PHYTOPLANKTON COMPOSITION AND ITS RELATED ENVIRONMENTAL CONDITIONS IN SANTUBONG AND SAMARIANG

ESTUARIES, KUCHING, SARAWAK

Siti Zubaidah Binti Kamarudin

Master of Science (Marine Science)

2012

Pusat Khidmat Maklumat Akademik [! NiVFRSM MALAYSIA SARAWAK

PHYTOPLANKTON COMPOSITION AND ITS RELATED ENVIRONMENTAL

CONDITIONS IN SANTUBONG AND SAMARIANG ESTUARIES, KUCHING,

SARAWAK

Siti Zubaidah Binti Kamarudin

This thesis is submitted in fulfillment of requirement for the degree of

Master of Science

(Marine Science)

Faculty of Resource Science and Technology

Universiti Malaysia Sarawak

2012

DECLARATION

I hereby declare that no portion of the work referred to this thesis have been submitted in

support of an application for another degree of qualification to this or any other university

of institution of higher learning.

(Siti Zubaidah Binti Kamarudin)

Date:

11

ACKNOWLEDGEMENTS

Most and foremost I would like to thank God for giving me the strength and capability in

doing this research. I would like to thanks Universiti Malayisa Sarawak for the Zamalah

Postgraduates Scholarship. I am also extraordinary grateful to my supervisor and my co

supervisor, Dr Lim Po Teen and Dr Leaw Chui Pin for their encouragements, critical

comments, recommendation, and guideline as well as discussion throughout the study. Not

forgetting, people from the Fisheries Department in Santubong and Samariang. Thank you

very much for the approval and accessibility to the sampling sites and the cooperation.

I am grateful to Hartina, Lim Hong Chang, Suriyanti, Fareha Hj Hilaluddin, Tan

Toh Hii, Teng Sing Tung, Hii Kien Soon and all lab mates for giving and sharing

knowledge and priceless experiences with me. My special appreciation due to the aquatic

laboratory assistant, Mr. Nazri Latip, and science officer, Mr. Shafri, for their assistances.

To my family, for always be by my side and strengthen me up; my housemates and friends,

for giving me supports, comment and encouragement, thank you very much. God bless

you.

ill

ABSTRACT il

ýA study was carried out to investigate phytoplankton composition in two estuaries of Kuching, Sarawak. Two hundred and two samples of phytoplankton were collected from

the estuaries of Samariang and Santubong fortnightly from 2008 to 2011 by using a Van

Dorn water sampler and plankton net. Water parameters such as pH, salinity, and

temperature were determined in-situ. Selected macronutrients concentration (phosphate,

nitrate-nitrogen and silicate) of water samples were analyzed in the laboratory.

Phytoplankton enumeration and identification were undertaken by using light and electron

microscopy. Ninety five taxa, composed of seventy eight species of diatoms and seventeen

species of dinoflagellates were identified from the study area Phytoplankton composition

was dominated most by diatom group in both sampling sites but dominated six times in

Samariang. The species frequently found predominantly in the water samples of the both

sampling sites were Nitzcshia spp., Coscinodiscus spp., Thalassionema spp. and Pleurosigma spp.. The predomination of single genus of phytoplankton at single time

resulted in higher cell density and shift in species composition was observed throughout

the sampling period. Seasonal occurrence of diatom cell density was recorded from April

to August in Santubong and from July to September and April to May and Samariang.

Compared to Santubong, dinoflagellates cell density found higher in Samariang. No bloom

event of diatom or dinoflagellates was recorded during the sampling period but low

Diatom to Dinoflagellates (DD) ratios in some occasion could serve as the window of

opportunity for dinoflagellate to bloom. However at this juncture no bloom was observed

which could be limit by the change in environmental conditions (pH, salinity, temperature

and macronutrients. Statistical analysis with One Way ANOVA showed that higher cell

density of diatom and dinoflagellates in both sampling site have significant difference with

environmental conditions. However, analysis with Principle Components Analysis (PCA)

showed that only pH, phosphate and temperature have positively correlated with cell

abundance of diatom and dinoflagellates. In this study, potentially harmful algae such as

Pseudo-nitzschia species were frequently detected in the plankton samples. Thus a laboratory experiment to determine the salinity tolerance of Pseudo-nitzschia species was

undertaken. In culture experiment, a total of nine strains of four species of Pseudo-

nitzschia showed different, salinity tolerance, in decreasing order of P. pungens > P.

brasiliana >P. cuspidata > P. circumpora. Maximal growth rate was greatest in P.

pungens (2.21), followed by P. cuspidata (1.57), P. brasiliana (1.52) and P. circumpra

iv

(1.01). Morphological abnormality of Pseudo-nitzschia was documented at different

salinities and shown to be varied among the species. Irregularity in cell shape was

documented in all species while appearance of bulges was observed only in P. pungens and

P. cuspidata. Cell abnormality was highest in P. circumpora and lowest in P. cuspidata. In

addition, reduction in cell length was observed at all strains but not significant since short

duration of experiment applied in this study. Further study using microscosm experiment is

important to evaluate the influence of environmental conditions to the changes of

phytoplankton compositions especially the abundance and growth of potential harmful

species.

Keywords: phytoplankton, macronutrient, salinity, diatom, dinoflagellates

V

KOMPOSISI FITOPLANKTON DAN KEADAAN PERSEKITARAN YANG

BERKAITANNYA DI MUARA SUNGAI SANTUBONG DAN SAMARIANG,

KUCHING, SARA WAK

ABSTRAK

Satu kajian telah dijalankan untuk mengkaji komposisi fitoplankton di dua muara sungai

Sarawak. Dua ratus dua sampel fitoplankton telah dikumpulkan dari setiap dua kali

sebulan penyampelan dilakukan dari muara Sungai Santubong dan Samariang dari 2008

ke 2011 dengan menggunakan penyampel air Van dorn dan faring Plankton. Parameter

air seperti pH, kemasinan dan suhu dilakukan in situ, manakala analisis makronutrien

terpilih (fosfat, nitrat-nitrogen dan silika) dilakukan di makmal. Pengecaman dan

pengiraan fitoplankton dilakukan menggunakan mikroskop cahaya dan elektron. Daripada

Sembilan puluh lima taksa, tujuh puluh lapan taksa diatom dan tujuh belas taksa

dinoflagella telah dikenal pasti di kawasan kajian. Komposisi fitoplankton kebanyakannya

didominasi oleh kumpulan diatom di kedua dua kawasan kajian tetapi dinoflagela

mendominasi sebanyak tujuh kali di Samariang. Spesies yg kerap menjadi pradominasi di

dalam sampel air di kedua dua kawasan kajian adalah Nitzcshia spp., Coscinodiscus spp.,

Thalassionema spp. and Pleurosi =a spp.. Pradominasi oleh satu genus fitoplankton pada

satu- satu masa mengakibatkan kepadatan sel yang tinggi dan perubahan dalam komposisi

spesies diperlihatkan sepanjang tempoh kajian. Kejadian bermusim bagi sel diatom adalah

tinggi direkodkan dari April ke August di Santubong dan Julai ke September dan April ke

Mci di Samariang. Berbanding Santubong, kepadatan sel dinoflagela adalah lebih tinggi

di Samariang. Tiada ledakan diatom atau dinoflagella diperhatikan semasa penyampelan

dilakukan namun beberapa nisbah diatom kepada dinoflagela (DD) yang rendah di

diperhatikan mampu membuka peluang kepada ledakan dinoflagela tetapi telah dihadkan

oleh keadaan alam sekitar (pH, suhu, kemasinan air dan makronutrien). Analisis statistik

dengan One Way ANOVA menunjukkan bahawa sel diatom dan dinoflagela di kedua dua

tapak penyampelan mempunyai signifikan terhadap keadaan alam sekitar. Walau

bagaimanapun, analisis dengan Principle Component Analysis . (PCAý menunjukkan

bahawa hanya pH, fosfat dan suhu mempunyai kaitan positif dengan kepadatan sel diatom

dan dinofalgela. Kehadiran alga yang berpotensi berbahaya seperti spesies Pseudo-

vi

nitzschia di perairan juga telah dikesan dalam sampel plankton. Justeru itu, satu ujikaji

makmal telah dilakukan bertujuan mengkaji toleransi kemasinan air Pseudo-nitzschia

spesis. Dalam kajian kultur, sembilan klon yang terdiri daripada empat spesies Pseudo-

nitzschia menunjukkan variasi dalam toleransi terhadap kemasinan air iaitu P. up ngens >

P. brasiliana > P. cuspidata > P. circumpora. Kadar pertumbuhan maksimum pula tinggi

pada P. up ngens (2.21), diikuti P. cuspidata (1.57), P. brasiliana (1.52) dan P. circumpora

(1.01). Morfologi abnormal Pseudo-nitzschia spesies pada kemasinan air yang berbeza

telah didapati dan menunjukkan perbezaan antara spesies. Bentuk sel yang luar biasa

telah didapati pada setiap spesies manakala bonjolan-bonjolan hanya didapati pada P.

pungens dan P. cuspidata. Sel abnormal tinggi pada P. circumpora dan rendah pada P.

cuspidata. Pengurangan dalam panjang sel diperhatikan pada setiap klon tetapi tidak

signifikan kerana tempoh eksperimen yang singkat digunakan dalam kajian ini. Kajian

menggunakan eksperimen mikrokosma adalah penting untuk mengenalpasti kesan alam

sekitar terhadap komposisi fitoplankton terutamanya kepadatan dan pertumbuhan spesies

berpotensi berbahaya.

Kata kunci: fitoplankton, makronutrien, kemasinan, diatom, dinoflagela

vii

Pusat Khidmat Maklumat Akademik UNIVERSTfi MALAYSIA SARAWAK

TABLE OF CONTENTS

DECLARATION ACKNOWLEDGEMENTS ABSTRACT ABSTRAK TABLE OF CONTENTS LIST OF TABLES LIST OF FIGURES

CHAPTER I INTRODUCTION

CHAPTER II PHYTOPLANKTON COMPOSITION IN THE TWO ESTUARIES OF KUCHING, SARAWAK

2.1 INTRODUCTION

2.2 MATERIALS AND METHODS 2.2.1 Study sites and sampling 2.2.2 Species identification and enumeration 2.2.3 Macronutrients analysis

2.2.3.1 Nitrate - Nitrogen (N03- - N) 2.2.3.2 Orthophosphate (P043" - P) 2.2.3.3 Silicate (Si02)

2.2.4 Rainfall distribution 2.2.5 Statistical analysis of the data

2.3 RESULTS 2.3.1 Phytoplankton composition in Santubong estuary 2.3.2 Phytoplankton composition in Samariang estuary 2.2.3 Physio-chemical data

Page ii

iv vi

viii x

xi

I

6

7

8 11 11 11 12 12 12 12

13 17 18 29

2.2.3.1 Water parameter 29 2.2.3.2 Macronutrients 32 2.2.3.3 Fortnightly rainfall data 35

2.2.4 Principle Components Analysis (PCA) 37

2.4 DISCUSSION 40 2.4.1 Phytoplankton composition in Santubong and 40

Samariang estuaries 2.4.2 Effects of macronutrients on phytoplankton 41

composition 2.4.3 Effects of physical factors on phytoplankton 45

composition 2.4.4 Potential harmful algae 49

viii

CHAPTER III EFFECTS OF SALINITY ON GROWTH OF PSEUDO- 52 NITZSCHIA SPECIES UNDER CONTROLLED TEMPERATURE AND LIGHT INTENSITY

3.1 INTRODUCTION 52

3.2 MATERIALS AND METHODS 54 3.2.1 Sampling sites 54 3.2.2 Culture media preparation 57 3.2.3 Growth -salinity experiment 57 3.2.4 Cell size reduction measurement 58 3.2.5 Scanning electron microscope preparation 59 3.2.6 Statistical analysis of One Way ANOVA 59

3.3 RESULTS 3.3.1 Effect of salinity on the growth of Pseudo-nitzschia 60

species 3.3.1.1 Pseudo-nitzschia pungens 60 3.3.1.2 Pseudo-nitzschia brasiliana 67 3.3.1.3 Pseudo-nitzschia circumpora 75 3.3.1.4 Pseudo-nitzschia cuspidata 78

3.4 DISCUSSION

CHAPTER IV CONCLUSION

REFERENCES

APPENDICES

83

88

91

108

ix

LIST OF TABLES

Table Page

Table 2.1 Dinoflagellates taxa as there are those identified to the genus or to 14 the genus level present in Santubong and Samariang estuaries.

Table 2.2 Diatom taxa as there are those identified to the genus or to the genus 15 level present in Santubong and Samariang estuaries.

Table 2.3 Summary of PCA for the first two axes. 38

Table 3.1 Strains of Pseudo-nitzschia species used in the study. 55

Table 3.2 Components used in preparing SWII medium. 57

Table 3.3 Initial length and cell length of Pseudo-nitzschia pungens at the end 66 of exponential phase.

Table 3.4 Initial length and cell length of Pseudo-nitzschia brasiliana at the 74 end of exponential phase.

Table 3.5 Initial length and cell length of Pseudo-nitzschia circumpora at the 78 end of exponential phase.

Table 3.6 Initial length and cell length of Pseudo-nitzschia cuspidata at the 82 end of exponential phase.

X

LIST OF FIGURES

Figure Page

Figure 2.1 Map of Kuching, Sarawak showing locations of Santubong and 10 Samariang estuaries where samples were collected.

Figure 2.2 Micrographs of phytoplankton found in Santubong and Samariang. 16 Diatom of (a) Coscinodiscus sp., (b) Chaetoceros sp., (c) Thalassionema sp., (d) Pseudo-nitzschia sp., (e) Nitzschia sp., (f) Pleurosigma sp,. (g) Guinardia sp., (h) Skeletonema sp., and (i) Navicula sp., and dinoflagellates of (j) Bacillaria sp., (k) Ceratium sp., (1) Peridinium sp., (m) Dinophysis sp., (n) Alexandrium sp., (o) Prorocentrum sp. (p) and Pyrophacus sp.

Figure 2.3 Average cell density of diatom and dinoflagellates (cells L-') and 20 diatom to dinoflagellates ratio (cells L") of Santubong (a) and Samariang (b) from July 2008 until February 2011.

Figure 2.4 Species composition in Santubong from July, 2008 to February, 21 2011.

Figure 2.5 Species composition in Samariang from July, 2008 to February, 25 2011.

Figure 2.6 Water parameter of Santubong (a) pH, (b) temperature, (c) salinity 31 and Samariang (d) pH, (e) temperature and (f) salinity taken from July, 2008 until February, 2011.

Figure 2.7 Macronutrients (Nitrate- nitrogen, Phosphorus, Silicate) reading at 34 Santubong and Samariang from July, 2008 to February 2011.

Figure 2.8 Average fortnightly precipitation rate of Kuching district from July, 36 2008 to February, 2011 (Source: Department of Meteorology, Malaysia).

Figure 2.9 Diagram of PCA showing relationships among nine variables at two 39 study sites. Both axes were significantly correlated with all variables with the total variance of 44%.

Figure 3.1 Sexual reproduction of pennate diatom (Source: Hasle and 52 Syvertsen, 1997).

Figure 3.2 Map of Malaysia showing location of the sampling sites. 56

Figure 3.3 Average cell. density per day (Cells mL') of P. pungens from Kudat, 62 PnKd03 (a), P. pungens from Santubong, PnSb02 (b), and P. pungens from Kota Kinabalu, PnKk24 (c) under salinity treatments of 5 to 35 PSU.

X1

Figure Page

Figure 3.4 Growth rate per day, p and maximum cell density of P. pungens 63 from Kudat, PnKd03 (a), P. pungens from Santubong, PnSb02 (b), and P. pungens from Kota Kinabalu, PnKk24 (c) under salinity treatments of 5 to 35 PSU.

Figure 3.5 Cell abnormality of Pseudo-nitzschia pungens, in stationary phase 64 of culture at salinity 10 PSU and 20 PSU observed under SEM. (a - c), cells with the appearance of bulges at the central part of valves. Scale bar = 10 µm. (d - f), close up of bulges.

Figure 3.6 Cell abnormality of Pseudo-nitzschia pungens, at salinity 30 PSU in 65 stationary phase observed under SEM. Scale bar for a-b= 10 µm. (a - c): cells with the appearance of bulges at the central part of the cell. (d and e): close up images of bulges at the central part of the cell.

Figure 3.7 Average cell density per day (Cells mL') of P. brasiliana from 70 Pulau Mamutik, PnPm05 (a), P. brasiliana from Samariang, PnSm26 (b), P. brasiliana from Kota Kinabalu, PnKk25 (c), and P. brasiliana from Santubong, PnSb43 (d), under salinity treatments of 5 to 35 PSU.

Figure 3.8 Growth rate per day, µ and maximum cell density of P. brasiliana 71 from Pulau Mamutik, PnPm05 (a), P. brasiliana from Samariang, PnSm26 (b) P. brasiliana from Kota Kinabalu, PnKk25 (c) and P. brasiliana from Santubong, PnSb43 (d) under salinity treatments of 5 to 35 PSU.

Figure 3.9 Cell abnormality of Pseudo-nitzschia brasiliana, at salinity 15 PSU 72 during stationary phase observed under SEM. (a - d): valve view of intruded and undulated valves at the central part of the cells and bent at apical end. (e - g): close up images of deformed parts of cells. Scale bar for a-d=5 gm; e-g =1 gm.

Figure 3.10 Cell abnormality of Pseudo-nitzschia brasiliana, at salinity 30 PSU 73 during stationary phase observed under SEM. (a - e): valve view of irregular shape, undulated and intruded valves at the central part of the cell. Scale bar for a-d=5 µm.

Figure 3.1 1 Average cell density per day (Cells mL-1) of P. circumpora from 76 Santubong, PnSb58 under salinity treatments of 5 to 35 PSU.

Figure 3.12 Growth rate per day, u and maximum cell density of P. 76 circumpora from Santubong, PnSb58 under salinity treatments of 5 to 35 PSU.

xii

Figure Page

Figure 3.13 Cell abnormality of Pseudo-nitzschia circumpora, clonal culture 77 from Santubong, Sarawak at salinity 25 PSU (a - c) and 30 PSU (d - f) during stationary phase observed under SEM. (a -d and f): images showing cells undergo deformed shape such as undulated valve, protruded and intruded valve along the cell. (e) : two similar deformed cells which came after cell division. Scale bar for a-c= 5 µm.

Figure 3.14 Average cell density per day (Cells mL-1) of P. cuspidata from 79 Likas, PnLk02 under salinity treatments of 5 to 35 PSU.

Figure 3.15 Growth rate per day, p and maximum cell density of P. cuspidata 80 from Likas, PnLk02 under salinity treatments of 5 to 35 PSU.

Figure 3.16 Cell abnormality of Pseudo-nitzschia cuspidata, clonal culture from 81 Likas, Sabah at stationary phase observed under SEM. (a - d): valve view of cells at salinity of 20 PSU. (e - h): valve view of cells at salinity of 30 PSU. (a -c and f- g): showing deformation occurred along the cell with the appearance of bulges, intruded and undulated valve and non linear shape. (d and e): close up images of deformed part of the cells. Scale bar for a-c and f-g= 10 µm; for d and e =1 µm.

xiii

CHAPTER I

INTRODUCTION

Phytoplankton community contributes to more than 95% of marine primary production

(Steeman Neilson 1975) which is important to higher trophic levels. It influences the

efficiency of carbon and energy transfer between trophic levels in the aquatic ecosystems.

Diatoms are the major contributors to primary production, they occur everywhere in the

sea but most abundant in colder, nutrient- rich waters. Dinoflagellates are the second

contributor to the marine primary production and also widespread across the oceans. Other

groups of phytoplankton are coccolithophorids, silicoflagellates, blue-green algae and

prochlorophytes (Valiela 1995).

The importance of phytoplankton is largely due to the very large area of the earth

covered by open sea. Mixtures of microalgae often can be found growing in or anchored to

sands, and mud bottoms in shallow waters. In tropical regions, some are found living

within the tissue of corals such as symbiotic dinoflagellates, zooxanthellae (Valiela 1995).

Abundance of phytoplankton is regulated by various environmental factors. In

temperate region, phytoplankton abundance most likely is controlled by irradiance and

water temperature (Day et al. 1989). Elevation of irradiance and water temperature

contributed to the peak of phytoplankton production in summer season (Mallin et al.

1991). Increase in nutrient loading and temperature were the major factors that contributed

to the shift of Pseudo-nitzschia species composition (Lundhlom et al. 2010). Wang et al.

(2008) showed that increase of human activities in coastal systems affects nutrient loading

I

and consequently affect phytoplankton composition. The bloom of Cochlodinuim sp.

occurred after a short period of dry season followed by heavy rains and strong winds with

an enhancement of nutrient received from aquaculture activities (Anton et al. 2008).

About two percent of total phytoplankton genera are known as potent phycotoxin

producers. Rapid growth of these microalgae resulting in high densities could result in

negative impact to the surrounding environments. Harmful algal bloom (HAB) is a natural

phenomenon caused by drastic increase of a single species in the water bodies resulting

from favorable conditions for growth. This event is detected at the water surface as colour

changes to be greenish, reddish or brownish depending the on pigments of the blooming

species.

Paralytic shellfish poisoning (PSP) is the most pivotal form of worldwide seafood

toxicity due to HABs (Hallegraeff 1993). The first report of HABs and shellfish toxicity in

Malaysia was documented back in 1976 when the marine dinoflagellate, Pyrodinium

bahamense var. compressum bloomed in the Brunei Bay on the west coast of Sabah (Roy

1997). Several people were poisoned during this event and the bloom eventually spread to

other parts of the west coast of Sabah (Usup et al. 2002). After 14 years of the first

incident, once again three people were taken ill after consuming green mussel (Perna

viridis) cultured at a newly established mussel farm in Sebatu, Malacca. PSP took place

again in Tumpat, Kelantan in September 2001 with six people hospitalized including one

death after consuming `lokan' (Polymesoda sp. ) collected from a coastal lagoon (Sungai

Ubi) (Lim et al. 2004). The symptoms and further toxin analysis confirmed the poisoning

was due to algal toxins produced by the dinoflagellate, Alexandrium minutum (Lim et al.

2004). In 2009, blooms of P. bahamense recurred in the Kota Kinabalu coast and the

shellfish mollusks were banned from harvesting due to shellfish contamination with

toxicity of more than 7,000 mouse units (Daily Express 2009).

2

Amnesic Shellfish Poisoning (ASP) or domoic acid (DA) poisoning is one of the

life-threatening syndromes of HABs caused by diatoms (Bates and Trainer 2006). ASP

included gastro-intestinal and neurological symptoms (Hallegraeff 2003). The first incident

of ASP was reported in 1987 with over 100 people sickened and three deaths after

consuming DA-contaminated mussels from Eastern Prince Edward Island (Bates et al.

1989; Wright et al. 1989). Since then, the number of poisoning cases related to ASP has

increased worldwide. Following the numerous incidence of ASP worldwide, monitoring of

plankton and shellfish toxicity have been implemented in shellfish areas of affected

countries.

Several species of the genus Pseudo-nitzschia are known to produce DA

(Lundholm et al. 2003). DA and its derivatives are water-soluble and heat-stable

tricarboxylic amino acid (Van Dolah 2000; Rue and Bruland 2001). DA may accumulate in

the food web in a number of organisms like crabs, lobsters, fish and shellfish that serve as

the vector. DA is accumulated in the digestive system of vector organisms and is present in

the flesh in low concentrations. Mass mortalities of more than 100 brown pelicans,

cormorants and marine mammals involving hundreds of sea lions accussed after eating the

toxic mackerel and anchovies in the coast of California and Mexico (Busse et al. 2006).

Shellfish farms in Scottish waters terminated harvesting in 1999 due to elevated DA in the

King scallop that exceeded the regulatory limit (Campbell et al. 2001). There is no known

antidote for ASP thus far (Hallegraeff 2003).

Since year 2000, HAB events have been increasingly reported in the country both

in the Straits of Malacca (Usup et al. 2002) and South China Sea (Anton et al. 2008).

Bloom of Prorocentrum minimum was reported to have lasted for months in the semi-

enclosed Tebrau Strait (Usup et al. 2002). One of the species responsible for the blooms in

Sabah coasts was Cochlodinium polykrikoides that resulted in mortality of cage cultured

3

fish (Anton et at. 2008). The same species was also reported to form bloom in aquaculture

site of Prai, Penang. In 2007, Neoceratium furca (previously as Ceratium furca) was

identified in the algal bloom in Pangkor and Penang (The Star 2007). Latest occurence of

HABs in Malaysia was reported in 2010 with fish-kills in Tanjung Adang, Johore where

the waters turned yellowish and murky (The Star 2010). The culprit of the discoloration

was still unknown.

Diatoms are valuable indicators of the environmental condition in a river or stream

because they reproduce and respond rapidly to environmental changes and provide early

warning for pollution (Stevenson and Pan 1999). Diatoms react rapidly to environmental

changes by shifting its composition, as the composition is affected by eutrophic conditions

(Case et al. 2008). They serve as indicators of anthropogenic influences on ecological

quality of streams, and therefore numerous diatom indices were developed for status

assessment (Kovacs 2007). Cooper (1999) suggested that diatoms are useful indicators in

estuarine system since they have specific adaptation and are able to provide clues to the

past environment of the area. Diatoms have been applied as bioindicator to measure water

quality of the Keelung River in Taiwan (Wu 1999), and radionuclide contamination in the

Swedish nuclear power plant (Snoeijs 1999). The ability of diatoms to respond rapidly to

actual changes of radionuclide concentration in the waters was higher compared to other

phytoplankton groups (Snoeijs 1999).

Salinity was one of the factors that influenced the distribution and abundance of

diatoms in the coastal waters including the diatom Pseudo-nitzschia species. Pseudo-

nitzschia was known to be widely distributed with records from around the globe (Halle

2002; Lundholm et al. 2003; Casteleyn et at. 2008) and presented in the water column

whole year round in some. areas (Qi et al. 1996; Yap-Dejeto 2008; Klein et at. 2010).

Numerous toxic and non toxic Pseudo-nitzschia species are known to be widespread in

4

Pusat Khidmat Maklumat Akademik UNIVERSITI MALAYSIA SARAWAK

coastal regions with variable salinity regimes due to euryhaline characteristic of this

species (Brand 1984; Hasle 2002; Thessen et al. 2005). Thessen et al. (2009) also reported

broad ecological niche of Pseudo-nitzschia species with its ability to grow under various

environments.

Studies on the influence of environmental conditions on growth and distribution of

Pseudo-nitszchia have been conducted by various researchers. The effect of pH (Vrieling

et al. 1999; Lundholm et al. 2004), salinity regimes (Gordon and Brodland 1990; Vrieling

et al. 2000,2007; Thessen et al. 2005; Leterme et al. 2010), ambient nutrient concentration

(Caroppo et al. 2005) and photoperiod (Fehling et al. 2005) on ecophysiological adaptation

on Pseudo-nitzschia was well documented in the last decade. However, these studies were

conducted on temperate Pseudo-nitzschia strains/species at higher latitude region.

Ecophysiological characteristics of tropical strains in warm water regions are still poorly

understood. Therefore, in this study effects of salinity on growth and morphology of

Pseudo-nitzschia species were investigated.

In Malaysia, studies on phytoplankton composition in the estuaries of Sarawak are

still lacking. There were previous phytoplankton studies documented in Malaysian waters

(Boonyapiwat 1998; Shamsudin 1990; Wan Maznah et al. 2000; Su 2011). However,

studies of Boonyapiwat (1998) did not state any relationship of phytoplankton composition

with environmental factors. In addition, Shamsudin (1990) focused on the ecology of

diatoms, and Wan Maznah et al. (2000) focused on the freshwater planktons. Meanwhile,

studies of Su (2011) had documented the abundance of Pseudo-nitzschia species in relation

to the environmental conditions in the estuaries of Kuching, Sarawak. The study had

shown significant correlation between species abundance and several environmental

factors. However, this study only focused on a single genus, viz. Pseudo-nitzschia, without

examining the community structure of phytoplankton species. This finding encouraged

5

further investigation into the succession of these potential ASP producers and the

phytoplankton assemblages in the same habitats.

The present study aims to obtain data on phytoplankton composition and

environmental factors that regulate the phytoplankton composition in the two estuaries of

Kuching, Santubong and Samariang focusing on the two main phytoplankton groups, the

diatom and dinoflagellates. This information is important and could provide insights into

understanding the succession of phytoplankton community which may relate to algal

blooms. The specific objective of the study is to investigate the compositon of

phytoplankton in the two estuaries, focusing on the two main phytoplankton groups, the

diatoms and dinoflagellates (Chapter II).

To date, there is no record of ASP in Malaysian waters. However, several studies

have been done to understand the emerging factors of ASP and its causative organism,

Pseudo-nitzschia species (Lim et al. 2010; Su 2011). Furthermore, little is known of the

physiological adaptation of its species. In Chapter III, a study on the effects of different

salinity regimes on growth and frustules morphology of Pseudo-nitzschia species were

undertaken. Frustules morphological observation of Pseudo-nitzschia species was

undertaken using scanning electron microscope (SEM).

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CHAPTER II

PHYTOPLANKTON COMPOSITION IN TWO ESTUARIES OF KUCHING,

SARAWAK

2.1 INTRODUCTION

Phytoplankton is a group of photosynthetic microorganisms occupying the euphotic zone

of water column, which supports the food chain in aquatic ecosystem. Generally, diatom

and dinoflagellates are the two main groups of phytoplankton broadly found in high

abundance in the marine, brackish as well as freshwater systems. The abundance of

phytoplankton in the water bodies is regulated by their environmental surroundings such as

light, temperature, salinity, water column stability, and nutrients (Anderson et al. 2002).

Sudden increase in abundance or density of certain groups of phytoplankton can lead to

algal blooms, which cause degradation of water quality, adverse effects to human health,

marine organism fatality, and other economic losses (Backer 2009).

One of the factors that trigger algal blooms in the water ecosystem is

eutrophication. Eutrophication can be defined as loading of macronutriens such as

phosphorus, nitrate and silicate into water ecosystems that promoting the growth and

bloom of algae especially diatoms and dinoflagellates (Livingston 2001). Algal bloom

triggered by eutrophication also resulted in higher primary productivity in affected area

(Hall and Smol 1999). Harmful algae bloom events are usually dominated by a single

species of phytoplankton (Usup et al. 1989; Lim et al. 2005). Among the sources of

eutrophication, are increase in coastal inhabitants, anthropogenic activities, mariculture and

terrestrial derived nutrient loading (Anderson et al. 2002). High concentration of nutrient

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loading into water ecosystem can be either direct or indirect from surrounding environment

(Larry 1996). Snoeijs (1999) reported that the abundances of biological species changed as

the environment changed due to human activities, and community composition can be used

as an indicator for the direction and rate of change caused by these activities.

Currently, studie on harmful algal blooms (HABs) focused mainly on the growth

physiology of the target organism and its response to environmental conditions. Somehow,

this has underestimated the role of other phytoplankton species co-occuring in the same

habitat, since most phytoplankton share similar niches in the ecosystem. Thus, this study

aims to establish baseline data on the phytoplankton composition of Santubong and

Samariang estuaries. Beside, the effects of environmental factors on phytoplankton

composition were also investigated in both estuarine waters.

2.2 MATERIALS AND METHODS

2.2.1 Study site and sampling

In this study, phytoplankton and water samples were collected fortnightly during

high tide from Santubong (1°117'50" N 110° 19'34" E) and Samariang (1°36'33" N 110°

19'27" E) estuaries (Figure 2.1) from July 2008 to Feb 2011. The two estuaries, Santubong

and Samariang were selected due to their distinctive character of estuarine waters where

Santubong is situated in the river mouth of Santubong while Samariang was situated 3 km

upstream of the estuary. Sampling site of Santubong located at the station of Fisheries

Department adjoining to the river mouth of Santubong. In Samariang, the sampling site is

located at the jetty of Sarawak Inland Fisheries station. A cage culture is operating nearby

of the sampling site. The area is surrounded by mangrove forests, with habitats for many

aquatic living, such as mudskipper fish, and crocodiles. Shrimp and fish cage cultures are

operating in this area to meet the increasing demands for protein source. Futhermore,

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Santubong and Samariang are the main route of transportation for fishermen to commute

daily to the sea. The two estuaries were subjected of nutrient loading to the nearby

receiving waters due to anthropogenic activities and coastal development in the areas.

Water samples were collected by using a Van Dorn water sampler. During the

sampling, duplicates of one liter sample were collected at Im depth during high tide. One

liter of phytoplankton samples were concentrated filtration unit fitted with glass fiber filter.

Samples were back wash into centriguge tube with final volume 50 mL. All samples were

preserved in acidic-Lugol solution and stored in dark at ambient temperature. Selected

water parameters such as salinity, temperature and pH from the two sampling locations

were determined in-situ by using HANNA electronic pH meter (HANNA Instrument,

USA) and AGATO hand refractometer (Agato, Japan).

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Figure 2.1: Map of Kuching, Sarawak showing locations of Santubong and Samariang estuaries where samples were collected.

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