30 days.

For toxicity assessment, the survival of D. magna in the concentration of 50% in both influent

and effluent were used (EC50) to examine the environmental relevance. The LC50 was converted

to TU (toxicity units) by using the equation:

TU = 100/EC50

Table 7. Wastewater toxicity classification (Persoone et al., 2003)

Toxic Unit (TU) Class Toxicity

TU < 0.4 I Non toxic

0.4 ≤ TU < 1.0 II Low toxic

1.0 ≤ TU < 10 III Toxic

10 ≤ TU < 100 IV Very toxic

TU > 100 V Extremely toxic

4 STATISTICAL ANALYSIS

EC50 was calculated by trimmed Spearman-Karber software version 1.5.

Spearman rank correlation (Systat 13) was applied to examine the relationships between main

groups of phytoplankton, zooplankton and environmental factors.

S-plus software was used for data treatment. Wilcoxon rank sum test was applied to calculate

statistically significant difference of survival rate, maturation and reproduction


21

IV. RESULTS

1 Water quality

1 Physical parameter

In Table 8, the main average water quality parameters were displayed. Surface water temperature

in the ponds is in the range from 28 – 34oC, with lowest temperature is in sedimentation ponds

and highest temperature is in the inlet. pH of the treated ponds during the period of study had

values from neutral to slightly alkaline (7.0 – 8.3), with lowest value is in maturation pond 8 and

highest value is in maturation pond 10. Meanwhile, the electrical conductivity of the water in

these ponds ranged from 582 – 913 µS/cm, with lowest value is in aerated pond and highest

value is in inlet. Dissolved oxygen in these ponds had values from 0.9 – 6.4 mg/l, lowest is in

inlet and highest is in maturation pond 10. For N-NH4+, there is a gradual decrease in

concentration in the ponds system, from 13.1 – 3.6 mg/l.

Table 8. Pond performance as an average of one month’s result.

Parameter Unit W A 1 A 2 S1 S2 Pond 8 Pond 9 Pond 10

Temperature oC 29.5 29.2 29.1 28.9 28.9 29.0 29.2 29.3

EC µS/cm 794.7 738.1 751.1 747.7 727.9 716.9 707.8 683.6

pH

7.5 8.0 7.7 7.6 7.6 7.5 7.6 7.8

DO mg/l 1.4 5.4 5.0 2.4 2.2 4.1 4.6 5.3

BOD mg/l 63 13 13 5 8 8

N-NH4+ mg/l 13.1 13.7 6.3 5.0 4.5 3.6

2 Plankton communities

2.1 Phytoplankton

2.1.1 Phytoplankton composition

Over 60 phytoplankton species were recorded in three maturation ponds during the study (Table

9), belonging to six classes: Cyanophyceae (cyanobacteria), Chlorophyceae (green algae),

Bacillariophyceae (diatoms), Euglenophyceae (euglenoids), Dinophyceae (dinoflagellates) and

Cryptophyceae.


22

Table 9. List of phytoplankton taxa identified in BHH wastewater treatment plant

Cyanobacteria

Merismopedia tenuissima

Gomphosphaeria sp. Pseudanabaena sp.

Oscillatoria sp.

Oscillatoria tenuis Lyngbya sp.

Planktothrix cf. agardhii

Aphanocapsa sp.

Bacillariophyceae Aulacoseira muzzanensis

Cyclotella sp.

Synedra sp. Eunotia sp.

Navicula sp.

Gomphonema sp. Nitzschia cf. palea

Eunotia sp.

Pinnularia sp.

Pinnularia cf. viridis

Chlorophyceae

Pandorina morum

Eudorina elegans Pleodorina sp.

Chlamydomonas sp.

Coelastrum microporum

Pediastrum duplex Pediastrum tetras

Dictyosphaerium pulchellum

Oocystis sp. Nephrocytium sp.

Ankistrodesmus sp.

Kirchneriella lunaris Monoraphidium contortum

Closteriopsis longissima

Selenastrum bibraianum

Scenedesmus acuminatus v. biseratus Scenedesmus arcuatus

Scenedesmus armatus

Scenedesmus armatus v. bicaudatus Scenedesmus dimorphus

Scenedesmus denticulatus

Scenedesmus javanensis

Selenastrum bibraianum

Crucigenia apiculata Crucigenia rectangularis

Tetrastrum heterocanthum

Actinastrum hantzschii Closterium acutum

Scenedesmus longus v. naegelii

Scenedesmus producto-capitatus

Scenedesmus ecornis Scenedesmus quadricauda

Scenedesmus smithii

Scenedesmus spp. Closterium cf. lunula

Closterium cf. moniliferum

Cladophora sp.

Oedogonium sp.

Cryptophyceae Cryptomonas sp.

Euglena acus

Euglena cf. caudata

Euglena ehrenbergii

Euglena spirogyra

Euglena spirogyra v. acuminata

Euglena texta

Euglena oxyuris

Euglena sp.

Lepocinclis acuta

Lepocinclis ovum

Lepocinclis fusiformis

Phacus acuminatus

Phacus pleuronectes

Phacus swirenkoi

Trachelomonas sp.

Strombomonas cf. napiformis v. brevicollis

Dinophyceae

Peridinium sp.

In particular, Chlorophyceae had the highest species (37 species), folowing by Euglenophyceae

(16 species), Bacillariophyceae (9 species), Cyanophyceae (8 species) and two lowest-recorded

species groups (only 1 species): Cryptophyceae and Dinophyceae (Table 10). Among three


23

maturation ponds, pond 8 recorded highest number of species (66 species) and pond 9 had lowest

number of species (61 species).

Table 10. Structure of phytoplankton

Class Pond 8 Pond 9 Pond 10

Cyanobacteria 8 5 5

Bacillariophyceae 9 8 6

Chlorophyceae 37 35 35

Cryptophyceae 1 1 1

Euglenophyceae 10 11 16

Dinophyceae 1 1 1

Total 66 61 64

Figure 5. Phytoplankton succession in taxonomic orders

Figure 5 represents the phytoplankton succession in three ponds. As seen in the results, pond 10

had high number of individuals in most groups (except for Chlorophyaceae and Cryptophyceae).

For Cyanobacteria and Bacillariophyceae, pond 10 had a number of individuals more than 2

times compared to pond 8 and pond 9. The dominant species in Cyanobacteria groups is Meris-

mopedia tenuissima and for Bacillariophyceae is Cyclotella sp. (Figure 6)

020000400006000080000

100000120000140000

Pond 8

Pond 9

Pond 10


24

Figure 6. Microscopic pictures of a) Merismopedia tenuissima and b) Cyclotella sp.

2.1.2 Diversity indices

The species diversity indices and eveness of phytoplankton in three ponds were shown in Table

11. As seen in the results, Shannon – Wiener index was highest in pond 8 and lowest in pond 9;

Simpson index was highest in pond 10; Peilou index was highest in pond 8 and lowest in pond 9.

Table 11. Diversity indices and eveness of phytoplankton in three maturation ponds

Index Pond 8 Pond 9 Pond 10

Shannon – Wiener 4.980 2.075 2.605

Simpson 0.708 0.708 0.856 Peilou 0.824 0.350 0.434

2.2 Zooplankton

2.2.1 Zooplankton composition

During the investigation period, 43 species and 3 larva were identified. Their qualitative and

quantitative composition varied depending on locations. At this period, the Rotifera group was

dominant (27 species), follwed by Arthropoda (5 species), Rhizopoda (5 species), Ciliophora (4

species) and Zoomastigina (1 species) (Table 12). In three maturation ponds, pond 10 recorded

the lowest number of species (34 species and 3 larva), pond 8 had the highest number of species

(39 species and 3 larva) (Table 12).

Table 12. Structure of zooplankton

Class Pond 8 Pond 9 Pond 10

Arthropoda 5 5 5

Rotifera 26 24 21

Rhizopoda 3 4 5

Ciliophora 4 4 2

Zoomastigina 1 1 1

Larva 3 3 3

Total 42 41 37


25

Figure 7 described the shap of Asplanchna amphora and Lecane robusta, two species that have

not been published in Vietnam.

Figure 7. Two zooplankton species firstly recorded in Vietnam. a) Asplanchna amphora and b) Lecane robusta

Figure 8. Zooplankton succession in taxonomic orders

A list of zooplankton species recorded during the period of study is given in Table 13, and Figure

8 represents the zooplankton succession in three ponds. Highest zooplankton density was in pond

0

200

400

600

800

1000

1200

Arthropoda Rotifera Rhizopoda Ciliophora Zoomastigina Lavar

Pond 8

Pond 9

Pond 10


26

10 (1160 individuals/l), lowest zooplankton density was in pond 8 (383 individual/l). The

dominant species recorded in three ponds over the study period were: Brachionus angularis, B.

caudatus, B. calyciflorus, B. urceous, Filinia longiseta and Testudinella elliptica (Figure. 9)

Table 13. A list of zooplankton recorded in three maturation ponds during the study period

ARTHROPODA Moina brachiata (Jurine)

Diaphanosoma sarsi Richard

Leydigia acanthocercoides Fishcer Microcyclops varicans Sars

Physocypria cf. crenulata Sars

ROTIFERA

Philodina cf. cristata Donner Rotaria neptunia Ehrenberg

Testudinella elliptica Gosse

Filinia brachiata Rousselet Filinia longiseta Ehrenberg

Asplanchna amphora Hudson

Anuraeopsis fissa (Gosse) Brachionus angularis Gosse

Brachionus budapestinensis Daday

Brachionus calyciflorus Pallas

Brachionus caudatus Apstein Brachionus falcatus Zacharias

Brachionus forficula forficua Wierzejski

Brachionus plicatilis O.F.Muller Brachionus quadridentatus Hermann

Brachionus urceus (Linnaeus)

Platyllias patulus O.F.Muller Lecane closterocerca Schmarda

Lecane robusta

Lecane curvicornis Murray Lecane elsa Hauer

Mytilina unguiper Lucks

Polyarthra vulgaris Carlin Synchaeta sp.

Trichocerca minuta Olofsson

Trichocerca pussila Jennigns

Trichocerca longirostris Schrank

RHIZOPODA

Arcella vulgaris Ehrenberg

Centropyxis aculeata Ehrenberg Centropyxis ecornis Ehrenberg

Difflugia limnetica

Euglypha alveorata Dujardin

CILIOPHORA

Didinium nasutum

Epistylis plicatilis Ehrenberg

Opisthonecta henneguyi Faur&Fremiet Vorticella campanula Ehrenberg

ZOOMASTIGINA

Tokophrya infusionum (Stein)

LARVA

Chironomidae - Diptera

Nauplius copepoda Nematoda


27

Figure 9. a) Brachionus angularis, b) B. caudatus, c) B. calyciflorus, d) B. urceous, e) Filinia longiseta

and f) Testudinella elliptica

2.2.2. Diversity indices and the Brachionus – Trichocerca quotient

The results of Shannon – Wiener, Simpson and Peilou indices were shown in Table 14. As we

can see, highest value of Shannon – Wiener is in pond 8 and lowest value is in pond 10.

However, there is a contradiction for Simpson index, highest value is in pond 10 and lowest

value is in pond 8. For Peilou index, highest value is in pond 8 and lowest value is in pond 9.

Table 14. Diversity indices and eveness of zooplankton in three maturation ponds

Index Pond 8 Pond 9 Pond 10

Shannon – Wiener 2.423 2.309 1.635

Simpson 0.711 0.765 0.942 Peilou 0.449 0.143 0.314

Table 15 showed the results of Brachionus – Trichocerca quotient, all three ponds were categorized

eutrophic group, and pond 10 had the highest value, since pond 9 had the lowest value of the quotient.

Table 15. Results of Brachionus – Trichocerca quotient in three maturation ponds

Pond 8 Pond 9 Pond 10

Brachionus/Trichocerca 83.3 73 115.6


28

2.3 Correlation between phytoplankton, zooplankton and environmental factors

Table 16 showed the Spearman rank correlation between phytoplankton, zooplankton and

environmental factors. As we concerned about the dominant groups of phytoplankton

(cyanobacteria, bacillariophyceae, chlorophyceae and cryptophyceae) and zooplankton (rotifera) in

three maturation ponds, we only needed to focus on the correlation between these groups and

environmental fators. From the results, cyanobacteria and bacillariophyceae showed negative

correlation with chlorophyceae, cryptophyceae and N-NH4+. However, chlorophyceae and

cryptophyceae correlated negatively with rotifera, temperature, pH and DO.


29

Table 16. Spearman’s correlation between main groups of phytoplankton and zooplankton and water parameters

Cyano-

bacteria

Bacillari-

ophyceae

Chloro-

phyceae

Crypto-

phyceae

Eugleno-

phyceae

Dino-

phyceae

Arthro-

poda

Rotifera Rhizo-

poda

Cilio-

phora

Zoomas-

tigina

Larva Temper-

ature

EC pH DO BOD N-NH4+

Cyanobacteria 1.0

Bacillariophyceae 1.0 1.0

Chlorophyceae -0.5 -0.5 1.0

Cryptophyceae -1.0 -1.0 0.5 1.0

Euglenophyceae 0.5 0.5 -1.0 -0.5 1.0

Dinophyceae 1.0 1.0 -0.5 -1.0 0.5 1.0

Arthropoda 0.5 0.5 0.5 -0.5 -0.5 0.5 1.0

Rotifera 1.0 1.0 -0.5 -1.0 0.5 1.0 0.5 1.0

Rhizopoda 1.0 1.0 -0.5 -1.0 0.5 1.0 0.5 1.0 1.0

Ciliophora -1.0 -1.0 0.5 1.0 -0.5 -1.0 -0.5 -1.0 -1.0 1.0

Zoomastigina -0.866 -0.866 0.866 0.866 -0.866 -0.866 0.0 -0.866 -0.866 0.866 1.0

Larva -0.5 -0.5 1.0 0.5 -1.0 -0.5 0.5 -0.5 -0.5 0.5 0.866 1.0

Temperature 1.0 1.0 -0.5 -1.0 0.5 1.0 0.5 1.0 1.0 -1.0 -0.866 -0.5 1.0

EC -1.0 -1.0 0.5 1.0 -0.5 -1.0 -0.5 -1.0 -1.0 1.0 0.866 0.5 -1.0 1.0

pH 1.0 1.0 -0.5 -1.0 0.5 1.0 0.5 1.0 1.0 -1.0 -0.866 -0.5 1.0 -1.0 1.0

DO 1.0 1.0 -0.5 -1.0 0.5 1.0 0.5 1.0 1.0 -1.0 -0.866 -0.5 1.0 -1.0 1.0 1.0

BOD 0.5 0.5 0.5 -0.5 -0.5 0.5 1.0 0.5 0.5 -0.5 0.0 0.5 0.5 -0.5 0.5 0.5 1.0

N-NH4+ -1.0 -1.0 0.5 1.0 -0.5 -1.0 -0 -1.0 -1.0 1.0 0.866 0.5 -1.0 1.0 -1.0 -1.0 -0.5 1.0


30

3 Toxicity test

3.1 Survival rate

After 30 days of experiment, the survival rate of Daphnia magna in the control was 100%. For

influent, at the concentration of 10%, the survival of D. magna decreased slightly after 17 days

of exposure and continued to decline in the following days. At concentration of 50%, the survial

rate of D. magna decreased after 3 days of exposure and the following days. At 100% of influent,

the survival rate decreased after 1 day of exposure and continued to decline in the next days of

exposure. In addition, after 30 days of chronic exposure, the results showed survival rate of 53%,

29% and 0% for the influent concentrations of 10%, 50% and 100%, respectively (Figure 10a).

When using ascute toxicity test, LC50 of influent was 13%. For influent, the survival rate

decreased after 15 days of exposure in concentrations of 10 and 50%. At 100% effluent, number

of D. magna decreased slightly after 22 days of exposure. In addtion, after 30 days of

experiment, the survival of studied organisms in all three concentrations was 93% (Fig. 10b)

Figure 10. Survival rate of D. magna: a) influent, b) effluent


31

3.2 Maturation

At 20 ° C, the maturation of the organism Daphnia magna in the control occurred at day 5 or day

6 of experiment, an average of 5 ± 0.5 days. For influent, at a concentration of 10%, 13

organisms matured on day 4 or 5, the average of 4 ± 0.5 days. Maturation time of D. magna in

concentration of 50% fluctuated from day 4 to day 8, average of 4 ± 0.16 days. However, the

maturation time of D. magna at concentration of 100% was slower than the control, Daphnia

matured from day 7 to day 9, average 8 ± 0.7 days (Figure 11).

Figure 11. Maturation age of the Daphnia magna (days ± standard deviation).. n = 15: number of

experimental animals.

For the effluent, in concentration of 10%, 10 organisms matured from day 4 to day 6 of the

experiment, an average of 5 ± 0.5 days. At concentration of 50%, 13 D. magna matured on day 4

or 5 of the experiment, the average was 4 ± 0.7 days. Maturation of D. magna at 100% occurred

early, 12 mature animals simultaneously on day 4 of the experiment (Fig. 11).

3.3 Reproduction

After 30 days of experiment, Daphnia magna in the control reproduced 7 – 8 times, an average

11 ± 5 offsprings/clutch, a total of 1226 offsprings. During exposure to influent, at a

concentration of 10%, D. magna reproduced 5 – 11 times, and average 12 ± 7 offprings/clutch, a

total of 1561 offsprings. At 50% influent concentration, D. magna reproduced 4 – 11 times, an

average 23 ± 13 offsprings/clutch, a total of 1558 offsprings. The reproduction of D. magna at


32

100% of influent was low, reproduced only 2 – 8 times, and average 20 ± 12 offsprings/clutch, a

total of 678 offsprings (Figure 12a).

Figure 12. Number of D. magna offsprings born. a) influent, b) effluent

During the experiment of exposure to effluent, at effluent concentration of 10%, D. magna

reproduced 3 – 8 times, average 12 ± 7 offsprings/clutch, total 760 offsprings. Reproduction in

D. magna increased in effluent concentration of 50%, reproduced 7 – 9 times, averaging 19 ± 8

offsprings/clutch, a total of 1943 offsprings. At 100% effluent, the reproduction of the D. magna

continued to rise, from 5 – 9 times, an average of 23 ± 13 offsprings/clutch, a total of 2208

offsprings (Figure 12b).

Reproduction helps organisms survive over time. In small animals, they often reproduce in large

numbers to adapt to harsh environment. When comparing to control, the reproduction of

organisms in all concentrations influent did not have any significant difference (Wilcoxon rank

sum test, p < 0.05). Reproductive rates of D. magna were higher in effluent concentrations of 50

and 100%, and decreased at concentrations of 10% compared to the control. However, there was

no significant difference when comparing control to all the concentrations of effluent (Wilcoxon

rank sum test, p < 0.05). In general, the reproduction in Daphnia magna depends on the degree

of wastewater toxicity.

3.4 Fecundity and malformation

The phenomenon that eggs/embryos were destroyed in Daphnia magna mother (the the abortion)

were also recorded in the concentration of 10, 50, 100% of influent and 100% of effluent. The

abortion rates at concentration of 10 and 50% of influent were higher than those of concentration


33

of 100% of inluent (3 individuals, 20%) and 100% of effluent (1 individual, 7%) (Table 17).

Particularly, during the exposure, the results also showed the malformation in offspring, such as

incomplete development of large antennae (for swimming) and un-rejected tail spine. The

malformation of offsprings was also recorded in effluent, especially at concentration of 50% and

100% (3 individuals, 20%) (Table 16).

Table 17. The number of individual Daphnia magna mother miscarriage and birth defects

Number of D. magna mother

Abortion Malformation in offspring Control 0 0

Influent 10% 3 0

50% 3 0

100% 1 2 Effluent 10% 0 1

50% 0 3

100% 1 3


34

V. DISCUSSION

1 Water quality

1 Physical and chemical parameters

Temperature has been concerned as the most important factor in biological wastewater treatment

since it affects the metabolic rate of microorganisms in the system, and thus the degradation of

organic matter (Gray, 1992). In addtion, temperature also afffects on the development of

phytoplankton and zooplankton. According to Reynolds (1984), optimum temperature for most

trains of planktic algae and cyanobacteria is in the range 25 – 35oC, this range is also the same

for zooplankton (Heinle 1969). The temperature results in BHH wastewater treatment system is

suitable for the growth of phytoplankton and zooplankton.

Electrical conductivity is dominated by the ions and valences of the compounds in water. EC

value depends on surface water temperature. Most of dissolved inorganic compounds in water

are good conductor. In contrast, organic molecules are not or hardly conductivity. In freshwater,

EC is usually range from 50 – 1500 µS/cm (APHA, 2005). Comparing to Tri An resevoir, the

dissolved ions in the effluent is 12 times higher (Dao, 2010). There was a general decrease of EC

through the treatment process (from 794.7 to 683.6) which can be accounted for the effectiveness

of the system.

Generally, pH values of ponds ranged in the optimum for microbial degradation (Parawira,

2004). In addition, pH values was also inside the optimum range for futher treatment of

wastewater (removal of COD and nutrients) (Ramadan and Pounce, 2004). However, due to

photosynthetic activity in maturation ponds, there is a slightly change in pH and DO values in

these ponds.

Dissolved oxygen is essential for living organisms, especially aquatic organisms. In water,

oxygen is mainly come from the air and from photosynthesis of phytoplankton. Due to high

nutrient in inlet, DO values is very low (from 0.9 to 2.0). Respiration of organisms (especially

microorganisms) also contributed to the low DO, this maybe a reason for low DO in

sedimentation ponds (1.6 – 3.5). For maturation ponds, the DO was in the range of 3.0 to 6.4,

which was suitable for the development of organisms (Boyd et al., 1978). For aerated ponds, DO

depends on the rate of aeration, which is range from 3.4 to 8.0.


35

Results of BOD indicate that BOD was within the acceptable limit of Vietnam for discharged

wastewater. However, there was an increase of BOD after going through maturation ponds due

to the development of algae (Hogan, 2001). BOD of inlet was expected high due to high organic

contents.

There was a decrease of NH4+ levels after going through the ponds. This may be due to the

nitrification-denitrification processes (Ramadan and Pounce, 2004) and to the nitrogen

assimilation and utilization of phytoplankton for their development (Padisak, 2003). The results

shown that NH4+ after the treatment process was within permissible limits with regard to

Vietnamese standards.

In summary, during the monitoring in the treatment plan, all concerned physical and chemical

parameters were in the standard limits of Vietnam. However, futher studies on light intensity,

stratification of the ponds (mainly maturation ponds), trace elements and nutrient cycle in the

water body are suggested for more understanding on the dynamics of the treatment process.

2 Plankton communities

2.1 Phytoplankton

2.1.1 Phytoplankton composition

The phytoplankton assemblage in BHH wastewater treatment ponds consisted of most major

taxonomic groups of freshwater algae such as green algae, golden algae, diatoms, dinoflagellates,

euglenoids and cyanobacteria (Fig. 5). This record was similar to that in a previous investigation

of phytoplankton in tropical ponds from Malaysia (Yusoff and McNabb, 1997), Ecuador

(Janssens, 2010). Green algae had highest species richness followed by euglenoids, diatoms, and

cyanobacteria; whereas dinoflagellates and golden brown algae contributed smallest species

number of all (Table 9) . This could be explained as green algae have higher competition

capacity to phosphorus, nitrate and light intensity (Horne and Goldman, 1994).

Species diversity and plankton succession in ponds is dependent on organic load, temperature,

day length, pH, and grazing intensity of zooplankton (Neel and Hopkins, 1956; Boutin et al.,

1988). There is a succession of dominant algal species through the ponds, only a few species will

be dominant. The most commonly recorded genera are: Scenedesmus and Euglena. These two

are also common on literature concerning WPS (Mara and Pearson, 1998). The most abundant


36

recorded species are: Cyclotella, Merismopedia, Scenedesmus, Cryptomonas and Euglena.

During the study, a potential harmful phytoplankton was found: Oscillatoria tenius (Fristachi

and Sinclair, 2008). This species was only found in pond 8 (qualitative data). In general, diatoms

had the highest number of individuals, followed by green algae, cyanobacteria, golden algae,

euglenoids and dinoflagellates; this is not in line with Janssens (2010) where euglenoids was the

highest and cyanobacteria was the lowest.

As phototrophs, algae produce oxygen, creating aerobic condition for the ponds. In addition, al-

gae also use nutrients in water for their growth and development; as a result, nutrients are re-

duced. Through photosynthesis, pH of water changes leading to precipitating of nutrients. These

findings corresponded with physico-chemical data.

Since the biomass of cyanobacteria and diatoms in pond 10 had two times higher than in pond 8

and pond 9 (Appendix 2). As a result, there was surface scum in pond 10 when the samples were

taken. The dominant species in Cyanobacteria groups is Merismopedia tenuissima and for Bacil-

lariophyceae is Cyclotella sp. (Figure 6). The explanation for the dominant of diatoms is as

phosphorus is limiting (Tilman et al, 1986). Moreover, during the study period, highest number

of phytoplankton was recorded in pond 10 (248.971 individuals/l) and lowest number of phytop-

lankton was in pond 8 (115.711 individuals/l).

2.1.2 Diversity indices

Diversity indices serve as important tools for various algae (Shanthala et al., 2009). There is neg-

ative correlation between species diversity and pollution of water. The distribution of species

concurs to the type of pollution. Species tolerate pollution can survive in the water. The more

diversity of organisms in the pond, the more significant role of purification. Species diversity is

used to indicated a more complex and healthier community since greater variety of species al-

lows more interaction among species; therefore, attaining greater stability and indicates good en-

vironmental conditions.

Based on Shannon – Wiener index, pond 8 had highest quality of water (unpolluted), this was

constrast to the results of physico-chemical data. Stiling (1996) suggested that the Shannon

Wiener value increase as the abundant of all species in community tend to be the same. In

polluted water, Shannon – Wiener index depends not only on the number of species present, but

also depends on the number of individuals of each species present. Besides, in some cases,


37

Shannon – Wiener value can give a result that is constrast to nutritional status of water bodies

(Hellawell, 1986). Therefore, a combination of Shannon – Wiener index with other indices like

Pielou index is necessary to assess water quality. Based on Pielou index, pond 8 had an possible

equal diversity of species compared to pond 9 and pond 10; therefore, pond 8 had such a high

value of Shannon – Wiener index. In addition, this pond also had the highest number of species.

From results of three indices, the waste stabilization ponds showed moderate level of pollution.

2.2 Zooplankton

2.2.1 Zooplankton composition

Zooplankton are also useful indicators for water quality. The species composition and abundance

shows changes in water quality as zooplankton vary depending on physico-chemical variables

and phytoplankton (Raymont, 1980). Since then, many studies have been investigating the effect

of environmental stresses on the response of zooplankton and their use as biological indicators

(Casé et al., 2008; Jeppesen, 2011).

The rotifer species recorded mainly belonged to Brachionidae, Lecanidae and Trichocercidae

families. They are good indicators for nutrient-rich environment, such as Brachionus

calyciflorus, they live in shalow water, organic polluted environment and highly resistant to

temperature (Gannon and Stemberger, 1978); B. angularis is an indicator for nutrient-rich

environment, well-developed in an alkaline environment and present throughout the year

(Sládeček, 1983).

Five species of Rhizopoda group were recorded as highly adapted to bottom environment,

including Arcella vulgaris, Centropyxis aculeate, Centropyxis ecornis, Difflugia limnetica and

Euglypha alveorata. Species in Ciliophora group, including Didinium nasutum, Epistylis plicati-

lis, Opisthonecta henneguyi and Vorticella campanula live in organic-rich environment (Thorp

and Covich, 2001). In addition, the results showed low distribution and species composition in

Arthropoda and Larva.

Generally, the distribution of zooplankton in BHH wastewater treatment plant is similar to a

study in industrial wastewater in Birla Nagar (India) (Mishra and Saksena, 1990). The results is

also the same compared to studies on zooplankton communities in tropical and sub-tropical

region (Magged and Heikal, 2006; Yildiz et al., 2007), as well as findings of Shirota (1966) on

zooplankton in freshwater in South Vietnam. Moreover, in this study, there are two species that


38

have not been published in Vietnam, namely Asplanchna amphora and Lecane robusta (Figure

7).

As was showed in Figure 8, in all three ponds, Rotifera group is the dominant group due to their

rapid adaptation to environmental changes and their reproductive cycle. In addition, the Rotifera

is also seen as good indicator organisms for water quality and nutritional status due to their short

generation period and rapid population replacement (Sládeček, 1983). Moreover, The dominant

species recorded in three ponds (also in Rotifera group) over the study period were: Brachionus

angularis, B. caudatus, B. calyciflorus, B. urceous, Filinia longiseta and Testudinella elliptica

(Figure 9). These species are indicators for nutrient-rich and organic polluted environment

(Sládeček, 1983).

Highest zooplankton density was in pond 10 (1160 individuals/l), lowest zooplankton density

was in pond 8 (383 individual/l). This result was also correspond with phytoplankton result as

the fact that phytoplankton is a food source for zooplankton; therefore, higher number of

phytoplankton in the pond can support more zooplankton.

2.2.2. Diversity indices and the Brachionus – Trichocerca quotient

According to the result from diversity indices, based on Afli et al. (2009), water quality in pond 8

and in pond 9 are moderate polluted; however, the polluted level in pond 10 is very high. These

results were contradict to those of physico-chemical analysis; however, this finding was similar

to that of phytoplankton. Again, we need to combine different indices to objectively evaluate the

results. By combining three indices, the water qualities in three ponds were in moderate level of

pollution.

In this study, there were twenty-seven species of rotifers were enumerated, which include nine

species of Brachionus, four species of Lecane, three species of Trichocerca, two species of

Filinia and one species each of Anuraeopsis, Asplanchna, Mytilina, Philodina, Platyllias,

Polyarthra, Rotaria, Synchaeta and Testudinella. The maximum number of rotifer were recorded

in pond 10 (1126 individuals/l). Based on the Brachionus – Trichocerca quotient, three

maturation ponds could be categorized eutrophy group (Table 15) since Brachionus is related to

eutrophic water (Blancher, 1984; Gannon and Stemberger, 1978, Sharma, 1983; Pejler,

1983) and Trichocerca are indicator species of oligotrophic water (Sládeček, 1983).


39

A look at Rotifera group, Brachionus was dominant in all three ponds. Amongst the rotifer

population, Brachionus angularis was number dominant over other species. This finding was

correspond with other studies (Pandey et al., 1992; Williams, 1966) that more than one genus

show only one dominant species. The abundance of rotifers is used as an indication of

eutrophication (George, 1966). The abundance of rotifer depends on the level of dissolved

oxygen, the results of zooplankton analysis was also supported this. According to Sládeček

(1983), B. angularis and B. calyciflorus are indicators for mesosaprobic condition, and having a

strong affinity to strong alkaline water. However, the study did not support this since pH values

of the three maturation ponds were still in the range of neutral. Filinia longiseta is recorded as

oligosaprobic or mesosaprobic preferred species. This species was present in all three maturation

ponds.

2.3 Correlation between phytoplankton, zooplankton and environmental factors

As we can see, cyanobacteria and diatoms positively correlated with temperature, this was in line

with finding of Ahmadi et al. (2005). The positive correlation between DO, pH, and

cyanobacteria and diatoms was a result of photosynthetic activity especially at favorable

temperature as mention above. BOD also showed a positive correlation with cyanobacteria and

diatom, since there was high biomass of these two groups in the ponds which furthermore

released organic compounds leading to be decomposed by bacteria.

In this study, both cyanobacteria and diatoms negatively correlated with inorganic nitrogen (in

form of ammonium), this may due to the high dissolved inorganic nitrogen concentration (> 100

µM) in the ponds and this may explain why the non-nitrogen-fixing colonial genera,

Merismopedia was dominating. In addition, Cyclotella is capable of nitrogen fixation, but grow

more rapidly when inorganic nitrogen source was supplied (Pahl et al., 2012).

In constrast to cyanobacteria and diatoms, green algae and golden brown algae negatively

correlated with temperature, pH and DO; however, possitively correlated with N-NH4+. This

maybe due to the fact that these two groups are autotrophic so they are more or less affected by

environmental factors.

Rotifers showed possitive correlation with cyanobacteria and diatoms; in constrast to negative

correlation with green algae and golden brown algae. This is due to the fact that cyanobacteria is


40

not an adequate food source for zooplankton (Lürling, 2003). However, in this study, a possitive

correlation between diatoms and rotifers is a significant remark since Cyclotella is also a

preferable food source for Brachionus (Pagano, 2008) (Cyclotella and Brachionus were the most

abundant species in diatoms and rotifers in this study, respectively). Since ammonium is a strong

candidate toxicant in Brachionus (Isidori et al., 2003), this is not surprise that there is a negative

correlation between ammonium and rotifers.

3 Toxicity test

3.1 Survival rate

The survability in the control after 30 days of experiment was 100%, this is reasonable since

media and conditions in control experiment are the same as those when D. magna are cultured in

the laboratory. As expected, differences between different concentrations of influent and effluent

occurred. The survival rate of D. magna changed between concentrations and decreased when

organisms were exposed to higher concentration; compared to the control, all concentrations of

wastewater (both influent and effluent) showed significant differences (Wilcoxon rank sum test,

p < 0.05). Based on EC50 value, the influent was classified in Class III according to Persoone et

al. (2003) (Table 7).

The effect of wastewater on D. magna showed that there were differences in survival rate in

influent and effluent. The influent had strong effect on the viability of D. magna, especially at

concentrations of 50% and 100%. In the effluent, the concentration that caused effect increased.

This showed that the acute toxicity in wastewater was significantly reduced after treatment,

effluent quality had been improved.

3.2 Maturation

In chronic experiments, all organisms in the control were maturated. However, for influent, at a

concentration of 10%, the percentage of matured organisms reached 87%. At concentrations of

50 and 100%, the rate of maturation of the organism decreased strongly, and reached 53%.

However, in the effluent, the rate of maturation of D. magna at concentrations of 10% was 67%.

At 50% effluent concentration, the percentage of matured organisms increased, reached 89%. At

100% effluent, the maturation rate decreased to 80%. The maturation rates of D. magna in

influent and effluent at a concentration of 10, 50 and 100% are significantly different, compared


41

to the control. In particular, the rate of unsuccessful maturation ones after 6 days in influent is

lower than the effluent.

In addition, the results recorded the differences in organisms that could not mature in influent

and effluent. The organisms exposed to influent that could not mature, died before the end of

chronic experiment (30 days). The organisms that were immature died earlier in higher

concentrations. In particular, at a concentration of 10%, 2 immature animals died on day 25 of

the experiment. At concentrations of 50%, 7 immature animals died on the 4th day of the

experiment. At concentration of 100%, 7 immature animals died from day 2 to day 5 of the

experiment. In the effluent, the rate of immaturation was high at low concentration, particularly

at a concentration of 10%, 5 animals did not mature, the rate of immaturation was 33%. At

concentration of 50%, two organisms were immature and at concentration of 100%, 3 organism

could not mature. The animals that could not mature in the effluent were able to survive to day

30 of chronic experiments.

The development of D. magna occurs after each molt (APHA, 2005). The toxic compounds in

wastewater can affect biological mechanisms of living organisms by hindering molting, delaying

growth and development, or increasing the growth and development which is faster than normal

levels (Hassold and Backhaus, 2009). This suggests that the maturability of organisms depends

on the concentration of wastewater: when concentrations of wastewater are insufficient to cause

inactivation in organisms will impact on the mechanisms that control molting leading to

abnormal development. Therefore, at the concentrations of influent and effluent, the maturation

occurs earlier or slower, compared to the control. In addtion, the body size of D. magna at

concentration of 100% is larger than that of in concentration of 10 and 50% (by observation).

This may be a key factor promoting the early maturation in the organism in response to the

environment (Hassold and Backhaus, 2009).

3.3 Reproduction

During the experiment of exposure to effluent, at effluent concentration of 10%, D. magna

reproduced 3 – 8 times, average 12 ± 7 offsprings/clutch, total 760 offsprings. Reproduction in

D. magna increased in effluent concentration of 50%, reproduced 7 – 9 times, averaging 19 ± 8

offsprings/clutch, a total of 1943 offsprings. At 100% effluent, the reproduction of the D. magna


42

continued to rise, from 5 – 9 times, an average of 23 ± 13 offsprings/clutch, a total of 2208

offsprings (Figure 12b).

Reproduction helps organisms survive over time. In small animals, they often reproduce in large

numbers to adapt to harsh environment. When comparing to control, the reproduction of

organisms in all concentrations influent did not have any significant difference (Wilcoxon rank

sum test, p < 0.05). Reproductive rates of D. magna were higher in effluent concentrations of 50

and 100%, and decreased at concentrations of 10% compared to the control. However, there was

no significant difference when comparing control to all the concentrations of effluent (Wilcoxon

rank sum test, p < 0.05). In general, the reproduction in Daphnia magna depends on the degree

of wastewater toxicity.

3.4 Fecundity and malformation

The decrease of offspring happened via two possibilies: a) mother aborted their eggs/embryos; b)

dead neonates in brood chamber or malformed neonates, incomplete development of antennae

and un-rejected tail spine, which usually died some hours after birth. This phenomenon was also

observed in D. magna which were exposed to cyanobacterial toxins (Dao, 2010). These two

possibilities were recorded when D. magna were exposed to different concentrations of influent

and effluent. In particular, the abortion occured at higher rate in influent concentrations of 10%

and 50%, the concentrations causing early maturation. The malformation in offsprings occured

more in effluent concentrations of 50% and 100%, the concentrations that caused non-lethal

effect on organisms, but affected on the fecundity of the organisms. This suggested that toxicity

was not only in influent but also in effluent. The potential source of pollution may be due to the

small-scaled industries around the canals; however, in the future, the problems with these

industries can be solved if governmental regulations are taken seriously.


43

VI. CONCLUSIONS AND PERSPECTIVES

6.1 Conclusions

In Binh Hung Hoa waste stabilization ponds, basic environmental parameters, phytoplankton and

zooplankton occurence were monitored in August 2011. The water temperature and pH were

almost stable, while DO, EC, BOD and N-NH4+ were different among ponds. Benifiting the

environmental conditions, major group of phytoplankton such as green algae, diatoms,

euglenoids, dinoflagellates and cyanobacteria coexisted in all three maturation ponds. In these

groups of phytoplankton, Chlorophyceae was the most abundant group. During the monitoring

period, seventy-two algal species were recorded.

For zooplankton analysis, 43 species and 3 larva were identified. The most abundant group was

rotifer, followed by Arthropoda, Rhizopoda, Ciliophora and Zoomastigina. The density

fluctuations of zooplankton in three maturation ponds was mainly by the change in population of

Rotifera. During the monitoring period, among twenty seven species of Rotifera, two species

were described the first time for Vietnam (Asplanchna amphora and Lecane robusta).

Based on Shannon – Wiener, Simpson and Pielou indices, the level of pollution in the pond

system was moderate. However, by using Brachionus – Trichocerca quotient, all three

maturation ponds showed eutrophic status.

Chronically exposed to influent showed dramatic influences on survival rates, number of

neonates, days of maturation and fecundity of D. magna mother and malformation of neonates

compared to chronically exposed to effluent.

6.2 Perspectives

Besides the monitored parameters, other factors such as light intensity, stratification of the ponds

(mainly maturation ponds), trace elements and nutrient cycle in the water body would also

influence the phytoplankton and zooplankton development. Therefore, futher studies on these

factors are suggested for more understanding on the dynamics of the treatment process as well as

correlation between phytoplankton, zooplankton and environmental factors.

Since there was sight of toxic cyanobacteria in the pond system, an investigation needs to be

carried on as surface water is used for drinking water supply which cyanotoxins could not be

completely removed during purification process in Vietnam.


44

VII. REFERENCES

Afli A., Boufahja F., Sadraoui S., Benmustapha K., Aïssa P. And Mrabet R. (2009) Functional

organization of the benthic macrofauna in the Bizerte lagoon (SW Mediterranean Sea), semi-

enclosed area subject to strong environmental/anthropogenic variations. Cahiers de Biologie

Marine. 50: 105 – 117.

Ahmadi A., Riahi H. and Noori M. (2005) Studies of the effects of environmental factors on the

seasonal change of phytoplankton population in municipal waste water stabilization ponds.

Toxicological & Environmental Chemistry. 87: 543–550.

APHA (2005) Standard Methods For The Examination Of Water And Wastewater, 21st Edition,

American Public Health Association, American Water Works Association and Water

Environment Federation, Washington, pp. 101-104.

Amahmid O., Asmama S., and Bouhoum K. (2002) Urban wastewater treatment in stabilization

ponds: occurrence and removal of pathogens. Urban water. 4: 255-262.

Berger W.H. and Parker F.L. (1970) Diversity of planktonic Foraminifera in deep sea sediments.

Science. 168: 1345-1347.

Blancher E.C. (1984) Zooplankton-trophic state realitionships in some North and central Florida

Lakes. Hydrobiologia. 109: 251-263.

Blomqvist P. and Olsin P. (1981) Växtplankton kompendium. Uppsala, 186p.

Botero L., Montiel M., Estrada P., Villalobos M. and Herrera L. (1997) Microorganism removal

in wastewater stabilisation ponds in Maracaibo, Venezuela. WATER SCIENCE AND

TECHNOLOGY. 35: 205-209.

Bourrelly P. (1970) Les algues d’eau douce - Tom III: Les algues bleues et rouges - Les

Eugléniens, Peridiniens et Cryptomonadines. Boubée et Cie, Paris, 512p.

Boutin P., Racault Y., and Douat J. (1988) Seasonal variation in concentration and flux of pollu-

tants in stabilization pond: Relations between parameters. Tribuna CBEDEAU. 41: 13–25, 1988.

Boyd C.E., Romaire R.P. and Johnson E. (1978) Predicting early morning dissolved oxygen

concentration channel catfish ponds. Transactions of the American Fisheries Society. 107: 484-

492.


45

Casé M., Leça E.E., Leitão S.N., Sant’Anna E.E., Schwamborn E. and de Moraes Junior A.T.

(2008) Plankton community as an indicator of water quality in tropical shrimp culture ponds.

Marine Pollution Bulletin. 56: 1343–1352.

Cripp S. and Kumar M. (2003) Environmental and other impacts of aquaculture. In: Lucas J.S.

and Southgate P.C. (eds) Aquaculture: farming aquatic animals and plants. Fishing New Books,

Oxford, pp: 74-99.

Dao T.S. (2010) Toxicity of cyanobacteria and cyanobacterial compounds from Tri An

Reservoir, Viet Nam, to Daphnids. Doctor of Philosophy thesis, Humboldt University.

Davis S. (1972) Dairy Waste Ponds Effectively Self-Scaling. Annual Meeting of the American

Society of Agricultural and Biological Engineers. Paper No. 72–222.

Dillaha T.A. and Zolan W.J. (1983) The effects of increased salinity levels on the reaction rates

of biological wastewater treatment. Water and Energy Research Institute of the Western Pacific,

Technical Report No. 37.

Dinges R. (1973) Ecology of Daphnia in Stabilization Ponds. Texas State Department of Health,

Division of Waste Water Technology and Surveillance, Austin, 155 pp.

Dor I., Schechter H., and Bromley H.J. (1987) Limnology of a hypertrophic reservoir storing

waste water effluent for agriculture at Kibbutz Na’an, Israel. Hydrobiologia. 150: 225– 241.

Environment Canada (1974). http://www.weatheroffice.gc.ca/canada_e.html.

Federal Water Quality Administration Municipal Waste Facilities in the United States, No.CWT

– 6. FWQA, Washington, DC (1970).

Fristachi A. and Sinclair J.L. (2008) Occurrence of Cyanobacterial Harmful Algal Blooms

Workgroup Report. In: Hudnell H.K. (Ed) Cyanobacterial Harmful Algal Blooms: State of the

Science and Research Needs. Springer Science + Business Media, LLC. P45 – 104.

Gannon J.E. and Stemberger R. (1978) Zooplankton (especially crustaceans and rotifers) as

indicators of water quality. Transactions of the American Microscopical Society. 97: 16-35.

George M.C. (1966) Comparative plankton ecology of five fish tank ponds. Hydrobiologia. 32:

47-60.

Gojdics M. (1953) The genus Euglena. University of Wisconsin Press, 268p.


46

Gray N.F. (1992) Biology of Wastewater Treatment. Oxford University Press: Oxford. 828p.

Hassold E. and Backhaus T. (2009) Chronic toxicity of five structurally diverse demethylase-

inhibiting Fungicides to the Crustacean Daphnia magna: A comparative assessement.

Environmental Toxicology and Chemistry. 28: 1218-1226.

Hathaway C. and Stefan H. (1995) Model of Daphnia populations for wastewater stabilization

ponds. Water Research. 29: 195–208.

Heinle D.R. (1969) Temperature and zooplankton. Chesapeake Science. 10: 186-209.

Hellawell M.J. (1986) Pollution monitoring series, biological indicators of freshwater pollution

and environmental management, Elsevier, New York. 546p.

Hodgson I.O.A. (2000) Treatment of domestic sewage at Akuse (Ghana). WATER SA. 26: 413-

415.

Hogan N. (2001) Extensive water-based post-treatment systems for anaerobically pre-treated

sewage. In: Lens P., Lettinga G. and Zeeman G. (Eds) Decentralised sanitation and reuse.

Concepts, systems and implementation. IWA Publishing, 650p.

Horne A.J. and Goldman C.R. (1994) Limnology (second edition). McGraw-Hill, Inc. pp. 111-

139.

Isidori M., Lavorgna M., Nardelli A., and Parrella A. (2003) Toxicity identification evaluation of

leachates from municipal solid waste landfills: a multispecies approach. Chemosphere. 52: 85-

94.

Janssens N. (2010) Characterization of algae diversity and kinetics in waste stabilization ponds.

Master thesis, Ghent University.

Jeppesen E., Nõges P., Davidson T.A., Erik Haberman J., Nõges T., Blank K., Lauridsen T.L.,

Søndergaard M., Sayer C., Laugaste R., Johansson L.S., Bjerring R., and Amsinck S.L. (2011)

Zooplankton as indicators in lakes: a scientific-based plea for including zooplankton in the

ecological quality assessment of lakes according to the European Water Framework Directive

(WFD). Hydrobiologia. 676:279–297.

Kirk J.T.O. (1994) Light and photosynthesis in Aquatic Ecosystems, 2nd

edition. Cambridge

University Press, Cambridge, 530pp.


47

Komárek J. and Anagnostidis K. (1989) Modern approach to the classification system of

Cyanophytes 4 - Nostocales. Arch. Algological Studies/Archiv für Hydrobiologie, Supplement

Volumes 56: 247-345.

Komárek J. and Anagnostidis K. (1999) Cyanoprokaryota 1. Teil: Chroococcales. In: Büdel B.,

Gärtner G., Krienitz L. and Schagerl M. (Eds) Süβwasserflora von Mitteleuropa. 19/1. Gustav

Fischer Verlag Jena. 548p.

Komárek J. and Anagnostidis K. (2005) Cyanoprokaryota 1. Teil: Oscillatoriales. In: Büdel B.,

Gärtner G., Krienitz L. and Schagerl M. (Eds) Süβwasserflora von Mitteleuropa. 19/2. Gustav

Fischer Verlag Jena. 759p.

Krammer K. and Lange-Bertalot H. (1997a) Süβwasserflora von Mitteleuropa, Bacillariophyceae

1. Teil: Naviculaceae. Gustav Fischer Verlag Jena. 876 p.

Krammer K. and Lange-Bertalot H. (1997b) Süβwasserflora von Mitteleuropa, Bacillariophyceae

2. Teil: Bacillariaceae, Epithemiaceae, Surirellaceae. Gustav Fischer Verlag Jena, 610p.

Krammer K. and Lange-Bertalot H. (2004a) Süβwasserflora von Mitteleuropa, Bacillariophyceae

3. Teil: Centrales, Fragilariaceae, Eunotiaceae. Gustav Fischer Verlag Jena, 598p.

Krammer K. and Lange-Bertalot H. (2004b) Süβwasserflora von Mitteleuropa, Bacillariophyceae

4. Teil: Achnanthaceae. Gustav Fischer Verlag Jena, 468p.

Llorens M., Saez J. and Soler A. (1992) Influence of thermal stratification on the behaviour of a

deep wastewater stabilization pond. Water Research. 26: 569-577.

Lürling M. (2003) Effects of microcystin-free and microcystin-containing strains of the

cyanobacterium Microcystis aeruginosa on growth of the grazer Daphnia magna. Environmental

Toxicology. 18: 202-210.

MAF (2005) General Description of Oxidation Pond Functions, Processes, and Performance,

Ministry of Agriculture and Forestry, Wellington, New Zealand.

Magged A.A.A. and Heika, M.T. (2006) Factors affecting seasonal patterns in epilimnion

zooplankton community in one of the largest man-mad lakes in Africa (Lake Nasser, Egypt).

Limnologica. 36: 91-97.


48

Mara D. (2003) Domestic Wastewater Treatment in Developing Countries. Earthscan publisher,

310p.

Mahassen M.E. and Sami E. F. (2011) Acute and chronic toxic effects of a Waste stabilization

pond wastewater on Daphnia magna”. Australian Journal of Basic and Applied Sciences. 5:

1371-1376.

Mara D.D. and Pearson H.W. (1998) Design manual for waste stabilization ponds in

Mediterranean countries. Lagoon Technology International Ltd, Leeds.

Mbwele L., Rubindamayugi M., Kivaisi A. and Dalhammar G. (2003). Performance of a small

wastewater stabilisation pond system in tropical climate in Dar es Salaam, Tanzania. WATER

SCIENCE AND TECHNOLOGY. 48: 187-191.

Mishra R.S. and Saksena N.D. (1990) Seasonal Abundance of the Zooplankton of Waste Water

from the Industrial Complex at Birla Nagar (Gwalior), India. Hydrobiologia. 18: 215-229.

Neel J.K. and Hopkins G.H. (1956) Experimental lagooning of raw sewage. Sewage and Indus-

trial Wastes. 28: 1326–1356.

Noumsi I.M.K., Nya J., Akoa A., Eteme R.A., Ndikefor A., Fonkou T. and Brissaud F.

(2005) Microphyte and macrophyte-based lagooning in tropical regions. WATER SCIENCE

AND TECHNOLOGY. 51: 267-274.

Padisak J. (2003) Phytoplankton. In: O’Sullian P.E. and Reynolds C.S. (Eds) The Lakes

Handbook: Volume 1 Limnology and Limnetic Ecology. Blackwell, pp. 251-308.

Pagano M. (2008) Feeding of tropical cladocerans (Moina micrura, Diaphanosoma excisum) and

rotifer (Brachionus calyciflorus) on natural phytoplankton: effect of phytoplankton size–

structure. Journal of Plankton Research. 30: 401-414.

Pahl S.L., Lewis D.M., King K.D., and Chen F. (2012) Heterotrophic growth and nutritional

aspects of diatom Cyclotella cryptica (Bacillariophyceae): effect of nitrogen source and

concentration. Journal of Applied Phycology. 24: 301-307.

Pandey, B. N., Mishra A.K., Jha A.K. and Lal R.N. (1992) Studies on qualitative composition

and seasonal fluctuation in plankton composition of River Mahananda, Katihar (Bihar). Journal

of Ecotoxicology and Environmental Monitoring. 2: 93-97.


49

Parawira W (2004). Anaerobic treatment of agricultural residues and wastewater – Application

of high rate reactors. Media-Tryck, Lund, Sweden. Pp 62-79.

Pearson H.W., Mara D.D., Mills S.W., and Smallman D.J. (1987) Physicochemical parameters

influencing the fecal bacterial survival in waste stabilization ponds. Water Science &

Technology. 19: 145-152.

Pejler B. (1983) Zooplanktic indicators of trophy and their food. Hydrobiologia. 101: 111-114.

Persoone G., Marsalek B., Blinova I., Torkokne A., Zarina D., Manusadzianas L., Nalecz-

Jawecki G., Tofan L., Stepanova N., Tothova L., Kolar B. (2003) A practical and user-friendly

toxicity classification system with microbiotests for natural waters and wastewaters.

Environmental Toxicology. 18: 395–402.

Pescod M.B. (1992) Wastewater treatment and use in agriculture. FOOD AND AGRICULTURE

ORGANIZATION OF THE UNITED NATIONS Rome, 1992.

Pielou E.C. (1966) The measurement of diversity in different types of biological collections.

Journal of Theoretical Biology. 13: 131-144.

Pietrasanta Y. et Bondon D. (1994) Le lagunage écologique. Economica. Paris. 112p.

Prescott G.W. (1951) Algae of the western great lakes – exclusive desmids and diatoms. The

Cranbook Press. 443p.

Ramadan H. and Pounce V.M (2004). Design and performance stabilization ponds.

http://stabilizationponds.sdsu.edu

Raschke R.L. (1970) Algal periodicity and waste reclamation in a stabilization pond ecosystem.

Journal Water Pollution Control Federation 42: 518-530.

Raymont J.E.G. (1980) Plankton and productivity in the oceans. In: Phytoplankton, vol. 1. Pergamon

Press, Oxford. 502p.

Reynolds C. S. (1984) The Ecology of Freshwater Phytoplankton. Cambridge: Cambridge

University Press.

Reynolds C.S. and Irish A.E. (1997) Modelling phytoplankton dynamics in lakes and reservoirs:

the problem of in-situ growth rates. Hydrobiologia, 349, 5–17.


50

Rohwer C. (1931) Evaporation from Free Water Surfaces. In US Department of Agriculture

Technical Bulletin. Vol 271.

Schultz T.E. (2005) Biotreating process wastewater: airing the options. Chemical engineering

magazine.

Shammas N.K., Wang L.K., and Wu Z. (2009) Waste Stabilization Ponds and Lagoons. In:

Handbook of Environmental Engineering, Volume 8: Biological Treatment Processes. Edited by:

Wang L.K., Pereira N.C., Hung Y.T., and Shammas N.K. Humana Press publisher.

Shanthala M., Hosmani S.P. and Hosetti B.B (2009) Diversity of phytoplanktons in a waste

stabilization pond at Shimoga Town, Karnataka State, India. Environmental Monitoring and

Sssessment. 151:437–443.

Sharma B.K. (1983) The Indian species of the genus brachionus (Eurotatoria: Monogononta:

Branhionidae). Hydrobiologia. 104: 31-39

Shiny K.J, Remani K.N., Nirmala E., Jalaja T.K. and Sasidharan V.K. (2005) Biotreatment of

wastewater using aquatic invertebrates, Daphnia magna and Paramecium caudatum.

Bioresource Technology. 96: 55-58.

Shirota A. (1966) The plankton of South Vietnam: Freshwater and marine planktons, Oversea

Technical Cooperation Agency, Japan.

Sládeček V. (1983) Rotifers as indicators of water quality. Hydrobiologia. 100: 169-201.

Smet J.D., Vasel J.L. and Dung L.Q. (2006) Wastewater treatment plant by aerated lagoon and

stabilization pond technology for Den canal in Ho Chi Minh City - Operation and Maintenance

Manual. Tan Hoa - Lo Gom Canal Sanitation and Urban Upgrading Project in Ho Chi Minh City,

Vietnam.

Smith G.M. (1924) Phytoplankton of the inland lakes of Wisconsin. Bulletin of the University of

Wisconsin, 227p.

Smith, D. G. 2001. Pennak's Freshwater Invertebrates of the United States: Porifera to

Crustacea, 4th Edition. John Wiley & Sons, 664p.

Sournia A. (1978) Phytoplankton manual. UNESCO, UK. p.77.


51

Steele, J.H., 1962. Environmental control of photosynthesis in the sea. Limnology and

Oceanography. 7: 137-150.

Stiling P.B. (1996) Ecology: Theories and Applcations, Prentice Hall International Editions, 2nd

Edition, pp. 539.

Storey M.V., van der Gaag B., and Burns B.P. (2011) Advances in on-line drinking water quality

monitoring and early warning systems. Water Research. 45: 741-747.

Suthers I.M. and Rissik D. (2009) Plankton: a guide to their ecology and monitoring for water

quality. CSIRO Publishing, 256p.

Tilman D., Kiesling R., Sterner R., Kilham S.S. and Johnson F.A. (1986) Green, bluegreen and

diatom algae: Taxonomic differences in competitive ability for phosphorus, silicon and nitrogen.

Archives fur Hydrobiologie. 106: 473-485.

Thorp J.H. and A.P. Covich. (2001) Ecology and Classification of North American Freshwater

Invertebrates, 2nd Edition. Academic Press.

Utermöhl H. (1958) Zur Vervollkommnung der quantitativen Phytoplankton-Methodik, Issue 9

of Mitteilungen, International Association of Theoretical and Applied Limnology, E.

Schweizerbart'sche.

Voigt M. (1956) Rotatoria Die Radertiere Mitteleuropas II Tafelband Gebruder Borntraeger.

BerlinNikolasse.

von Sperling M. (1996) Comparison Among the most Frequently Used Systems for Wastewater

Treatment in Developing Countries’, Water Science and Technology. 33: 59–72.

Webber M., Edwards-Myers E., Campbell C., and Webber D. (2005) Phytoplankton and

zooplankton as indicators of water quality in Discovery Bay, Jamaica. Hydrobiologia. 545:177–

193.

West W. and West G.S. (1904) A monograph of the British Desmidiaceae. Vols I – V. Johnson

Reprint Corporation.

Williams L.G. (1966) Dominant planktonic rotifers of major water ways of the United States.

Limnology and Oceanography. 11: 83-91.


52

Yamagishi T. and Akiyama M. (1994a) Photomicrographs of the freshwater algae. Uchida

Rokakuho. 12: 1-100.

Yamagishi T. and Akiyama M. (1994b) Photomicrographs of the freshwater algae. Uchida


Yamagishi T. and Akiyama M. (1995) Photomicrographs of the freshwater algae. Uchida


Yildiz S., Altindag A. and Ergonul B.E. (2007) Seasonal fluctuations in the zooplankton

composition of a eutrophic lake: Lake Marmara (Manisa, Turkey). Turkish Journal of Zoology.

31: 121-126.

Yusoff F.M. and McNabb C.D. (1997) The effects of phosphorus and nitrogen on phytoplankton

ominance in tropical fish ponds. Aquaculture Research. 28: 591-597.

Zeng Y., Fu X., and Ren Z. (2012) The effects of residual chlorine on the behavioural responses

of Daphnia magna in the early warning of drinking water accidental events. Procedia

Environmental Sciences. 13: 71-79.

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August 2012. Promoters Prof. Dr. Eng. Peter Goethals Prof ... · Ecological study of a large scale treatment pond system in Ho Chi Minh City (Vietnam) PREFACE This work is the reflection