Antimicrobial Activity of Azadirachta Indica (Neem) Against Bacteria Isolated from Urinary Tract Infected Patients in Khartoum State

Abstract This was analytical study conducted during the period from January to April 2017, to determine the antimicrobial activity of Azadirachta indica leaves methanolic extract against different bacteria isolated from urinary tract infection. A total of 130 urine samples were collected from patients with Urinary tract infection. These specimens were inoculated into Cystine Lactose Electrolyte Deficiency (CLED) media and incubated aerobically at 37 C for 24 hrs. The clinical isolates were then identified using conventional method. Ninety seven (75%) samples showed bacterial growth. The antibiotic susceptibility testing was performed using standard disk diffusion method. The results showed that Ceftazidime had higher resistant rates 70.6% followed by Ciprofloxacin 64.7%, Cotrimoxazole 62.7%, Gentamycin 54.9% and Imipenem 15.7 %. A. indica leaves methanolic extract showed antibacterial activity against all pathogenic and reference strains, 50 (% w/v) concentration was the most effective one, but 100 (% w/v) didn‟t produce any effect. Gram negative bacteria were more susceptible to A. indica leaves methanolic extract than Gram positive bacteria. MIC of Neem leaves methanolic extract for all tested bacteria was 3.125 (% w/v) except for E. coli ATCC 25922 which was 12.5 (% w/v). Gas chromatography analysis revealed that 45 chemical active compounds of A. indica (Neem), 29 of them have antibacterial activity.

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I األٚــــــــــــــــت

Dedication II

Acknowledgement III

Abstract (English) IV

Abstract (Arabic) V

Table of content VI

List of figure VII

List of table VIII

CHAPTER ONE: INTRODUCTION

1.1. Introduction 1-2

1.2. Rational 3

1.3. Objectives 4

1.3.1. General objective 4

1.3.2. Specific objectives 4

CHAPTER TOW : LITERATURE REVIEW

2. Literature review 5

2.1. Azadirachta indica (Neem) 5

2. 2. Classification of A. indica 5

2. 3. Origin of the tree 6-7

2. 4. Botanical description 7

2. 5. Neem general features 8

2. 6. Secondary metabolites 8-9

2. 7. Extraction of the secondary metabolites 9

2. 8. Medicinal usage 10-11

2. 9. Melluscicidal effect 12

2. 10. Other uses 13

2. 11. Dosing 13

2. 12. Neem benefits and sides effects 13

VII

2. 13. Toxicity 13

2. 14. Urinary tract infections (UTIs) 14

2. 15. Recurrent urinary tract infection 14

2. 16. Incidence 14-15

2. 17. Risk factors 15

2. 18. Most common bacteria that cause urinary tract infections 15

2. 18. 1. Escherichia coli (E. coli) 15-16

2. 18.2. Pseudomonas aeruginosa (P. aeruginosa) 16

2. 18. 3. Klebsiellae pneumoniae (K. pneumoniae) 17

2. 18. 4. Proteus species 17

2. 18. 5. Citrobacter species 17

2. 18. 6. Enterococcus feacalis (E. faecalis) 18

2. 18. 7. Staphylococcus aureus (S. aureus) 18

2. 19. Multidrug resistant (MDR) pathogens 19

2. 20. Classification of MDR pathogens 19

2. 20. 1. Primary Resistance 19

2. 20. 2. Secondary resistance 19-20

2. 20. 3. Clinical resistance 20

Chapter three: material and method

3.1. Study design 21

3.2. Study area 21

3.3. Study population 21

3.4. Study period 21

3.5. Sample size 21

3. 6. Collection and proceeding of urine samples 21-22

3.7. Identification of the clinical isolates 22

3. 7. 1. Colonial morphology 22

3. 7. 2. Preservation and storage of isolated organisms 22

VIII

3.7. 3. Gram stain 22-23

3. 7. 4. Biochemical tests 23-24

3. 8. Antibacterial susceptibility testing 24

3. 9. Collection and identification of plant material 24

3. 10. Preparation of A. indica extract 24-25

3.11. Antibacterial activity of A.indica 25

3. 11. 1. Cup plate method 25

3.11. 2. Determination of minimum inhibitory concentration (MIC)

0f Neem extract by agar diffusion method

25

3. 12. Quality control 25-26

3. 12. 1. Control of culture media 26

3. 12. 2. Control susceptibility testing method 26

3. 12. 3. Batch quality control 26

3. 13. Photochemical screening of A. indica 27

3. 13. 1. Method of analysis 27

3. 14. Statistical Analysis 27

3. 15. Ethical consideration 27

CHAPTER FOUR : RESULTS

4. 1. Gender 28

4. 2. Bacterial growth 29

4. 3. Growth of bacteria on culture media 30

4. 4. Gram stain 30

4. 5. Biochemical tests of different bacterial isolates 30-31

4. 6. Antibacterial Susceptibility 32

4. 7. Antibacterial activity of A. indica 36

4. 8. Interpretation of results 36

4. 9. Minimum Inhibitory Concentration (MIC) of A. indica

methanol extract obtained by agar diffusion method

38

4.10. Gas chromatography results 40

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CHAPTER FIVE: DISCUSSION

5.1 Discussion 42-44

5.2 Conclusion 45

5.4 Recommendations 46