ISOLATION AND CHARACTERIZATION OF ANTIBIOTIC PRODUCING BACTERIA FROM SOIL RECEIVING ABATTOIR EFFLUENT IN UMUAHIA, ABIA STATE.

ABSTRACT

Soil samples were collected from different location within Ubakala Abattoir, Umuahia and serially diluted followed by pour plate technique to screen for antibiotic producing colonies. The media used for the screening were nutrient agar, Tryptic soy agar, Cysteine Lactose Electrolyte deficient agar and DeMan, Rogosa and Sharpe agar. Antibiotic- sensitivity test was performed on Mueller-Hinton agar and the inhibitory effect of the isolated microorganism was found out by screening them against pathogenic bacteria Staphylococcus aureus, Escherichia coli, Streptococcus spp, Proteus spp, Klebsiella spp, Salmonella spp, and Pseudomonas spp. Characterization was done by performing various biochemical tests and staining methods. The antibiotic producing bacteria isolated were Bacillus spp, Lactobacillus spp, Proteus spp and Micrococcus spp.  

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

Objectives 2

Background of Study 3

CHAPTER TWO: LITERATURE REVIEW

2.1 Soil 6

2.2 Soil Colour 6

2.3 Soil Types 7

2.4 Physiochemical parameters of soil 8

2.5 Soil Bacteria 10

2.6 Antibiotics 12

2.6.1 Antibiotic producing Bacteria 13

2.7 Classification of Antibiotics 14

2.8 Production of Antibiotics 15

2.9 Abattoir 16

2.9.1 Activities that occur in abattoirs 17

2.9.2 Abattoir Waste 18

CHAPTER THREE: MATERIALS AND METHODS

3.1 Collection and preparation of soil samples 19

3.2 Isolation of the microorganism 19

3.3 Identification of Bacteria 20

3.3.1 Gram staining 20

3.3.2 Spore staining 21

3.3.3 Motility test 21

3.4 Sub-culturing 21

3.5 Biochemical tests 21

3.6 Antibiotic sensitivity tests 23

CHAPTER FOUR: RESULTS

CHAPTER FIVE: DISCUSSION AND CONCLUSION

5.1 Discussion 32

5.2 Conclusion 34

REFERENCES

APPENDIX

LIST OF TABLES

Table Title Page

1 Some Antibiotics 4

2 Different physiochemical parameters of soil 9

3 List of common soil bacteria 11

4 Mean Colony Count 28

5 Morphology and Biochemical Characterization 28

of the Bacterial Isolates from Ubakala abattoir

soil samples that showed Clear Zone of Inhibition

6 Microorganisms isolated from abattoir receiving 29

soil effluent, Ubakala and their % occurrence

7 Antibiotic producing bacteria isolated from soil 30

receiving abattoir effluent, Ubakala

8 Antibacterial Activity of Isolated Bacterial 31

Strains from Ubakala abattoir against different

Pathogenic strains of Bacteria

CHAPTER ONE

1.0 INTRODUCTION

Soil refers to the outer loose material of the earth crust. It is a primary source of microorganisms. The numbers and species of microbes in soil is dependent on environmental conditions like nutrient availability, soil texture, presence of moisture in soil and type of vegetation cover and their number varies according to the type of environmental conditions (Atlas and Bartha, 1998). The presence of microorganisms like bacteria, fungi and Actinomycetes help to decompose the dead and decaying organic matter in soil into simple molecules which they use for their nutrition. From ancient times, is well understood that natural products have a key role in the discovery and development of many antibiotics (Newman and Cragg, 2007). One of the best approaches to the discovery of new antimicrobial agents from natural sources has been to use folklore or historical records to guide the collection of samples or a good research work on the soil of that area (Cordell et al., 1994).

There are many different potential sources where antibiotics can be discovered. However, the soil remains the most important target for most researchers in their efforts to discover novel antibiotics which have pharmaceutical values. This is because many microbes especially bacteria that reside in the soil have the ability to produce biologically active secondary metabolites such as useful antibiotics. Antibiotics have immense importance for humans as they help fight many bacterial infections and diseases. Bacillus and Actinomycetes are predominantly present in the soil and are known for producing inhibitory substances like Bacitracin and Actinomycin respectively (Abdulkadir and Waliyu, 2012). Streptomyces griseus is also an important soil microorganism which produces streptomycin that fights several pathogenic bacteria like Mycobacterium tuberculosis (Thakur et al., 2007). The growing resistance of pathogens towards already available antibiotics has posed as major problem (Eddie, 2013).

Antibiotics are one of the important pillars of modern medicines (Ball et al., 2004) but old antibiotics lose their efficacy and are necessarily replaced with new ones for many species of pathogenic bacteria (Hancock, 2007). Microorganisms that are able to produce secondary metabolites have a diverse chemical structure and biological activities and are produced only by some species of a genus Bacillus (Stachelhaus et al., 1995).

Effluent is defined by the United States Environmental Protection Agency as ‘waste water treated or untreated that flows out of a treatment plant, sewer or industrial outfall. Generally, refers to waste discharged into surface waters’ (EPA, 2006). The abattoir is a specialized facility approved and registered by the regulatory authority for inspection of animals, hygienic slaughtering, processing and effective preservation and storage of meat products for human consumption (Alonge, 2001). Waste water or effluent generated from the abattoir is characterized by the presence of a high concentration of whole blood of slaughtered food animals and suspended particles of semi-digested and undigested feeds within the stomach and intestine of slaughtered and dressed food animals (Cooker et al., 2001).

1.2 Objectives

A.) Isolate antibiotic-producing organisms from soil receiving abattoir effluent from Ubakala abattoir in Umuahia, the capital city of Abia State, Nigeria.

B.) Examine the effects of different antibiotic-producing organisms on known microbes.

C.) Quantify the sensitivity of known organisms to antibiotics.

D.) To determine the variety of organisms that live in soil.

1.3 Background of Study

Antibiotics have made it possible for people in the 20^th century to live longer, healthier lives than previous generations. Antibiotics also are now used in agriculture to prevent infection, a practice that some believe has led to an increase in the number of drug resistant organisms.

Antibiotics from the term anti-bios or ‘against life’ are compounds that will either inhibit growth (bacteriostatic) of cell or kill (bactericidal) the cell by blocking an essential cellular function such as transcription, translation or cell wall synthesis.

Table 1: Some Antibiotics

(Source: AHDE, 2011)

Many antibiotics have been isolated from soil microbes, the most important Genera of these being the prokaryotes Streptomyces and Bacillus and the eukaryotes (molds) Penicillium and Cephalosporin. Soil bacteria can be rod (bacilli), cocci (spherical), and spirilla (spirals), of these, Bacillus are more numerous than the others. They are one of the major groups pf soil bacteria population and are very widely distributed (Bhagabatiet et al., 2004). The number and type of bacteria present in a particular soil would be greatly influenced by geographical location such as soil temperature, soil type, soil pH, organic matter contents, cultivation, aeration and moisture content (Davies et al., 1999).

Historically, the Streptomyces and related Actinomyces were thought to be related to fungi because they do not grow as single cells but instead produce long rope-like mats called mycelium (made up of individual filaments called hyphae). Hyphal growth begins underground or in an agar and later aerial hyphae are produced which will contain spores. Streptomyces looks like stringy, whitish material in the soil. Further, the earthy, fresh soil smell is due in part to compounds such as geosmin that the Streptomyces produce. Streptomycete is responsible for the formation of more than 60% of known antibiotics (Williams et al., 1993). Actinomyces were originally considered to be an intermediate group between bacteria and fungi but now are recognized as prokaryotic organisms which are widely distributed in soil, water and found colonizing plants (Jensen and Fenical, 2000; Ogunmwonyi, 2008).

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