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  • UNIVERSITI PUTRA MALAYSIA

    APPLICATION OF BACILLUS SPHAERICUS UPMB10 FOR GROWTH ENHANCEMENT OF BANANA (MUSA SPP. VAR. BERANGAN) AND

    ITS EFFECT ON FUSARIUM WILT

    ILLANI ZURAIHAH BINTI IBRAHIM

    FP 2008 32

  • APPLICATION OF BACILLUS SPHAERICUS UPMB10 FOR GROWTH

    ENHANCEMENT OF BANANA (MUSA SPP. VAR. BERANGAN) AND ITS

    EFFECT ON FUSARIUM WILT

    ILLANI ZURAIHAH BINTI IBRAHIM

    MASTER OF SCIENCE UNIVERSITI PUTRA MALAYSIA

    2008

  • ii

    APPLICATION OF BACILLUS SPHAERICUS UPMB10 FOR GROWTH

    ENHANCEMENT OF BANANA (MUSA SPP. VAR. BERANGAN) AND ITS

    EFFECT ON FUSARIUM WILT

    By

    ILLANI ZURAIHAH BINTI IBRAHIM

    Thesis submitted to the School of Graduate Studies, Universiti Putra

    Malaysia, in Fulfillment of the Requirement for the Degree of Master of

    Science

    SEPTEMBER 2008

  • iii

    DEDICATION

    Special dedication to:

    Zull and Danial

    My lovely family…. For all the supports, endless loves and advices throughout

    the long journey to complete my study…. Keeps me alive!

  • iv

    Abstract of thesis presented to the Senate of Universiti Putra Malaysia in fulfillment of the requirement for the degree of Master of Science

    APPLICATION OF BACILLUS SPHAERICUS UPMB10 FOR GROWTH

    ENHANCEMENT OF BANANA (MUSA SPP. VAR. BERANGAN) AND ITS

    EFFECT ON FUSARIUM WILT

    By

    ILLANI ZURAIHAH BINTI IBRAHIM

    September 2008

    Chairman : Professor Zulkifli Hj Shamsuddin, Ph.D. Faculty : Agriculture

    A locally isolated strain Bacillus sphaericus UPMB10, categorized as plant

    growth promoting rhizobacteria (PGPR), was reported to have beneficial

    effects in stimulating growth and yield of many agricultural crops. This study

    explores the indirect mechanism of B. sphaericus UPMB10 that would support

    the hypothesis that this PGPR could act as a biocontrol agent. Through in-

    vitro study, B. sphaericus UPMB10 was observed to inhibit the growth of

    Fusarium oxysporum f.sp cubense race 4 (FOCR4) mycelium by 70% in

    culture medium. Malformation of the fungal hyphae by vacuole formation,

    swelling and thickening of the hyphal strands occurred with the presence of B.

    sphaericus UPMB10. In addition, B. sphaericus UPMB10 showed positive

  • v

    results in the tests for production of cell wall-degrading enzymes and

    phosphate solubilizing activity by producing clear zones or halos. It can also

    produce a plant growth regulating hormone, indole acetic acid (IAA). An initial

    glasshouse study showed that inoculation with B. sphaerius UPMB10

    increased plant growth up to 30% in plant height and increased nutrient

    uptake namely for N, P, K, and Ca, and also reduced the disease severity

    through visual symptoms and vascular discoloration of the corm. In the last

    glasshouse experiment, the appearance of the disease incidence was

    observed. All of the treatments with FOCR4 application were infected by

    FOCR4 including those treated with B. sphaericus UPMB10. Significantly, the

    infection occurred at the late stage of the experiment indicating that B.

    sphaericus UPMB10 could delay the infection process. Solid inoculum

    inoculation with B. sphaericus UPMB10 showed a delayed onset of

    symptoms, lower percentage in disease incidence and epidemic rate. As

    liquid and solid substrate were selected for evaluation against the disease in

    the glasshouse, it showed that soil application of B. sphaericus UPMB10 with

    solid inoculum provided the best suppression of Fusarium wilt of banana. In

    this study, an inoculum dose of 100 g of solid formulation per plant was

    optimum, because it suppressed the disease to the same extent as higher

    doses of the antagonist. This showed B. sphaericus UPMB10 has a high

    potential for its use as a plant growth enhancer and biocontrol agent in the

    disease management of Fusarium wilt in bananas.

  • vi

    Abstrak tesis yang dikemukakan kepada Senat Universiti Putra Malaysia sebagai memenuhi keperluan ijazah Master Sains

    APLIKASI BACILLUS SPHAERICUS UPMB10 UNTUK MENINGKATKAN

    PERTUMBUHAN PISANG (MUSA SPP. VAR. BERANGAN) DAN

    KESANNYA KE ATAS PENYAKIT LAYU FUSARIUM

    Oleh

    ILLANI ZURAIHAH IBRAHIM

    September 2008

    Pengerusi : Professor Zulkifli Hj Shamsuddin, Ph.D.

    Fakulti : Pertanian

    Strain bakteria tempatan Bacillus sphaericus UPMB10 yang dikategorikan

    sebagai rhizobakteria penggalak pertumbuhan pokok (PGPR) dilaporkan

    mempunyai keupayaan dalam menggalakkan pertumbuhan dan hasil dalam

    pelbagai tanaman pertanian. Kajian ini merupakan explorasi yang

    menunjukkan secara tidak langsung mekanisme yang dapat menyokong

    hipotesis PGPR ini untuk bertindak sebagai agen kawalan biologi. Melalui

    kaijan in vitro, B. sphaericus UPMB10 telah diperhatikan dapat merencatkan

    pertumbuhan Fusarium oxysorum f.sp.cubense race 4 (FOCR4) sebanyak

    70% di dalam medium kultur. Ketidakseragaman pertumbuhan hifa fungi yang

    disebabkan oleh pembentukan vakuol, pembengkakkan dan penebalan

  • vii

    jaluran hifa telah berlaku dengan kehadiran B. sphaericus UPMB10.

    Tambahan pula, B. sphaericus UPMB10 juga telah menunjukkan keputusan

    yang positif di dalam ujikaji penghasilan enzim pelupus dinding sel dan ujikaji

    aktiviti pelarutan fosfat dengan menunjukkan kesan zon jernih yang jelas atau

    halo. Ia juga berupaya menghasilkan hormon pertumbuhan iaitu indole asetik

    asid (IAA). Pada permulaan eksperimen rumah kaca menunjukkan yang

    inokulasi dengan B. sphaericus UPMB10 telah meningkatkan pertumbuhan

    pokok sebanyak 30% ketinggian pokok dan kadar pengambilan nutrient

    terutamanya N, P, K dan Ca. Ia juga telah mengurangkan kadar keterukan

    penyakit melalui pemerhatian simptom luaran pokok dan perwarnaan umbisi

    pokok. Pada akhir eksperimen, kadar insiden penyakit ditunjukkan melalui

    simptom luaran dengan menunjukkan kesemua rawatan yang melibatkan

    Fusarium telah dijangkiti walaupun dirawat dengan B. sphaericus UPMB10.

    Secara signifikannya, serangan hanya bermula lewat di akhir eksperiment

    dan ini menunjukkan keupayaan B. sphaericus UPMB10 untuk melambatkan

    serangan penyakit tersebut. Inokulum pepejal B. sphaericus UPMB10 telah

    menunjukkan perlambatan kemunculan simptom, peratus insiden penyakit

    dan kadar epidemik yang rendah. Aplikasi kepada tanah telah menunjukkan

    B. sphaericus UPMB10 dengan cara pepejal telah memberikan ketahanan

    terbaik kepada penyakit layu fusarium. Kombinasi samada menggunakan

    aplikasi substrat cecair atau pepejal telah dipilih untuk penilaian terhadap

    penyakit di eksperimen rumah kaca. Di dalam kajian ini, didapati bahawa

  • viii

    inokulum dengan formulasi 100 g pepejal per pokok adalah optimum untuk

    memberikan ketahanan terbaik pada tahap antagonis pada dos yang sama.

    Kesimpulan menunjukkan B. sphaericus UPMB10 berpotensi tinggi untuk

    digunakan sebagai penggalak pertumbuhan pokok dan agen kawalan biologi

    dalam pengurusan penyakit layu Fusarium pada tanaman pisang.

    .

  • ix

    ACKNOWLEDGMENTS

    All praises and thanks to Almighty Allah SWT, the beneficent and merciful for

    the blessing and the strength in completion of my study. Firstly, I would like to

    express my deepest and sincerest appreciations with a heartfelt gratitude to

    Professor Dr. Zulkifli Hj Shamsuddin, the Chairman of Supervisory Committee

    for his invaluable advice, constructive criticism, support, encouragement,

    patience and understanding that made this study possible. Not forgetting to

    Professor Dr. Sariah Meon and Associate Professor Dr. Zakaria Wahab as

    members of the Supervisory Committee, for their guidance, suggestions and

    invaluable advice and comments throughout the duration of this study and

    preparation of this thesis.

    Also with deep sense of honour, I wish to extend my sincere gratitude to all

    staffs of the Department of Land Management especially in the Soil

    Microbiology Laboratory and to Mrs Zarinah and Mr Dzulkifli Duaji for their

    time and assistance in the preparation of materials. Heartfelt thanks are also

    extended to my friends Wahida, Noraini, Maliza, Zakry, Prema, Negash, Ina,

    Palie, Yani, Ziana, Siti and others.

    I also would like to thank the Ministry of Science, Technology and Innovation

    (MOSTI) for the research grant no. 01-02-04-PR0010/04-02 and financial

  • x

    support through PASCA Scheme that provided me the opportunity to pursue

    my master degree.

  • xi

    I certify that an Examination Committee met on 25th September 2008 to conduct the final examination of Illani Zuraihah Ibrahim on her thesis entitled “Application of Bacillus sphaericus UPMB10 for Growth Enhancement of Banana (Musa spp.var. Berangan) and Its Effects On Fusarium Wilt“ in accordance with Universities and University Colleges Act 1971 and the Constitution of the Univeristi Putra Malaysia [P.U.(A) 106] 15 March 1998. The Committee recommends that the student be awarded Master of Science. Members of the Thesis Examination Committee were as follows: Zaharah Abdul Rahman, Ph.D. Professor Faculty of Agriculture Universiti Putra Malaysia (Chairman) Jugah Kadir, Ph.D. Associate Professor Faculty of Agriculture Universiti Putra Malaysia (Internal Examiner) Halimi Mohd Saud, Ph.D. Associate Professor Faculty of Agriculture Universiti Putra Malaysia (Internal Examiner) Nik Masdek Nik Hassan, Ph.D. Lecturer Horticultural Research Centre Malaysian Agricultural Research and Development Institute (External Examiner)

    BUJANG KIM HUAT, PhD Professor and Deputy Dean, School of Graduate Studies, Universiti Putra Malaysia. Date: 19 February 2009

  • xii

    This thesis submitted to the Senate of Universiti Putra Malaysia has been accepted as fulfilment of the requirement for the degree of Master of Science. The members of the Supervisory Committee were as follows:

    Zulkifli Hj. Shamsuddin, Ph.D. Professor, Faculty of Agriculture, Universiti Putra Malaysia. (Chairman) Sariah Meon, Ph.D. Professor, Faculty of Agriculture, Universiti Putra Malaysia. (Member) Zakaria Wahab, Ph.D. Associate Professor, Faculty of Agriculture, Universiti Putra Malaysia. (Member)

    HASANAH MOHD GHAZALI, Ph.D.

    Professor and Dean, School of Graduate Studies, Universiti Putra Malaysia. Date: 9 April 2009

    .

  • xiii

    DECLARATION

    I hereby declare that the thesis is based on my original work except for quotations and citations, which have been duly acknowledged. I also declare that it has not been previously or concurrently submitted for any other degree at UPM or other institutions.

    ___________________________

    Candidate: ILLANI ZURAIHAH IBRAHIM Date:

  • xiv

    TABLE OF CONTENTS

    PAGE iii iv vi ix xi xiii xvii xviii

    DEDICATION ABSTRACT ABSTRAK ACKNOWLEDGEMENTS APPROVAL DECLARATION LIST OF TABLES LIST PLATES LIST OF FIGURES LIST ABBREVIATION

    xx xxi

    CHAPTER

    1 INTRODUCTION 1

    2 LITERATURE REVIEW 2.1 Overview of banana industry 2.2 Fusarium wilt

    2.2.1 Systematics and Characterization of Fusarium oxysporum

    2.2.2 Characterization of Fusarium oxysporum f.sp.cubense

    2.2.3 Fusarium Wilt of Banana 2.2.4 Disease Distribution and Losses 2.2.5 Epidemiology 2.2.6 Current Management of Fusarium Wilt

    2.3 Beneficial Microbes as An Alternatives for Biocontrol 2.3.1 Bacillus sphaericus

    2.4 Mechanisms of Biocontrol by PGPR 2.4.1 Antibiosis 2.4.2 Antifungal Metabolites 2.4.3 Enzymes 2.4.4 Competition 2.4.5 Plant Ethylene Levels 2.4.6 Induced Systemic Resistance 2.4.7 Direct Stimulation of Plant Growth

    and Development 2.5 Microbial Inoculum Performance

    6 6 7 7 9

    10 13 15 17 19 21 24 25 25 27 28 32 33 35

    36

  • xv

    3 4

    MATERIALS AND METHODS 3.1 Morphological and Metabolite Production

    Characteristics, and Biological Activity of B. sphaericus UPMB10 3.1.1 PGPR Strain 3.1.2 Fungal Strain 3.1.3 Culture, Media and Growth Condition 3.1.4 Dual Culture Technique 3.1.5 Hyphal Morphology 3.1.6 Production of Cell-wall Degrading Enzyme 3.1.7 Test for Phosphate Solubilizing Activity 3.1.8 Production of Indole Acetic Acid (IAA)

    3.2 B. sphaericus UPMB10 Inoculation on Growth of Banana Seedlings and Pathogenecity Toward Fusarium oxysporum 3.2.1 Tissue-cultured Banana Plantlets 3.2.2 Preparation of Fusarium Inoculant 3.2.3 Plant Tissue Analysis 3.2.4 Plant Growth Measurement 3.2.5 Disease Assessment

    3.3 Infectivity Studies and Effect of Different Rates of B. sphaericus UPMB10 Inoculation on the Disease Incidence Caused by Fusarium oxysporum 3.3.1 Inocula Preparation and Application Rate 3.3.2 Plant Growth Performance (based on height

    increment) 3.3.3 Evaluation on Different Rates of Inoculum on

    Disease Assessment 3.4 Statistical Analysis

    RESULTS AND DISCUSSION 4.1 Morphological and Metabolite Production

    Characteristics, and Biological Activity of B. Sphaericus UPMB10 4.1.1 Dual Cultural Studies 4.1.2 Hyphal Morphology 4.1.3 Secondary Metabolite Production of B.

    sphaericus UPMB10 4.1.3.1 Test for Phosphate Solubilizing Activity 4.1.3.2 Production of Indole Acetic Acid (IAA) 4.1.3.3 Production of Cell Wall-degrading Enzymes

    38

    38 38 38 39 40 41 41 42 42

    43 43 43 44 44 45

    48 48

    49

    50 53

    54

    54 54 56

    60 60 61 63

  • xvi

    4.2 Evaluation of B. sphaericus UPMB10 Inoculation on Plant Growth Performance 4.2.1 Evaluation on Nutrient Status of Different Plant

    Components 4.2.1.1 Nitrogen (N) 4.2.1.2 Phosphorus (P) 4.2.1.3 Potassium (K) 4.2.1.4 Calsium (Ca)

    4.2.1.5 Magnesium (Mg) 4.2.2 Evaluation on Plant Height Development

    4.2.3 Evaluation on Disease Progress 4.3 Effect of Different Rates of B. sphaericus UPMB10

    Inoculation on Disease Incidence of Fusarium oxysporum 4.3.1 Evaluation and Comparison Between Liquid vs.

    Solid Inocula and Application Rate on Growth Performance

    4.3.2 Evaluation on Disease Assessment (Disease Incidence and Symptom Development)

    66

    73 73 76 77 78 79 79 83

    89

    92

    96

    5 GENERAL DISCUSSION AND CONCLUSION 96

    REFERENCES/BIBLIOGRAPHY APPENDICES BIODATA OF THE STUDENT

    105 131 139

  • xvii

    LIST OF TABLES

    No. Table

    Page

    1 S Some free-living rhizobacteria (PGPR) that are also plant growth-promotive. (Note: Not all strains of the microorganisms can act as PGPR)

    20

    2 Summary of beneficial and antagonistic characteristic of B. sphaericus UPMB10

    65

    3 The nutrient status in root, pseudostem, corm and leaves of banana plantlets inoculated with B. sphaericus UPMB10 after 12 weeks. (Means having the same letter (s) in a column of a parameter do not differ significantly at 0.05% level by Tukey Test)

    74

    4 Total nutrient accumulation in banana plantlets inoculated with B. sphaericus UPMB10 after 12 weeks. (Means having same letter (s) in a column do not differ significantly at 0.05 levels by Tukey’s Test)

    75

    5 Disease severity (vascular discoloration of corm) index (DSI) of banana seedlings var. Berangan cv. Intan challenged with FOCR4 at 90 days. Each value is the mean of five replicates

    86

    6 Area Under Disease Progress Curve (AUDPC) and epidemic rate of Fusarium wilt under different treatments

    94

  • xviii

    LIST OF PLATES

    No. Plate

    Page

    1 Inoculated banana plants under evaluation for tolerance to Fusarium oxysporum in a glasshouse.

    53

    2 B. sphaericus UPMB10 colony in a ring inhibiting the growth of FOCR4 on NA plate after 5 days of incubation

    55

    3 5mm agar disc of FOCR4 on NA plate as the control after 5 days’ incubation

    55

    4 A streak colony of B. sphaericus UPMB10 bacteria inhibiting the growth of FOCR4 on NA plate after 5 days of incubation.

    56

    5 (a) (a) Light microscopy shows abnormalities of FOCR4 hyphae with B. sphaericus UPMB10 in the culture medium; thickened and vacuolar hyphal strand.

    57

    5 (b) Normal fungal growth with smooth hyphal wall (no swelling and vacuoles) in absence of B. sphaericus UPMB10.

    58

    5 (c) Photo from Chaurasia et al., 2005 showed (A) Normal hyphae macroconidia of F.oxysporum , (B) swelling of hyphal tip and deformation of conidia in F.oxysporum

    58

    6 Clear zone around B. sphaericus UPMB10 cultured on chitin agar plate after 5 days of incubation

    63

    7 Comparison of inoculation effect of B. sphaericus UPMB10 (T2) with control (T5) after 4 weeks of treatments.

    68

    8 Effect of B. sphaericus UPMB10 inoculation on plant height of banana tissue cultured plantlet at six weeks. Left B. sphaericus UPMB10 (T2) and right control (T5).

    81

    9 Infected banana plant (left) showing foliar symptoms of Fusarium wilt caused by FOCR4 compared to healthy control plant (right).

    82

  • xix

    10 Infected banana plant showing the development of foliar associated symptoms of Fusarium wilt of banana on cultivar INTAN caused by Fusarium oxysporum f. sp. cubense. Healthy plant (A); chlorosis or yellowing of lower leaves (B); desiccation of lower leaf and chlorosis of upper leaves (C); advanced stage of infection with severe chlorosis and wilting (D).

    85

    11 Disease severity index, (based on vascular discoloration of corm caused by FOCR4): (a) uninfected control, (b, c) less severe (inoculated with UPMB10) and (d) most severe (FOC4); of banana seedlings var. Berangan cv. Intan challenged with FOC4 after 60 days.

    87

    12 Culture of banana pseudostem on PCNB agar. New Fusarium colonies formed from tissue of an infected plant (a) and no fungal proliferation from tissue of a healthy plant (b).

    88

  • xx

    LIST OF FIGURES

    No. Figure

    Page

    1 Effect of B.sphaericus UPMB10 inoculation on fresh weight of banana tissue culture plantlets (pseudostem and corm, roots, leaves) at 12 weeks of treatments. (Means having same letter (s) in a column of a parameter do not differ significantly at 0.05 level by Tukey’s Test)

    67

    2 Effect of B.sphaericus UPMB10 inoculation on dry weight of banana tissue culture plantlets (pseudostem and corm, roots, leaves) at 12 weeks of treatments. (Means having same letter (s) in a column of a parameter do not differ significantly at 0.05 level by Tukey’s Test)

    69

    3 The effect of B.sphaericus UPMB10 inoculation on root volume of banana tissue cultured after 12 weeks of treatments. (Means having same letter (s) between treatment not differ significantly at 0.05 level by Tukey’s Test)

    71

    4 Effect of different treatments of inoculation in plant height of banana plantlets. (Each value represent mean of 5 replicates). Vertical bars on the top indicate LSD for treatment comparisons at a given week of treatment. Vertical bars on the right indicate LSD for comparisons of growth height over weeks within a treatment.

    80

    5 Disease development on banana seedlings challenged with FOCR4 based on foliar symptoms. (Each value is the mean of 5 replicates).

    84

    6 Figure 6: Effect of different rates of B. sphaericus UPMB10 inoculation to FOCR4 challenge on growth height of banana plants. (Each value represent mean of 12 replicates). Vertical bars on the right indicate LSD for comparisons of growth height over weeks within a treatment

    90

    7 Effect of different treatments on the development of Fusarium wilt in banana based on foliar symptoms. Each value represent mean of 12 replicates.

    93

  • xxi

    LIST OF ABBREVIATIONS

    ANOVA Analysis of variance

    AUDPC Area under Disease Progress Curve

    cfu Colony forming units

    DI Disease incidence

    DS Disease severity

    DSI Disease severity index

    FOCR4 Fusarium oxysporum f.sp. cubense race 4

    FSM Fusarium selective media

    FW Fresh weight

    GA Gibberelic acid

    IAA Indole acetic acid

    M monit

    PDB Potato Dextrose Broth

    PGPR Plant growth promoting rhizobacteria

    PIRG Percentage of inhibition of radial growth

    PSB Phosphate solubilizing bacteria

    RCBD Randomised Complete Block Design

    rm Epidemic rate

    TSA Tryptic Soy Agar

    TSB Tryptic Soy Broth

  • CHAPTER 1

    INTRODUCTION

    Non-symbiotic rhizobacteria, associated with plant roots and able to

    enhance plant growth, known as plant growth promoting rhizobacteria

    (PGPR) has generated much interest in a wide range of research. It has

    impacts in different fields of study because it provides enormous information

    on different aspects of plant growth promotion and development while at the

    same time promises possible biological control solutions to pytopathogens of

    some plant diseases. At present, phytopathogens have caused a great loss in

    crop productivity and had to depend largely on chemicals, fumigation, steam-

    treatment and solarization of soils in order to control it (Gamliel and Katan,

    1992). Earlier, it was believed that the advantages of chemical controls of

    diseases could be obtained without any deleterious environmental effects.

    Currently, researchers have realized that these chemical agents are both

    hazardous to animals and humans because they can persist and accumulate

    in natural ecosystems. Consequently, there is a strong desire to replace

    chemicals with biological approaches that are potentially more “friendly” to the

    environment in the long term. These biological approaches include the use of

    transgenic plants, which are resistant to one or more plant pathogens

    (Greenberg and Glick, 1993), or either to use of biocontrol plant growth-

  • 2

    promoting bacteria which can suppress or prevent the plant damage

    (O’Sullivan and O’Gara, 1992; Sivan and Chet 1992; Sutton and Pegg, 1993;

    Cook, 1993). Biocontrol seems to be a reliable alternative compared to

    chemical fungicides, which have raised serious concerns of food

    contamination and environmental pollution. Some saprotrophic bacteria can

    serve as excellent biocontrol agents against plant pathogens (Fernando et al.,

    2005).

    The introduction of beneficial microbes as biological control of soil

    borne plant pathogen as well as enhancement of plant growth has a

    considerable potential in agriculture. Biological control using antifungal

    rhizobacteria to suppress plant diseases offers a powerful alternative to the

    use of synthetic chemicals. Numerous studies have demonstrated the ability

    of several bacteria to suppress diseases caused by soil-borne and seed-

    borne plant pathogens (Weller, 1988; O’Sullivan and O’Gara, 1992; Whipps,

    1997). The anti-fungal abilities of these beneficial microbes have been known

    since a few decades ago and there have been extensive efforts to use them

    for plant disease control and more recently on a commercialized scale.

    The uniqueness of microorganisms has never been understood until

    their beneficial functions were discovered. It is a paradox that bacteria can be

    both beneficial and antagonistic towards a plant pathogen to maintain its life

  • 3

    cycle. In this manner, it is possible to introduce the beneficial microorganism

    as a biological control for soil borne plant pathogen as well as enhancement

    of plant growth. This creates a great implication on plant growth development

    through direct or indirect mechanisms (Glick et al., 1999). A direct promotion

    of plant growth by PGPR is generally based on the ability of the bacteria to

    synthesize and facilitate nutrient uptake from the environment. Meanwhile

    indirect mechanism is through its effect on the phytopathogenic organism

    (usually fungus) by one or several mechanisms such as production of

    antibiotic and antifungal metabolite or enzyme that could lyse the fungal cell

    wall. It was believed that this advantage would lead to the potential of PGPR

    as a biocontrol agent besides stimulating plant growth. A local rhizobacterial

    strain, isolated from oil palm roots and identified as Bacillus sphaericus

    UPMB10 (B. sphaericus UPMB10), has shown great potential in promoting

    plant growth development and has been characterized for its optimum growth

    requirement, inoculum carrier (ground oil palm frond) and inoculum production

    technique (Premalalatha, 2006). Bacillus sphaericus is categorized as a free-

    living soil bacterium that can function as a plant growth promoting

    rhizobacteria (Kloepper et al., 1989). A few studies have demonstrated that

    inoculation with B. sphaericus UPMB10 enhanced plant growth and fixed

    nitrogen with oil palm (Amir et al., 2001 and banana (Mia et al., 2000). The

    current research interest is to assess the ability of B. sphaericus UPMB10 as