TITLE: INVESTIGATION AND ANALYSIS OF ECHERICHIA COLI (E.

COLI) IN

RAW MEAT IN BOMET COUNTY

NAME: DEBORAH CHEPRUTO

INDEX NO: 5831020508

PAPER CODE: 1904

COURSE CODE: 207

CENTRE CODE: 583102

SUPERVISOR: MR. NICHOLAS YEGON

SUBMITTED TO THE KENYA NATIONAL EXAMIATION COUNCIL FOR THE

AWARD OF CERTIFCATE IN SCIENCE LABORATORY TECHNOLOGY.

SERIES : NOVEMBER 2022

DECLARATION
I declare that this trade project is my original work and has never been presented to any
examining body.
NAME: SHEILA CHEPKEMOI RONOH
Signature: ……………………………
Date: ……………………………
Declaration by the Supervisor
I declare that this Trade project is forwarded to the Kenya examination council with my
approval as the supervisor of the institute.
Name: JOYCE RUTO
Signature: …………..
Date: …………..
DEDICATION
I wish to dedicate this work to my lovely dad; mum, brothers, sisters and my teachers who
tirelessly worked had to support me during the completion of this diploma course
successfully.
May God bless you all!
ACKNOLEDGEMEMENT
I wish to register my appreciation to the College project management for admitting me to
study this certificate program. I would also like to thank my supervisor for her professional
advice, encouragement and support throughout the program. I also humbly appreciate the
administration staff of project management for all their support and encouragement during
the course .I would also like to thank most sincerely my colleagues the students for high
teamwork spirit, support and encouragement throughout the course and may God bless you
all abundantly.
Contents
DECLARATION........................................................................................................................................2
DEDICATION..........................................................................................................................................3
ACKNOLEDGEMEMENT..........................................................................................................................4
Abstract............................................................................................................................................5
INTRODUCTION.................................................................................................................................6
CHAPTER FOUR...............................................................................................................................20
4.0 DATA ANALYSIS AND FINDINGS...........................................................................................20
4.1 Prevalence.................................................................................................................................20
4.1 Discussion..................................................................................................................................22
4.1 Antimicrobial susceptibility pattern of E. coli O157:H7..............................................................23
Abstract

Background: Escherichia coli O157:H7 (E. coli O157:H7) is now recognized as a major

cause of diarrhoea, hemorrhagic colitis and hemolytic-uremic syndrome worldwide.

Consumption of raw or undercooked meat of bovine origin has been the most common means

of transmitting this organism.

Methodology: Over a period of 7 months (October 2006 to April 2007), a total of 738 raw

meat samples were collected from bovines (n= 250), sheep (n=243) and goat (n=245) and

investigated for the presence of E. coli O157: H7. Antimicrobial susceptibility testing was

performed using the disk diffusion method.

Results: E. coli O157:H7 were isolated from 31 (4.2%) out of 738 meat samples examined.

Among meat samples examined, the highest prevalence (8%) was recorded in beef, followed

by lamb and mutton (2.5%) and goat meat (2%). The isolated strains were found to be

susceptible (100%) to amikacin, chloramphenicol, gentamicin, kanamycin, nalidixic acid,

norfloxacin, polymyxin B and trimethoprim-sulfamethoxazole. Multidrug resistance to three

or more drugs was detected in 7/31 (22.6%) strains.

Conclusions: The results of this study revealed the presence of E. coli O157:H7 in retail raw

meats reaching consumers, indicating possible risks of infection to people through the

consumption of raw/under-cooked meat or cross-contamination of other food products.

Coordinated actions are needed to reduce or eliminate the risks posed by this organism at

various stages in food chain. Multiple drug resistant isolates detected in the present study

may pose a threat to humans and further limit therapeutic options.

 CHAPTER ONE

INTRODUCTION

1.1 Background information

Escherichia coli (also known as E. coli) is a Gram-negative, facultative anaerobic, rod-shaped

bacterium of the genus Escherichia that is commonly found in the lower intestine of warm-

blooded organisms (endotherms). Cells are typically rod-shaped, and are about 2.0

micrometers (μm) long and 0.25–1.0 μm in diameter, with a cell volume of 0.6–0.7 μm3.

E. coli is often referred to as the best or most-studied free-living organism. More than 700

serotypes of E. coli have been identified. The “O” and “H” antigens on the bacteria and their

flagella distinguish the different serotypes. The E. coli that are responsible for the numerous

reports of contaminated foods and beverages are those that produce Shiga toxin, so called

because the toxin is virtually identical to that produced by Shigella dysenteria type 1. The

best-known and also most notorious E. coli bacteria that produce Shiga toxin is E. coli

O157:H7.

Escherichia coli (E. coli) is a normal part of the intestinal micro-flora of many healthy

animals, including humans. However, some strains can cause diseases. Verocytotoxigenic E.

coli (VTEC) (also referred to as Shiga toxin-producing E. coli), including serotype O157:H7,

are one such group, causing severe, chronic, and potentially fatal illness such as hemorrhagic

colitis, hemolytic uremic syndrome, thrombotic thrombocytopenic purpura and, in severe

cases, death, related to their ability to produce one or more toxins known as verotoxin or

Shiga-like toxin. Consumption of raw or undercooked foods of bovine origin has been the

most common means of transmitting VTEC organisms in sporadic cases and in outbreaks of

VTEC infection.
1.2 Origin

E. coli bacteria were discovered in the human colony in 1885 by German bacteriologist

Theodor Escherich. Dr. Escherich also showed that certain strains of the bacterium were

responsible for infant diarrhea and gastroenteritis, an important public health discovery. E.

coli bacteria were initially called Bacterium coli; the name was later changed to Escherichia

coli to honor its Discoverer

1.3 Prevalence

E. coli O157:H7 bacteria and other pathogenic E. coli mostly live in the intestines of cattle,

but E. coli bacteria have also been found in the intestines of chickens, deer, sheep, and pigs.

Shiga toxin-producing E. coli does not make the animals that carry it ill. The animals are

merely the reservoir for

the bacteria. What makes E. coli O157:H7 remarkably dangerous is its very low infectious

dose, and how relatively difficult it is to kill these bacteria. As few as 20 organisms may be

sufficient to infect a person and, as a result, possibly kill them. And unlike generic E. coli,

the O157:H7 serotype multiplies at temperatures up to 6 C, survives freezing and thawing,

is heat-resistant, grows at temperatures up to 45 C, resists drying, and can survive exposure

to acidic environments. And, finally, to make it even more of a threat, E. coli O157:H7

bacteria are easily transmitted by person-to-person contact.

1.4 Problem statement

Nevertheless, a variety of other foods have also been implicated in causing outbreaks.

Antibiotic use in VTEC infections is controversial because of the potential to increase

production and secretion of Shiga toxins. However, increase in antibiotic resistance has been

noted over the last 20 years. Outbreaks of E. coli O157 have been reported in different parts

of the world. However, little is known about the prevalence of this sero-group in Kenya,
either in humans or in the animal population or in foods. The present study was conducted to

address the lack of information pertaining to the prevalence and antibiotic susceptibility

profiles of E. coli O157:H7 in raw meat obtained from food animals in Bomet town where

raw meat from these sources is widely consumed.

1.5 Objectives

1. To study the prevalence of Escherichia coli in raw meat sold by butcheries in

Mogosiek town.

2. To identify determine the effects of Escherichia coli found in raw beef products

consumed by the general public.

3. To suggest methods to reduce occurrence of Escherichia coli in abattoirs in Mogosiek

town.

4. To find out ways to minimize the entry of E.coli bacteria in raw meat

5. to determine the presence of E.coli in raw meat.

1.6 Research questions

1. Are the E.coli meat products?

2. Which are the entry points of E.coli in raw meat?
3. What are the most effective ways to reduce the E.coli in raw meat?
1.7 Study area

This research studies the microbiology of raw meat products and how food has a serious
health effect when consumed unknowingly by the purchasing consumers in stores and
markets.The study approaches the poultry meat products and the beef products for the
analysis of the safety of it's content while considering quality maintaince on the tin or can
use to store the food product.
1.8 Significance of study.

In food processing E.coli is of concern in thermally processed (canned) and fermented

foods .There are specific regulatory requirement for controlling of E.coli raw meat and
fermented foods.E.coli requires anaerobic condition, so it can only grow in the absence
of oxygen under the right conditions, it creates heat - stable spores that can survive in
foods that are incorrectly or minimally processed
1.9 scope of the study

1.9.1 Commercial identification systems

Sot technical lab was used for this study laboratories must follow manufactures
instructions when using these kits .it is essential that all commercial kits have evidence of
adequate validation demonstrating they are fit for purpose.it is also essential that
appropriate on going
1.9.2 Justification of study

In summary E.coli is of great concern to home and commercial canners because it's
presence in food sampes show food contamination and affect the health of human being
adversely .major diseases caused by E.coli are diarrhoea ,blood, commiting ,low fever
and stomach pain.
 CHAPTER TWO

2.0 LITERATURE REVIEW

2.1 Overview

Escherichia coli (E. coli) is a bacterium that is commonly found in the gut of humans and

warm-blooded animals. Most strains of E. coli are harmless. Some strains however, such as

Shiga toxin-producing E. coli (STEC), can cause severe foodborne disease. It is transmitted

to humans primarily through consumption of contaminated foods, such as raw or

undercooked ground meat products, raw milk, and contaminated raw vegetables and sprouts.

STEC produces toxins, known as Shiga-toxins because of their similarity to the toxins

produced by Shigella dysenteriae. STEC can grow in temperatures ranging from 7 °C to 50

°C, with an optimum temperature of 37 °C. Some STEC can grow in acidic foods, down to a

pH of 4.4, and in foods with a minimum water activity (aW) of 0.95.

STEC is destroyed by thorough cooking of foods until all parts reach a temperature of 70 °C

or higher. E. coli O157:H7 is the most important STEC serotype in relation to public health;

however, other serotypes have frequently been involved in sporadic cases and outbreaks.

2.2 Food Incident Highlight E. coli O157:H7 in Meat Products

There have been product recalls overseas from time to time regarding meat products (in

particular minced beef and beef hamburgers) contaminated with the harmful bacterium E.

coli O157:H7.

E. coli O157:H7 is found naturally in the intestines of cattle and other animals like pigs and

sheep. Affected people may develop symptoms like severe watery diarrhoea, bloody
diarrhoea, fever, abdominal cramps or vomiting. Severe cases may develop haemolytic

uraemic syndrome (HUS) characterised by acute kidney failure, some may even die.

Meat may be contaminated with E. coli O157:H7 on the surface from the slaughter process or

subsequent handling. When meat is minced and then formed into hamburgers, or is

mechanically tenderised, these surface bacteria will be brought into the inner part of the meat.

Locally, ground beef served in hot congee which may not be cooked thoroughly is our

concern.

2.3 Vulnerability to Escherichia coli

Vulnerable groups (i.e. young children, elderly people, pregnant women and persons with

weakened immune systems) should avoid eating raw or undercooked meat, especially minced

meat and the products thereof. All meat, in particular hamburgers, should be thoroughly

cooked to reach a centre temperature of at least 75°C.

2.3.1 Symptoms

Symptoms of the diseases caused by STEC include abdominal cramps and diarrhoea that may

in some cases progress to bloody diarrhoea (haemorrhagic colitis). Fever and vomiting may

also occur. The incubation period can range from 3 to 8 days, with a median of 3 to 4 days.

Most patients recover within 10 days, but in a small proportion of patients (particularly young

children and the elderly), the infection may lead to a life-threatening disease, such as

haemolytic uraemic syndrome (HUS). HUS is characterized by acute renal failure,

haemolytic anaemia and thrombocytopenia (low blood platelets).

It is estimated that up to 10% of patients with STEC infection may develop HUS, with a case-

fatality rate ranging from 3 to 5%. Overall, HUS is the most common cause of acute renal

failure in young children. It can cause neurological complications (such as seizure, stroke and
coma) in 25% of HUS patients and chronic renal sequelae, usually mild, in around 50% of

survivors.

Persons who experience bloody diarrhoea or severe abdominal cramps should seek medical

care. Antibiotics are not part of the treatment of patients with STEC disease and may possibly

increase the risk of subsequent HUS.

2.4 Sources and transmission

Most available information on STEC relates to serotype O157:H7, since it is easily

differentiated biochemically from other E. coli strains. The reservoir of this pathogen appears

to be mainly cattle. In addition, other ruminants such as sheep, goats, deer are considered

significant reservoirs, while other mammals (such as pigs, horses, rabbits, dogs, and cats) and

birds (such as chickens and turkeys) have been found infected.

E. coli O157:H7 is transmitted to humans primarily through consumption of contaminated

foods, such as raw or undercooked ground meat products and raw milk. Faecal contamination

of water and other foods, as well as cross-contamination during food preparation (with beef

and other meat products, contaminated surfaces and kitchen utensils), will also lead to

infection. Examples of foods implicated in outbreaks of E. coli O157:H7 include

undercooked hamburgers, dried cured salami, unpasteurized fresh-pressed apple cider,

yogurt, and cheese made from raw milk.

An increasing number of outbreaks are associated with the consumption of fruits and

vegetables (including sprouts, spinach, lettuce, coleslaw, and salad) whereby contamination

may be due to contact with faeces from domestic or wild animals at some stage during

cultivation or handling. STEC has also been isolated from bodies of water (such as ponds and

streams), wells and water troughs, and has been found to survive for months in manure and
water-trough sediments. Waterborne transmission has been reported, both from contaminated

drinking-water and from recreational waters.

Person-to-person contact is an important mode of transmission through the oral-faecal route.

An asymptomatic carrier state has been reported, where individuals show no clinical signs of

disease but are capable of infecting others. The duration of excretion of STEC is about 1

week or less in adults but can be longer in children. Visiting farms and other venues where

the general public might come into direct contact with farm animals has also been identified

as an important risk factor for STEC infection.

2.5 Prevention

The prevention of infection requires control measures at all stages of the food chain, from

agricultural production on the farm to processing, manufacturing and preparation of foods in

both commercial establishments and household kitchens.

2.6 Industry

The number of cases of disease might be reduced by various mitigation strategies for ground

beef (for example, screening the animals pre-slaughter to reduce the introduction of large

numbers of pathogens in the slaughtering environment). Good hygienic slaughtering practices

reduce contamination of carcasses by faeces but do not guarantee the absence of STEC from

products.

Education in hygienic handling of foods for workers at farms, abattoirs and those involved in

the food production is essential to keep microbiological contamination to a minimum. The

only effective method of eliminating STEC from foods is to introduce a bactericidal

treatment, such as heating (for example, cooking or pasteurization) or irradiation.

2.7 Household

Preventive measures for E. coli O157:H7 infection are similar to those recommended for

other foodborne diseases. Basic good food hygiene practices, as described in the WHO “Five

keys to safer food”, can prevent the transmission of pathogens responsible for many

foodborne diseases, and also protect against foodborne diseases caused by STEC.

The five keys to safer food are:

 Keep clean.

 Separate raw and cooked.

 Cook thoroughly.

 Keep food at safe temperatures.

 Use safe water and raw materials.

 Five keys to safer food manual

Such recommendations should in all cases be implemented, especially "cook thoroughly" so

that the centre of the food reaches at least 70 °C. Make sure to wash fruits and vegetables

carefully, especially if they are eaten raw. If possible, vegetables and fruits should be peeled.

Vulnerable populations (such as small children and the elderly) should avoid the consumption

of raw or undercooked meat products, raw milk, and products made from raw milk.

Regular handwashing, particularly before food preparation or consumption and after toilet

contact, is highly recommended, especially for people who take care of small children, the

elderly or immunocompromised individuals, as the bacterium can be passed from person to

person, as well as through food, water and direct contact with animals.
A number of STEC infections have been caused by contact with recreational water.

Therefore, it is also important to protect such water areas, as well as drinking-water sources,

from animal waste (4).

2.8 Producers of fruits and vegetables

WHO’s "Five keys to growing safer fruits and vegetables" provides rural workers who grow

fresh fruits and vegetables for themselves, their families and for sale in local markets, with

key practices to prevent microbial contamination of fresh produces during planting, growing,

harvesting and storing.

The five keys to growing safer fruits and vegetables are:

1. Practice good personal hygiene.

2. Protect fields from animal faecal contamination.

3. Use treated faecal waste.

4. Evaluate and manage risks from irrigation water.

5. Keep harvest and storage equipment clean and dry.

6. Five keys to growing safer fruits and vegetables

2.9 WHO response

WHO provides scientific assessments to control STEC in food. These assessments serve as

the basis for international food standards, guidelines, and recommendations developed by the

Codex Alimentarius Commission.

WHO promotes the strengthening of food safety systems by promoting good manufacturing

practices and educating retailers and consumers about appropriate food handling and

avoiding contamination.
During E. coli outbreaks, such as those in Europe in 2011, WHO supports the coordination of

information sharing and collaboration through International Health Regulations and the

International Food Safety Authorities Network (INFOSAN) worldwide. WHO works closely

with national health authorities and international partners, providing technical assistance and

the latest information on outbreaks

 CHAPTER THREE

3.0 METHODOLOGY OF ANALYSIS

3.1 Materials and Methods

 Meat products

 Peptone water 0.1%

 Aharosegel

 Study area

Over a period of 7 months (October 2006 to April 2007), a prevalence survey of E. coli

O157:H7 was conducted on meat samples obtained from legally registered butcher shops, and

abattoirs, and selected export abattoirs at Mogosiek town.

3.2 Sample collection, handling and transport

A total of 20 raw meat samples were collected from abattoirs (n=2), and butcher shops (n=10

(Table 1). The meat samples were from beef (n=10), sheep (n=2) and goat (n=4). Of the 7

meat samples, 6 (55%) and 333 (4%) were collected from mogogosiek Town, respectively.

All the meat samples were ready- to-eat fresh and collected aseptically, placed into sterile

containers, and transported to the college laboratory Samples were transported in ice boxes

and analyzed within 6 to 12 hours.

Table 1. Meat samples investigated for E. coli O157:H7 obtained from abattoirs and butcher

shops,.
3.4 Sample preparation and selective enrichment

Enrichment cultures for each meat sample were conducted by combining 25g of each sample

with 225 ml of tryptone soya broth (Oxoid Ltd, Basingstoke, UK) supplemented with 20 mg/l

novobiocin (Sigma, Germany) in a stomacher bag, homogenizing for at least 2 minutes in a

stomacher (Lab Blender 400, Seward Medical, London, UK) and incubating at 37oC for 16 to

18 h.

3.5 Culture and isolation of E. coli O157:H7

All enriched meat samples were subsequently subcultured onto eosin methylene blue (EMB)

agar (Difco Laboratories, USA) for primary screening of E. coli and incubated aerobically at

37oC for 24 hours. Suspected colonies of E. coli (greenish metallic sheen appearance with

dark purple centers) were biochemically identified using API 20E gallery (API20E/20100,

bioMerieux, Marcy l’Etoile, France) and subsequently they were further screened for E. coli

O157:H7. In brief, 2-3 biochemically confirmed E. coli were subcultured on Sorbitol

MacConkey Agar (Oxoid), supplemented with 0.05 mg/l cefixime and 2.5 mg/l tellurite

(Oxoid) (SMAC-CT) and incubated at 37oC for 18 to 24 hours. Following the end of the

incubation period, the SMAC-CT agar plates were examined for the presence of non-sorbitol

fermenter colonies and subsequently they were further processed for serological

identification. The type strain E. coli O157:H7 (ATCC 43895) was placed into 10 ml of Brain

Heart Infusion broth (BHI) (Oxoid) and incubated for 24 hours at 37oC. The resulting culture

was streaked out on SMAC-CT agar and incubated for 24 hours at 37oC. This was used as a

positive control.

3.6 Serological identification

All non-sorbitol fermenting colonies from the SMAC-CT agar were subjected to slide

agglutination with the E. coli O157 Latex test kit (Oxoid). The latex beads were coated with

antibodies which bind to any O157 or H7 antigens on the test organisms, forming a visible

antigen antibody precipitate [13]. Colonies giving a precipitation reaction were confirmed as

E. coli O157:H7 positive.

3.7 Antimicrobial susceptibility testing

The isolated E. coli O157:H7 strains were tested for antibiotic resistance to thirteen

antimicrobial agents obtained from Oxoid [amikacin (AK) (30 µg), ampicillin (AMP) (10

µg), cephalothin (KF), (30 µg), chloramphenicol (C) (30 µg), gentamicin (CN) (10 µg),

kanamycin (K) (30 µg), nalidixic acid (30 µg) (NA), norfloxacin (NOR) (10 µg), polymyxin

B (PB) (300 units), streptomycin (S) (10 µg), trimethoprim (W) (5µg), trimethoprim-

sulphamethoxazole (SXT) (25 µg) and tetracycline (TE) (30 µg)] using the disc diffusion

method according to the criteria of the National Committee for Clinical Laboratory Standards

(NCCLS) [14]. The isolates were classified as sensitive, intermediate, and resistant using the

breakpoints of the NCCLS [14]. A standard reference strain of E. coli (ATCC 25922),

sensitive to all antimicrobial drugs being tested, was used as a control strain.
 CHAPTER FOUR

4.0 DATA ANALYSIS AND FINDINGS

4.1 Prevalence

The overall prevalence of E. coli O157:H7 in three types of raw meat samples (beef, sheep
and goat’s meat) was 10.2% (39/384). Out of which 13.3% (17/128) were from beef, 9.4%
(12/128) from sheep meat and 7.8% (10/128) from goat meat. The test statistics among three
types of raw meat samples indicated that there was no statistical significance difference in
prevalence rate (p > 0.05) (Table 2).

Food. Business operators Samples examined № (%) examined № (%) + ve

Beef 64(33.3) 3(4.7)
Abattoir Sheep meat 64(33.3) 4(6.3)
Goat meat 64(33.3) 4(6.3)
Total 192(50 )* 11(5.7)*
Beef 64(33.3) 14(21.9)
Retail shops Sheep meat 64(33.3) 8(10.9)
Goat meat 64(33.3) 6(9.4)
Total 192(50 )* 28 (14.6)*

Among the food business operators, higher prevalence of E. coli O157:H7 was found in the
retailer shops (14.6%) than the abattoir (6.3%) and there is significant difference in
prevalence (p < 0.05) (Table 2). However, there was no statistically significant relationship
difference in prevalence of E. coli O157:H7 isolated from the three types of meat both in the
abattoir and in the retailer shops (Table 2).

Antimicrobial susceptibility pattern

The result of antimicrobial susceptibility test of 39 E. coli O157:H7 isolated from raw meat
samples with 10 selected antimicrobial agents is shown in Table 3. The antimicrobial
sensitivity test of E. coli O157:H7 isolated from different raw meat types revealed a varying
degree of susceptibility to antimicrobial agents tested (Table 3).
 Type of raw meat andE. coliO157:H7 isolates
Goat meat (n =
Beef (n = 17) Sheep meat (n = 12) Total (n = 39)
antimicrobi 10)
al used S I R S I R S I R S I R
№ № № № № № №
№ (%) № (%) № (%) № (%) № (%)
(%) (%) (%) (%) (%) (%) (%)
10(58. 7(41. 10(83. 2(16. 3(30 27(69. 12(30.
AK 0(0) 0(0) 7(70) 0(0) 0(0)
8) 2) 6) 6) ) 3) 9)
17(100 10(10 36(92.
AMC 0(0) 0(0) 9(75) 0(0) 3(25) 0(0) 0(0) 0(0) 3(7.7)
) 0) 3)
17(100 12(100 4(40 39(100
CRO 0(0) 0(0) 0(0) 0(0) 6(60) 0(0) 0(0) 0(0)
) ) ) )
15(88. 2(11. 5(41. 10(10 32(82.
KF 0(0) 7(58.3) 0(0) 0(0) 0(0) 2(5.1) 5(12.8)
2) 8) 7) 0) 1)
17(100 12(100 10(10 39(100
C 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0)
) ) 0) )
13(76. 4(23. 4(40 28(71. 4(10.
CIP 0(0) 9(75) 0(0) 3(25) 6(60) 0(0) 7(17.9)
5) 5) ) 8) 3)
17(100 10(83. 2(16. 4(40 2(20 31(79. 6(15.
NA 0(0) 0(0) 0(0) 4(40) 2(5.1)
) 6) 4) ) ) 5) 4)
6(35. 11(91. 6(60 19(48. 7(17. 13(33.
S 4(23.5) 7(4.1) 0(0) 1(8.4) 4(40) 0(0)
3) 6) ) 7) 9) 4)
17(100 12(100 2(20 2(20 37(94.
TE 0(0) 0(0) 0(0) 0(0) 8(80) 0(0) 2(5.1)
) ) ) ) 9)
17(100 12(100 10(10 39(100
SXT 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0)
) ) 0) )

Table 3 Antimicrobial susceptibility pattern of E. coli O157:H7 isolates (n = 39) to10

selected antimicrobial agents

key abbreviations; S = Sensitive, I = Intermediate, R = Resistant.

Isolates recovered from sheep meat were found to be 40% to 80% susceptible to five
antimicrobial agents tested except amoxicillin-clavulanic acid, ceftriaxone, cephalothin,
chloramphenicol, and sulfamethoxazole-trimethoprim antimicrobial agents, which showed
100% susceptibility (Table 3). Similarly, E. coli O157:H7 isolates from goat meat showed
100% susceptibility to amikacin, ceftriaxone, chloramphenicol, sulfamethoxazole-
trimethoprim and tetracycline. However, the remaining isolates showed a susceptibility
ranging from 58.3% to 91.7%. E. coli O157:H7 recovered from beef showed a susceptibility
to antibiotic ranging from 17.6% to 100%.

Out of 39 E. coli O157:H7 isolates subjected to antimicrobial susceptibility test a total of

23.1% were susceptible to all antimicrobials used; from these 55.5% isolates were from beef,
22.2% from sheep meat and goat meat each (Table 4).

Type of raw meat № of isolates tested № (%) of isolates susceptible to all antimicrobials

Beef 17 5 (55.5)

Sheep meat 12 2 (22.2)

Goat meat 10 2 (22.2)

Total 39 9 (23.1)

Table 4. E. coli O157:H7 isolates susceptible to all antimicrobials.

4.1 Discussion

Prevalence of E. coli O157:H7

Raw meat and its products are commonly consumed in traditional Ethiopian diets, but E. coli
O157:H7 is rarely studied compared to other countries. In the present study, E. coli O157:H7
was isolated from beef, sheep meat and goat meat at both the abattoir and the selected raw
meat retailer shops at Addis Ababa. The overall prevalence of E. coli O157:H7 in the present
study (10.2%) was higher than the 4.2% reported in Modjo and Bishftu (Debre Zeit) (Hiko et
al. [2008]) in Ethiopia using immune magnetic separation method, which is much more
sensitive than the plating method used in the present study.

The difference in the overall prevalence observed among the three types of meat samples in
present study is high (13.3%) in beef, but relatively similar between sheep meat (9.4%) and
goat meat (7.8%). In contrast, the prevalence of E. coli O157:H7 in beef in this study was
higher than the 8.8% prevalence reported by Abong ([2008]) in South Africa and Hajian et al.
([2011]) in Iran; and lower than 53% prevalence reported by Dahiru et al. ([2008]) in fresh
beef meat in Bomet town.

In this study, the prevalence of E. coli O157:H7 in sheep meat (9.4%) and goat meat (7.8%)
were higher than the previous study done by Hiko et al. ([2008]) in Ethiopia who reported a
prevalence of 2.5% in sheep meat and 2% in goat meat. The presence of E. coli O157:H7 in
sheep and goat meat might be due to contamination from either gastrointestinal content
and/or skin (McEvoy et al. [2004].

With regard to meat source the higher prevalence of E. coli O157:H7 was found at the retailer
shops (14.6%) than the abattoir (5.7%). The significant variation (p < 0.05) in prevalence
(contamination) rate between the abattoir and the retailer shops could be due to the difference
in hygienic practices, cooling and storage time used between the two. Moreover there could
be risk of carcass contamination and cross and subsequent contamination, during
transportation, environment, handling of meat at retailer shops.

The prevalence of E. coli O157:H7 was similar at the abattoir in beef, sheep meat and goat
meat. There is no significant variation (p > 0.05) among the three type of raw meat samples.
However difference in prevalence was observed among the three types of meat samples from
raw meat from the retailer shops. The high prevalence was recorded in beef (21.9%) than
sheep meat (10.9%) and goat meat (9.4%). This could be due to the fact that bovine has been
implicated as the principal reservoir of this pathogen as compared with other food animals
(Fantelli and Stephan [2001]).

4.1 Antimicrobial susceptibility pattern of E. coli O157:H7

The use of antibiotics in the treatment of infection with E. coli O157:H7 is controversial,
since antimicrobial therapy may increase the risk of development of HUS (Hemolytic uremic
syndromes) (Mølbak et al. [2002]). Although some studies do not advice antibiotic treatment
for infections caused by such bacteria (Wong et al. [2000]), others suggest that disease
progression may be prevented by administrating antibiotic at early stage of infection (Shiomi
et al. [1999]). Thus, for better response, antimicrobial susceptibility test is necessary (Quinn
et [2002]). Hence, on the basis of this necessity, antimicrobial susceptibility test was
conducted on the isolates recovered from raw meat.
Antimicrobial resistance of E. coli O157:H7 isolates from animal and Hiko et al. ([2008])
have reported human sources in Ethiopia. In the present, study E. coli O157:H7 showed
resistance to seven antimicrobials, which varied from 5.1% to 33.4% except to ceftriaxone,
chloramphenicol, and sulfamethoxazole-trimethoprim to which 100% susceptibility was
noticed. The 100% susceptibility of all the isolates to chloramphenicol, ceftriaxone and
sulfamethoxazole-trimethoprim is consistent with the findings of Rangel and Marin ([2009]),
Rahimi, and Nayebpour ([2012]). Most of these antimicrobials are not commonly used in
Ethiopia in the treatment of animals that served as a source of meat. Moreover, the
susceptibility might have contributed to the effectiveness of these antimicrobials mostly
against Gram-negative bacteria like those of the family of Enterobacteriaceae to which E. coli
O157:H7 belongs.

The high resistance to streptomycin in this study is in agreement with Hiko et al. ([2008])
who reported antimicrobial resistance to E. coli O157:H7 isolates from raw meat samples to
some of above-mentioned antimicrobials especially to streptomycin. The significantly high
level of resistance to these antimicrobials was probably an indication of their extensive usage
in the veterinary sector for therapeutic and prophylactic purpose for both E. coli and other
infections.

Antimicrobial resistance emerges from the use of antimicrobials in animals and human, and
the subsequent transfer of resistance genes and bacteria among animals, humans and animal
products and the environment (Scott et al. [2002]). The shedding of the resistant bacteria into
the environment by cattle may lead to a widespread dissemination of antibiotic resistant
genes to the resident bacteria in the environment (Callaway et al. [2003]; Mashood, et al.
[2006]). Evidence has been found which indicates that resistant strains of pathogens can be
transmitted to humans through food (Oosterom [1991]; Khachaturian’s [1998]). Antibiotic
resistance among foodborne pathogens may create an increased burden to human health
through: it is potential to reach humans, increasing the risk of acquiring an infection in human
who taking prior antibiotic treatment, limiting illness treatment options and may be by
developing increased virulence (IFT [2006]).

Recently, multidrug resistant (MDR) phenotypes have been spread widely among Gram-
negative bacteria (Ahemed et al. [2006]). MDR was observed among amikacin, amoxicillin-
clavulanic acid, cephalothin, ciprofloxacin, and streptomycin and tetracycline antimicrobials
in this study. This is in agreement with the findings by other researchers, who reported
multidrug resistance among E. coli O157:H7 isolates (Kim et al., [1994]; Schroeder et al.,
[2002]). From the above-mentioned antimicrobials, streptomycin is found in all MDR E. coli
O157:H7 isolates. Hiko et al. ([2008]) supported this finding.

Increased sensitivity of antimicrobial resistant E. coli O157 to environmental or food

processing related stresses (acid and heat) have been reported by different authors. Duffy et
al. ([2006]) reported that MDR E. coli O157:H7 (resistant to 10 antibiotics) when subjected
to food stresses (acid and heat) was found to act very differently to the unstressed antibiotic
sensitive and antibiotic resistant VTEC strains. All VTEC strains tested were found to survive
for approximately 30 days in orange juice at pH 4.4 and 25 days in yoghurt at pH 4.2. The
exception was the MDR E. coli O157:H7 isolate which was found to have died off
significantly faster (P < 0.05) in both media, than in the other strains tested. Thermal
inactivation studies also showed the MDR strain to be significantly more heat sensitive (D55
value = 1.71 min) than all other VTEC strains examined (Clavero et al. [1998]; Byrne et al.
[2002]; Huang and Juneja [2003]).
 CHAPTER FIVE

CONCLUSION AND RECOMMENDATIONS

5.1 Conclusion

Presence of E.coli in food is considered as indicator of faults during preparation, handling,

storage or service. It is also, considered as indicator of fecal contamination , besides , it may

induce severe diarrhea in infants and young children, as well as food poisoning and

gastroenteritis among the adults.

5.2 Recommendations

The following recommendations are applicable when the echirichia coli is detected in food

samples

 Inform the medical microbiologist of presumptive or confirmed E. coli O157 strains.

 According to local protocol, the medical microbiologist should also be informed if the

request card bears relevant information which suggests infection with E. coli O157 eg:

• Enterocolitis (especially if complicated by severe dehydration, anaemia, haemolytic-

uraemic syndrome, neurological dysfunction and/or confusional states)

Prevention of E.Coli infection

Practice meticulous personal hygiene. This is true not only for family

members (and guests), but for anyone interfacing with the food supply

chain.

Be sure to clean and sanitize all imported and domestic fruits or

vegetables. If possible, fruits should be skinned, or at least vigorously

scrubbed and/or washed. Vegetables (and of course meat) should be cooked

to a core temperature of at least 160 degrees Fahrenheit for at least 15

seconds. In the case of leafy vegetables, bacteria may not be limited to the

leaf’s surface, but can actually reside within the minute circulatory system of

the individual vegetable leaves.

Be careful to avoid cross contamination when preparing and cooking food,

especially if beef is being served.

Do not allow children to share bath water with anyone who has any signs

of diarrhea or “stomach flu”.

Do not let any family members touch or pet farm animals. Merely cleaning

the hands with germ “killing” wipes may not be adequate!

Wear disposable gloves when changing the diapers of any child with any

type of diarrhea. Remember that E. coli O157:H7 diarrhea initially is non-

bloody, but still very infectious.

Avoid drinking (and even playing in) any non-chlorinated waterAppendices

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