Isolation Of Bacillus Specie And Production Of Bacitracin

ABSTRACT

Bacillus species was isolated from soil samples using spread plate method on nutrient agar while peptone water was used as the production medium. The production of bacitracin was authenticated by antimicrobial activity against Klebsiella pneumoniae, Escherichia coli, Streptococcus pyogenes, Staphylococcus aureus and Pseudomonas aeruginosa. This was done using agar well diffusion method. After the production of bacitracin by Bacillus species, the minimum bactericidal concentration of the produced bacitracin was evaluated, using 70%, 60%, 50%, 30% and 20% each of the diluted bacitracin aliquot on Klebsiella pneumoniae, Escherichia coli, Streptococcus pyogenes, Staphylococcus aureus and Pseudomonas aeruginosa. The zone diameter against the respective was calculated in duplicates and the mean diameter was obtained. Highest zone inhibition diameter was observed against Streptococcus pyogenes with a mean inhibition diameter of 10mm. Next in line was Staphylococcus aureus with a mean inhibition diameter of 9mm. After this came Escherichia coli with a mean inhibition diameter of 8mm, Klebsiella pneumoniae with a mean inhibition diameter of 7mm and lastly, Pseudomonas aeruginosa with a mean inhibition diameter of 6mm.  For Escherichia coli, the MIC was at 50% and minimum bactericidal concentration was at 50%. For Pseudomonas aeruginosa MIC was at 60% and minimum bactericidal concentration percentage was at 60%. For Staphylococcus aureus MIC was at 30% and MBC was at 30%. For Klebsiella pneumoniae, MIC was at 60% while MBC was at 60%. For Streptococcus pyogenes, MIC was at 30% while MBC was at 30%. Bacitracin is very effective against Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Streptococcus pyogenes and Escherichia coli and as such can be used effectively in the treatment of diseases caused by these organisms.

TABLE OF CONTENTS

Certification ﾿ ii

Dedication ﾿ iii

Acknowledgements ﾿ iv

Table of Contents ﾿ v

List of Tables ﾿ viii

Abstract                                                                                                                                 ix ﾿

Chapter One

1.0 ﾿        Introduction…………………………………………………………………..1

1.1 ﾿ Aim …………………………………………………………………………..2

1.2 ﾿ Objectives………………………………………………………………….2

Chapter Two

2.0 ﾿        Literature review……………………………………………………………3

2.1 ﾿ Bacitracin…………………………………………………………………….3

2.1.1 ﾿ History ﾿ 3

2.1.2 ﾿ Synthesis ﾿ 3

2.1.3 ﾿ Composition ﾿ 4

2.1.4 ﾿ Spectrum of activity and susceptibility data ﾿ 4

2.1.5 ﾿ Mechanism of action ﾿ 4

2.1.6 ﾿ Clinical use ﾿ 5

2.2 ﾿ Bacillus subtilis ﾿ 9

2.2.1 ﾿ Habitat ﾿ 9

2.2.2 ﾿ Reproduction ﾿ 10

2.2.3 ﾿ Chromosomal replication ﾿ 10

2.2.4 ﾿ Genome ﾿ 11

2.2.5 ﾿ Transformation ﾿ 12

2.2.6 ﾿ Uses ﾿ 14

2.2.7 ﾿ Safety ﾿ 15

2.2.7.1 ﾿ In humans ﾿ 16

2.2.7.2 ﾿ In animals ﾿ 17

Chapter Three

3.0 ﾿ Materials and methods ﾿ 17

3.1 ﾿ Source of material and collection ﾿ 17

3.2 ﾿ Preparation of culture media ﾿ 17

3.3 ﾿ Sterilization of materials ﾿ 17

3.4. ﾿ Isolation of bacillus specie ﾿ 18

3.5. ﾿ Identification of the isolate ﾿ 18

3.5.1 ﾿ Morphological appearance ﾿ 18

3.5.2 ﾿ Gram reaction ﾿ 18

3.5.3 ﾿ Endospore stain ﾿ 19

3.5.4 ﾿ Biochemical characteristics ﾿ 20

3.5.5    Growth at 550C ﾿ 20

3.6. ﾿ Screening for crude bacitracin production by Bacillus specie ﾿ 20

3.6.1. ﾿ Antimicrobial testing of bacitracin produced by Bacillus specie ﾿ 20

3.7. ﾿ Minimum bactericidal concentration ﾿ 21

Chapter Four

4.0. ﾿ Results ﾿ 30

Chapter Five

5.1. ﾿ Discussion ﾿ 31

5.2. ﾿ Conclusion ﾿ 32

References ﾿ 34

Appendix