ANTIBIOTIC SUSCEPTIBILITY PATTERN OF ESCHERICHIA COLI ISOLATED FROM WASTE WATER IN UMUAHIA

ABSTRACT

The antibiotic susceptibility pattern of E.coli in waste water from Umuahia Metropolis was investigated. A total of 17 (85%) E.coli were isolated from the waste water samples. All 17 isolates were subjected to antibiotics susceptibility test using Kirby Bauer disc diffusion method. A total of 12 antibiotic discs namely; Ceftazidime (30mg), Cefuroxime (30mg), Gentamicin (10mg), Ciprofloxacin (5mg), Ofloxacin (5mg), Augumentin (30mg), Nitrofurantoin (300mg), Ampicillin (10mg) Imipenem (10mg), Menopenem (10mg), Aztreonam (30mg) and Ceftriaxone (30mg) were used. The E.coli isolates showed highest susceptibility to both Gentamicin and Ofloxacin (41.1%), followed by Ceftriaxone 76.5%. Also 17 (100%) of the isolates were found to be multi drug resistant (MDR). The E.coli isolates were screened for ESBLs in accordance to the CLSI breakpoint and 17(100) were positive. On subjecting the 17CLSI positive isolates to double disc synergy test (DDST), 12 (70.6%) were confirmed to be ESBL producers. This result shows that waste waters found in Umuahia Metropolis could serve as a reservoir for antibiotic resistant E.coli.

TABLE OF CONTENTS

CHAPTER ONE

INTRODUCTION

1.1 Aims and Objectives

CHAPTER TWO

2.1     Municipal Waste Water

2.1.1   Nature, growth and survival characteristics of E. coli

2.1.2   Epidemiology of E. coli

2.1.3   Pathogenicity, virulence and clinical features

2.1.3.1 ETEC (Enterotoxigenic E. coli)

2.1.3.2 EPEC (Enteropathogenic E. coli)

2.1.3.3 EHEC (Enterohemorrhagic E. coli) or VTEC (Vero cytogenic E. coli)

2.1.3.4 EIEC (Enteroinvasive E. coli)

2.1.3.5 EAEC (Enteroaggregative E. coli)

2.1.3.6 DAEC (Diffusely adherent E. coli)

2.1.3.7 UPEC (Uropathogenic E. coli)

2.1.3.8 MAEC (Meningitis-associated E. coli)

2.1.4   Clinical syndromes

2.1.5   Detection of E. coli

2.1.6   Antibiotic considerations and multi-drug resistance in E. coli

2.2      Antimicrobial agents

2.2.1   Major classes of antibacterial drugs

2.2.2   Mechanism of action of antibiotics

2.2.3   Mechanism of antibiotics resistance in enterobacteriacae

CHAPTER THREE

MATERIALS AND METHOD

3.1      Study area

3.2      Collection of samples

3.3      Sterilization of Materials

3.4      Media Used and Their Preparation

3.5      Microbiological Analysis of Samples

3.5.1   Serial Dilution

3.5.2   Isolation of E. coli  

3.6      Characterization and Identification of the Bacterial Isolates

3.6.1   Purification and storage of the isolates

3.6.2   Colonial morphology

3.6.3   Motility test

3.6.4   Gram staining

3.6.5   Biochemical tests

3.6.5.1 Indole test

3.6.5.2 Methyl red test

3.6.5.3 Voges Proskauer test

3.6.5.4 Catalase test

3.6.5.5 Citrate utilization test

3.6.5.6 Oxidase test

3.6.5.7 Ornithine decarboxylase test

3.6.5.8 Triple sugar iron test

3.6.5.9 Lysine decarboxylase test

3.6.5.10 Urease test

3.7      Standardization of inoculum

3.8      Antibiotic Susceptibility Testing

3.9      Clinical Laboratory Standard Institute (CLSI) Breakpoint Test for ESBLs Screening

3.10    Double disc synergy test (DDST) for ESBL confirmation

CHAPTER FOUR

RESULTS

CHAPTER FIVE

DISCUSSION, CONCLUSION AND RECOMMENDATION

5.1      Discussion

5.2      Conclusion

5.3      Recommendation

REFERENCES

APPENDIX 1: Disc diffusion supplemented tables: performance standards for antimicrobial susceptibility testing for entero bacteriaceae

APPENDIX 2

APPENDIX 3: Zones of inhibition for screening of the E. coli isolates for ESBLs based on CLSI breakpoint using Ceftriaxone (CTR), Ceftrazidine (CAZ) and Aztreonam (ATM) antibiotics

LIST OF TABLES

Table 1:        Cultural, Morphological and Biochemical characteristic of the Escherichia coli isolates

Table 2:        Incidence and Percentage Occurrence of the E.coli isolates in the Waste water samples

Table 3:        Antibiotics susceptibility pattern of the isolated E. coli isolates from the Waste water samples

Table 4:        The multiantibiotics resistance (MAR )index of the E. coli isolates

Table 5:        Percentage Occurrence of ESBL producing E. coli isolates based on CLS 1 break point

Table 6:        Percentage occurrence of ESBL producing E. coli isolates based on the double disc synergy test (DDST)

CHAPTER ONE

INTRODUCTION

Waste water is any water that has been affected by human use. It is “used water” from any combination of domestic, industrial, commercial or agricultural activities, surface runoff or storm water and any sewer inflow or sewer infiltration (Tilley et al., 2016). Waste water is a by-product of domestic, industrial, commercial or agricultural activities. Types of waste water include: domestic waste water from households municipal waste water from communities (also called sewage) and industrial waste water from industrial activities. According to WHO (2006), waste water can contain physical (sand, grits, metals, ceramics), chemical (heavy metals, pharmaceuticals, gases and pesticides) and biological pollutants (Bacteria, viruses, parasites and protozoa).

Waste water may be conveyed in a sanitary sewer which conveys only sewage or transported in a combined sewer which includes storm water runoff and industrial waste water. Waste water that is discharged into the environment without suitable treatment causes water pollution, (Tilley et al, 2016). The use of untreated waste water for irrigation purposes is of a great public health importance as waste waters has also been linked to the dispersal of antibiotic resistance. The term antibiotic resistance is a subset of antimicrobial resistance as it applies to only bacteria becoming resistant to antibiotics (WHO, 2014). Resistant microbes are difficult to treat requiring alternative medications or higher doses of antimicrobials. These approaches may be more expensive, more toxic or both (CDC, 2018).

Growing resistance has also been linked to the dumping of inadequately treated effluents from pharmaceutical industry and hospital effluents (sewage) especially in countries where bulk drugs are manufactured.  Also, antimicrobial resistance could be linked to the presence of antimicrobial agents in waste water (Gulberg et al., 2011).

1.1       Aims and Objectives

1.     To isolate and identify Escherichia Coli from municipal waste water

2.     To determine the antimicrobial profile of the E. Coli isolates

3.     To screen the E. coli for extended beta-lactamase producers (ESBLS)

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