ISOLATION AND SCREENING OF ANTIBIOTIC PRODUCING MICROORGANISMS FROM FOREST SOIL IN MICHAEL OKPARA UNIVERSITY OF AGRICULTURE, UMUDIKE (MOUAU)

ABSTRACT

The soil environment is rich in microorganisms capable of antibiotics production but the rate of occurrence and distribution of these microorganisms at ecologically significant levels has not been much clear. In the past few decades, microorganisms have been demonstrated as synthsizers of varieties of antibiotics even under field conditions in the rhizosphere. In the present work, soil smples were collected randomly from the forest environment in Michael Okpara University of Agriculture, Umudike, in Abia-state, Nigeria and were screened for antibiotics-producing microorganisms by the agar sensitivity assay using paper discs impregnated with the isolates from the forest soil. Forty-eight (48) isolates were gotten on the total, out of which thirteen 13 (27.1%) isolates active against different test organisms (Salmonella typhi , Staphylococcus aureus ATCC 25923TM, Escherichia coli ATCC 25922TM, Pseudomonas aeruginosa ATCC 27893TM). Nine (9) (18.8%) bacterial isolates were active against the test pathogenic bacteria and they were identified by their cellular characteristics, colonial morphology and biochemical tests. These active isolates sort into the Genus Bacillus and Micrococcus and the ones that were not fully characterized yet are probably members of the Streptomyces genera. Four (4) (8.3%) fungal isolates were also active against the test bacteria and were identified based on their fungal morphology as probably Penicillium spp. and Aspergillus spp. Of all the isolates, isolate 36 which is probably a Streptomyces spp., has the highest range of antibacterial activity with 12mm zone of inhibition on Pseudomonas aeruginosa, 12mm on Staphylococcus aureus 5mm on Salmonella typhi and 4mm on Escherichia coli. Isolates 35 and 40 had the highest zones of inhibition, both on Salmonella typhi. The present work has once more contributed to the existing truth in cases of isolation of antibiotics-producing microorganisms from the forest soil and promises to contribute hugely in all enquiries into the antibiotics producing micro-flora in the forest environment.

TABLE OF CONTENTS

Cover Page ﾿ i

Title Page ﾿ ii

Certification Page ﾿ iii

Dedication ﾿ iv

Acknowledgement ﾿ v

Table of Contents ﾿ vi-viii

List of Tables    ﾿                         ﾿ ix

Abstract ﾿ x ﾿

CHAPTER ONE

1.0 ﾿ INTRODUCTION ﾿ 1

1.1 ﾿ Factors Affecting Soil Microbial Composition ﾿ 1

1.2 ﾿ Definition of Antibiotics ﾿ 1

1.3 ﾿ Bacillus spp. and antibiotics ﾿ 3

1.4 ﾿ Aims and Objectives ﾿ ﾿ 4

CHAPTER TWO

2.0 ﾿ LITERATURE REVIEW ﾿ 5

2.1` ﾿ Soil and Antibiotics ﾿ 5

2.2 ﾿ Review on Different Soil Environment and Their Antibiotics-Producing Microbes Composition ﾿

2.3 ﾿ Fungi and Antibiotics ﾿ 7

2.4 ﾿ Sources of Nutrients for Soil Microbes ﾿ 9

2.5 ﾿ Formation of Antibiotics in the Soil ﾿ 9

2.6 ﾿ Factors Influencing Natural Stimulation of Secondary Metabolite Production ﾿ 9

2.7 ﾿ Class: Actinomycetales ﾿ 10

2.7.1 ﾿ Actinomycetes ﾿ 10

2.8 ﾿ Estimation of Soil Microbial Flora ﾿ 11

2.9 ﾿ Distribution of the Actinomycetes in the Soil ﾿ 13

2.10 ﾿ Classification of Antibiotics ﾿ 13

2.11 ﾿ Streptomyces: Wide-range Antibiotics Producers ﾿ 14

2.12 ﾿ Ecological and Geographical Distribution of Microorganisms in Soils ﾿ 14

2.13 ﾿ Problems Encountered In Determining Soil Microbial Distribution ﾿ 14

2.14 ﾿ Factors Encouraging Antibiotics Production by the Streptomycetes ﾿ 14

2.15 ﾿ Effects of Isolation Media in Antibiotics Production ﾿ 16

2.16 ﾿ Techniques in Identifying Antibiotic Producers in Media Plates. ﾿ 16

2.17 ﾿ Failure of In-use Antibiotics and the need for Novel Antibiotics ﾿ 16

2.18 ﾿ Concentration, Inactivation, pH and Adsorption as Factors Affecting Effectiveness of Antibiotics in the Soil ﾿

2.19. The Need for Specific Isolation Media in Screening for Antibiotics Production ﾿ 18

CHAPTER THREE

3.0 ﾿   MATERIALS AND METHODS ﾿ 20

3.1 ﾿ Study Area and Period ﾿ 20

3.2 ﾿ Collection of Samples ﾿ 20

3.3 ﾿ Sterilization of Equipment and Preparation of Media ﾿ 20

3.4 ﾿ Preparation of Sample ﾿ 21

3.5. ﾿ Sample Inoculation, Incubation and Isolation of Microbes ﾿ 21

3.6 ﾿ Colonial Examination ﾿ 22

3.7 ﾿ Subculturing and Storage of Isolates ﾿ 22

3.8 ﾿ Testing for Antibiotic Production ﾿ 22

3.9 ﾿ Gram Staining ﾿ 22

3.10 ﾿ Biochemical Tests ﾿ 23

3.10.1 ﾿ Starch Hydrolysis Test ﾿ 23

3.10.2 ﾿ Hydrogen Sulphide Test ﾿ 23

3.10.3 ﾿ Sugar Utilization Test ﾿ 24

CHAPTER FOUR

4.0 ﾿ RESULTS ﾿ 25

CHAPTER FIVE

5.1 ﾿ Discussion ﾿ 32

5.2 ﾿ Recommendations ﾿ 34

5.3 ﾿ Conclusion ﾿ 35

﾿ References ﾿ 36