ISOLATION AND CHARACTERIZATION OF MICRO ORGANISMS FROM SOIL SAMPLES

ABSTRACT

Soil is the major repository of microorganism that clinically useful antibiotics have been isolated for several group of microorganism including bacteria (Streptomyces sp, Bacillus sp). Bacillus and Actinomycetes are the most abundant microorganism present in Soil and inhibit the growth of other microorganisms. Present in soil and inhibit the growth of other microorganism. In this study 5 Soil samples from different ecological sites were obtained and Cultured in microbiological media to isolate microorganisms. Ten fold serial dilution of the Soil samples were prepared and inoculated on nutrient agar and Saboraud Dextrose agar. A total of 15 Isolated were recovered. Ten of the total isolated were sub cultured on nutrient agar and incubated at 37oc for 24hrs. Out of these ten isolates four are Micrococcus, four Bacillus species and two Actinomycetes were obtained. Five different isolates was tested against four test organisms. Namely: Staphylococcus aureus, Pseudomonas aeruginosa, Mycobacterium semgmatis and Escherichia coli   for antibacterial activity. Out of the five isolates, only two (NDDCfemale Hostel) exhibitory effect against one of the test organisms (Pseudomonas aeruginosa) with zones of inhibition of 9mm and 13mm respectively. The remaining isolates showed no antimicrobial activity against the test organisms. Therefore, it is confirmed that soil samples for microorganisms collected within the University Premises of Umudike were found to be potential source of OR one of the isolates (NDDC Hostel) that is Actinomyces was able to inhibit growth of Staphylococcus aureus with a clear inhibition range with a clear inhibition range 0-14mm, which indicates an antibacterial activity against S. aureus while others showed no antimicrobial activity against the test organisms. This shows that the isolation of (NDDC Hostel) from this study area may contribute to the discovery of with further extraction and purification of the isolate.

TABLE OF CONTENTS

Title Page                                                                                                                    i

Certification                                                                                                                ii

Dedication                                                                                                                  iii

Acknowledgements                                                                                                    iv

Table of Contents                                                                                                       v

List of Tables                                                                                                              vi

Abstract                                                                                                                      vii

CHAPTER ONE:  INTRODUCTION                                                                   1

1.1    Aims And Objective                                                                                         2

CHAPTER TWO LITERATURE

2.1            Soil Description and Classification                                                            4

2.2            Microbial Diversity and Sample Strategy                                                  4

2.3            Types of Microorganisms Found In the Soil                                              5

2.3.1         Fungi                                                                                                          5

2.3.2         Algae                                                                                                          6

2.3.3         Protozoa                                                                                                     6

2.4            Sample Collection and Preparation                                                            7

2.5            Fermentation Conditions                                                                            8

2.6           Soil Antimicrobial Agents Producing Microbes                                          9

2.7        Microbial Diversity In Soil                                                                             11

2.8       Micro-Organisms As Sources Of Natural Products                                         13

2.9       Causes of improper soil disposal                                                                     14

CHAPTER THREE: MATERIALS AND METHODS

3.1       Collection an Preparation of soil sample                                                         15

3.2       Media Used                                                                                                    15

3.3       Sterilization of Materials                                                                                15

3.4       Samples Preparation                                                                                        16

3.4.1   Culturing                                                                                                          16       

3.4.2   Colonial Examination                                                                                      16

3.4.3   Sub-culturing                                                                                                   17

3.5       Gram Staining                                                                                                 17

3.6       Biochemical Tests                                                                                           17

3.6.1.   Starch Hydrolysis                                                                                          17

3.6.2    Catalase Tests                                                                                                 18

3.6.3    Citrate Utilization Test                                                                                   18

3.6.4    Coagulase Test                                                                                                19

3.6.5    Oxidase Test                                                                                                   19

3.7       Physicochemical Analysis                                                                               19

3.8       Additional Test                                                                                               20

3.9       Biochemical Identification of Organisms Isolates                                          20

CHAPTER FOUR: RESULTS                                                                               21

CHAPTER FIVE: DISCUSSION, CONCLUSION AND RECOMMENDATION

5.1              Discussion                                                                                                       28

5.2       Conclusion                                                                                                      29

5.3       Recommendation                                                                                                        29

References                                                                                                      30

LIST OF TABLES

Table                                                     Title                                                              Page

1      Collection of Soil Samples from Different Location in MOUAU                     22

2     Physico-Chemical Properties of Soil Sample of the Different Locations           

        (Makut, 2011)                                                                                                     23

3    Microbial Count of Soil Samples                                                                          24

4    Percentage Occurrence of Microbial Isolated From Soil Sample in MOUAU     25

5   Biochemical Test Characterization of Isolated Bacterial from Soil

      Sample in MOUAU                                                                                              26

6  Cultural and Morphological of fungi identification                                               27

CHAPTER ONE

1.0    INTRODUCTION

The term soil refers to the outer loose material of the earth crust it maybe regarded as a three phase’s system composed of solids, liquids and dispersed to form a heterogeneous matrix. On the whole soil is composed of five major components, these includes; mineral water, matter, organic matter, air and living organisms. The various component of the soil environment components constantly changed and the quantity of these constituents are not the same in all soil body includes. Small animals and microorganism but is generally considered that it’s in microorganisms that plays the most important role in the release of Nutrient and Carbondioxide for plant growth. (Hamlon et al., 2007). The bacteria are the most abundant group usually more numerous than the four combined. Soil bacteria can be rod (bacilli) cocci (Sphereical) spirilla of these bacillus, are more numerous than the others. They are one of the major groups of soil bacteria population and are very widely distributed (Bhagabati el al., 2004) which can be found as single cells or micro colonies development around the soil particles (Torsuik,2002) they are responsible for many of the key processes in the biogeochemical cycling on earth such as nitrogen, sulphr and carbon cycle (Lenyeler et al., 1999) in addition they are a valuable sources of natural products providing important antibiotic for pharmaceuticals, important enzymes and bioactive compounds for industries (Strohl, 2000). The number and types of bacteria present in a particular soil would be greatly influenced by geographical location such as soil temperature, soil type, soil pH, organic matters contents, cultivation, aeration and moisture content (Davies et al., 1999). An ecological niche is composed of many microhabitats; each microhabitat is composed of a microscopic diversity which includes bacteria, protozoa, fungi, and macroscopic diversity that that includes plant and insects. Soil is a complex medium in which one can encounter many kinds of microbial communities. Application of nucleic acid-based techniques to analyze soil microbial communities has revealed high prokaryote diversity (Pandey et al., 2008).  The microbial diversity or communities present in soil principally depend on the composition of the soil and many physical chemical properties that the medium posses. Also the flora and decomposing organism matter on the surface of the biowill influence microbial diversity present. For example, the fallen trees, barks and flowers provide nutrients both the microbes and plant present, through microbial degradation of carbohydrates, lipid and proteins to sugars, fatty acids, glycerol and amino acids and respectively to mineralization. Besides providing these nutrients plant secondary metabolites that are generally toxic to microorganisms will need to be degraded or detoxified by certain microbes. These degraders (microbes) are selectively presented and ultimated evolved to produce novel secondary metabolites possibly to counteract the toxic plant secondary metabolites (Pandey et al., 2008). There are approximately 10⁶ -10⁹ colony forming unit per gram of soil. Microbes present in medium posses advantages that will permit or facilitate their survival in that medium. For examples research has demonstrated that in desert crust or his soil that has low water availability, gram positive and spore farm microbes are most abundant. The gram positive bacteria posses a thicker layer of Murein in their cell wall which makes the cell less vulnerable to the limiting conditions present in these habitat. Also, spore farming bacteria can resist long period of desiccation and limiting nutrient conditions smiles they compact and protect their genuine material in the bacteria spore, until conditions are favorable for sporulation to occur (Sadha et al.,2000).

1.1     Aims and Objective

1.       To isolate, identify and characterize bacteria from the soil sample.

2.       To isolate, identify and characterize fungi from the soil sample.

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