ISOLATION OF ENTERIC BACTERIA AND STAPHYLOCOCCUS AUREUS FROM

STREET-VENDED READY-TO-EAT FOODS SOLD AT THE UNIVERSITY OF

ZIMBABWE CAMPUS AND THEIR SENSITIVITY TO SELECTED ANTIBIOTICS

By

Talent Mangeya (R197835X)

A Thesis submitted in partial fulfilment of the requirements for the degree of Bachelor of
Sciences Honours in Biological Sciences

University of Zimbabwe

Faculty of Science

Department of Biological Sciences

Zimbabwe

May 2024

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 CHAPTER 1: INTRODUCTION

1.1 Background

Street food venting plays a significant role in urban living in developing countries as a result of
widespread unemployment and limited job prospects (Khairuzzaman et al., 2014). Street vendors
are particularly valued by low-income urban dwellers for the distinct taste and nutritional value
of the food they offer (Sabuj et al., 2020). Unfortunately, the safety of street food has been
observed to be unsafe due to inadequate adherence to hygienic practices during preparation,
handling, sales, and use of improper storage temperatures. These food items have been
associated with outbreaks of foodborne illnesses (Khairuzzaman et al., 2014).

Bacterial food contamination has been linked with the upsurge in food poisoning cases (WHO
2015). Bacterial food poisoning refers to a group of illnesses caused by ingesting viable
pathogenic bacteria or preformed toxins following bacterial growth in the food (Khairuzzaman et
al., 2014). In developing countries, most of food borne outbreaks occurs in institutional settings,
such as colleges and also generally in food eating places including hotels and restaurants (Yilmaz
and Çakıroğlu 2016), though cases have been recorded for home-made food. Some of these
foodborne disease outbreaks are due to bacterial contamination of ready-to-eat food sold in
public places (Mengistu and Tolera 2020)

Bacterial food contamination has been linked with the upsurge in food poisoning cases (WHO
2015). Bacterial food poisoning refers to a group of illness caused by ingesting viable pathogenic
bacteria or preformed toxins following bacteria growth in the food (Quinn et al. 1999). In
developing countries, most of food borne outbreaks occurs in institutional settings, such as
colleges (Yilmaz and Çakıroğlu 2016), though cases have been recorded for home-made food.
Most of these foodborne disease outbreaks are due to bacterial contamination of ready-to-eat
food sold in public places (Mengistu and Tolera 2020)

There are several factors associated with bacterial food contamination, including the poor
implementation of handling practices, undercooking, cross-contamination, insufficient personal
hygiene of food handlers, inadequate hygiene of equipment and food-contact surfaces, as well as
inadequate temperature and holding time of the ready- to eat food (Khairuzzaman et al. 2014). In

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all of these instances, bacteria are introduced into the food chain, and depending on the type of
food and subsequent storage temperature, they multiply to significant levels, with certain species
producing toxins before ingestion while some grow and produce toxins after ingestion. Food can
become contaminated by both pathogenic and non-pathogenic bacteria. The occurrence of
bacterial food contamination is primarily attributed to the lack of awareness among street
vendors regarding proper food safety practices during food preparation, processing, handling and
storage (Wandolo 2016, Valero et al. 2016).

Consuming street-vended foods that are contaminated with high levels of coliform such as
Escherichia coli, Klebsiella pneumoniae, Enterobacter aerogenes and Citrobacter freundii and
pathogenic bacteria, such as Salmonella species, Staphylococcus aureus, Escherichia coli,
Bacillus cereus, and Clostridium perfringens, not only results in illness but can also lead to
death, especially among the elderly and those with weakened immune systems (Institute Of Food
Technologists 2004). The risk of contracting microbial diseases depends on factors such as the
specific pathogen and its infective dose, the immune status of the host, the condition of the
stomach and upper small intestine at the time of exposure, and the composition of the food in
which the pathogen is present. Furthermore, Consuming foods containing these bacteria may
increase the risk of multidrug resistance in consumers, posing a significant health risk to the
general population (Anwar 2018, Khalif et al. 2018).

There is need to implement food safety programmes on street-vended foods, that identify the
source of food contamination and offer preventive measures in order to protect the health of
consumers. .To enhance food safety and mitigate the risk of foodborne illnesses caused by
bacterial contamination, it is crucial to first identify the issue of bacterial contamination in street-
vended ready-to-eat foods (Yilmaz and Çakıroğlu 2016). This necessitates a comprehensive
assessment of the presence of bacteria in such food items. By doing so, appropriate measures can
be implemented to minimize and prevent contamination, ensuring the safety of consumers. This
involves isolating and identifying indicator bacteria, such as enteric bacteria (such as Escherichia
coli, Salmonella, Shigella, and Campylobacter) and Staphylococcus aureus, in food using
standard microbiology procedures. The presence of hygiene indicator organisms in food, though
not inherently a hazard, serves as a warning sign that harmful bacterial pathogens may also be
present (Health Protection Agency 2009).
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1.2 Statement of the Problem

The occurrence of foodborne diseases due to bacterial pathogens food contamination has been a
concern worldwide (WHO 2015, Alelign et al. 2023,), and it has been postulated that street-
vended ready-to-eat foods contributes significantly to the problem (Khairuzzaman et al. 2014).

The University of Zimbabwe is one of the busiest tertiary institutions in Harare, and moreover, it
has been experiencing overcrowding at its hostel premises due to unauthorized occupation by
students. The net effect has been a rapid expansion in the number of street vendors on the
campus. These street vendors primarily consist of food vendors. This increase in the street stalls
selling ready-to-eat food has largely occurred without effective regulations on food safety and
hygiene thus posing a potential threat to students’ health.

1.3 Significance of the study

Extensive documentation has revealed a significant correlation between bacterial contamination

of food and the occurrence of diseases, leading to high rates of morbidity and mortality
worldwide (Alelign et al., 2023). In developing countries, foodborne outbreaks, particularly
within institutional settings, are prevalent (Khairuzzaman et al., 2014). At the University of
Zimbabwe, a considerable number of students rely on ready-to-eat food from informal street
stalls located on campus. Information from a personal interview with Dr Harris (16 October
2023) at University of Zimbabwe on-campus clinic reveal that at least 5 people suffer from
diarrhoea each month as a result of bacterial infection. Given the potential impact of
contaminated food on public health, it is therefore crucial to conduct a comprehensive
bacteriological quality assessment specifically focusing on the ready-to-eat foods sold by these
informal street vendors at the University of Zimbabwe.

Despite the prevalent concerns surrounding microbial food contamination at the University of
Zimbabwe, limited research has been conducted to investigate the prevalence and isolation of
bacteria, as well as their antibiotic susceptibility patterns. Therefore, this study aims to gather
essential data regarding the prevalence and isolation of bacterial strains found in ready-to-eat
street-vended foods at the University of Zimbabwe, while also analysing their antibiotic
susceptibility profiles.

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The establishment of food catering services in Zimbabwe should follow stringent food safety
regulations to ensure the health and well-being of consumers, as mandated by existing
government statutory laws (Food and Food Standards Act 2023). However, the continued
increase in the number of street food vendors at the University of Zimbabwe, combined with the
lack of knowledge among most food handlers regarding hygiene practices (Khairuzzaman et al.
2014), suggests a potential non-compliance with these food safety regulations or a need for
stronger enforcement measures. This situation poses a significant risk to consumer health.

Therefore, conducting a study to evaluate the bacteriological quality of ready-to-eat food sold on
the campus becomes crucial. This study aims to provide evidence to the responsible authorities
regarding the current state of food safety practices among street vendors and the associated risks.
By identifying any shortcomings in compliance with food safety regulations, the study will
underscore the urgency of crafting a system of providing formal spaces for vendors on campus,
rather than allowing them to simply park on the road. Ultimately, the study aims to contribute to
enhancing food safety standards among street vendors at the University of Zimbabwe, creating a
safer and healthier food environment for consumers.

1.4 Research Questions

1. What are the pathogenic bacteria contaminating street-vended ready-to-eat food served
from street stalls at the University of Zimbabwe campus?

2. What is the total number of coliforms and [Link] in street-vended ready-to-eat food at the
University of Zimbabwe campus?

3. What is the prevalence of bacteria isolates from street-vended ready-to-eat food at the
University of Zimbabwe campus?

4. Are the bacteria isolates from street-vended ready-to-eat at the University of Zimbabwe
campus susceptible to commonly prescribed antibiotics?

5. Is there any significant difference in total coliforms and [Link] counts present in street-
vended ready-to-eat food on different days at the University of Zimbabwe campus?

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1.5 Main objective of the study

The main objective of this study was to determine the bacterial quality of street-vended ready-to-
eat food (cooked chicken, beef and sausage, and vegetable salad) sold at the University of
Zimbabwe campus.

1.5.1 Specific objectives

1. To identify enteric bacteria and staphylococcus aureus in selected street-vended food

items (cooked chicken, beef, sausage, and vegetable salad) from three street vendors at
the University of Zimbabwe campus on three different days.
2. To estimate total coliforms and [Link] counts in selected street-vended food items
(cooked chicken, beef, sausage, and vegetable salad) from three street vendors at the
University of Zimbabwe campus on three different days.
3. To determine the prevalence of bacteria isolates from street-vended ready-to-eat food at
the University of Zimbabwe campus?
4. To determine the antibiotic susceptibility patterns of bacterial isolates from selected
street-vended food items (cooked chicken, beef, sausage and vegetable salad) from three
street vendors at the University of Zimbabwe campus on three different days.
5. To determine if there is any significant difference in total coliforms and [Link] counts
present in street-vended ready-to-eat food (cooked chicken, beef and sausage, and
vegetable salad) on different days at the University of Zimbabwe campus.

1.6 Hypothesis

It was hypothesized that:

H0₁: There are no pathogenic bacteria contaminating street-vended ready-to-eat food served
from street stalls at the University of Zimbabwe campus.

H0₂: The bacteria isolates from street-vended ready-to-eat food at the University of Zimbabwe
campus are not susceptible to commonly prescribed antibiotics.

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H0₃: There is no significant difference in total coliforms and [Link] counts present in street-
vended ready-to-eat food on different days at the University of Zimbabwe campus.

CHAPTER 2: LITERATURE REVIEW

2.1. Concept of food contamination

Food contamination refers to the presence of harmful substances or microorganisms in food that
can cause illness or injury when consumed (Balter 2006). It can occur through various sources,
including biological pathogens, chemical contaminants, and physical hazards. Biological food
contamination is of greater concern due to the potential for transmitting harmful pathogens that
can cause widespread illnesses (Kadariya et al. 2014). While chemical and physical contaminants
also pose risks, the presence of bacteria, viruses, parasites, and fungi in food can lead to severe
outbreaks of foodborne diseases (Mwove et al. 2023). Therefore, it is crucial to prioritize
measures to prevent, detect, and control biological contamination in the food supply chain to
ensure public health and safety (Balter 2006).

2.1.1 Biological food contamination

Biological food contamination refers to the presence of harmful microorganisms in food, such as
bacteria, viruses, parasites, and fungi (Balter 2006). These contaminants can enter the food
supply chain during different stages, including production, processing, transportation, or storage
(Kadariya et al. 2014). Pathogenic bacteria like Salmonella, Escherichia coli, Campylobacter,
Bacillus cereus, Shigella, and Clostridium perfringens, Listeria monocytogenes, and
Staphylococcus aureus are commonly associated with foodborne illnesses that affect the
gastrointestinal system (Balter 2006). Viruses like Norovirus and Hepatitis A can contaminate
food through infected individuals or contaminated water sources, leading to symptoms such as
nausea, vomiting, and diarrhoea. Parasites such as Toxoplasma gondii and Cryptosporidium can
be found in undercooked meat or contaminated water, causing various illnesses. Fungal
contamination, though less frequent, can occur when certain fungi produce mycotoxins that
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contaminate crops, posing health risks. Proper food handling, hygiene practices, and adherence
to food safety regulations are essential to prevent biological food contamination and protect
public health (Balter 2006, Kadariya et al. 2014).

2.1.2 Chemical food contamination

Pesticides, heavy metals, food additives, and packaging materials can all cause chemical
contamination in food. Pesticides used in agriculture may leave residues on food, potentially
causing health problems. Heavy metals, such as lead, cadmium, mercury, and arsenic, can
contaminate food and endanger human health through environmental contamination. Chemical
contamination can also be caused by food additives and packaging materials such as
preservatives, artificial colours, and chemicals such as BPA and phthalates. Proper regulation,
monitoring, and adherence to safety standards are necessary to minimize chemical contamination
risks in the food supply chain (Mwove at al. 2023).

2.1.3 Physical food contamination

Physical food contamination occurs when food contains foreign substances or objects that can be
harmful to consumers if ingested. These substances can include materials like glass, metal,
plastic, stones, or wood fragments that unintentionally enter the food at various stages of
production, processing, packaging, or distribution. The potential risks associated with physical
contamination mainly involve injuries such as choking, cuts, or harm to the digestive system.
Preventing physical contamination requires strict adherence to food safety practices, quality
control measures, and the use of detection technologies (Mwove at al. 2023).

2.2 Bacterial food contamination

Bacterial contamination in food is a major concern compared to other types of biological

contaminants because bacteria have the ability to produce toxins that directly impact human
health, leading to foodborne illnesses. Unlike viruses and parasites, bacteria can produce toxins
that affect the gastrointestinal tract, nervous system, and cells, causing a wide range of symptoms
and complications. This poses significant public health risks when pathogenic bacteria and their
toxins are present in food. Examples of pathogenic bacteria that can cause severe illnesses

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include Salmonella, Campylobacter, E. coli, and Listeria monocytogenes (Balter 2006, Kadariya
et al. 2014).

Bacterial food contamination occurs when harmful bacteria or their toxins are present in food,
posing a risk of foodborne illnesses upon consumption. This contamination can happen during
various stages of food production, processing, handling, or storage. Factors contributing to
bacterial contamination include poor hygiene practices, inadequate cooking or processing
temperatures, cross-contamination, and the use of contaminated water or ingredients (Sadiku et
al. 2020).

Bacteria have the ability to produce different types of toxins that can cause illness in humans.
These toxins can be classified into three main categories: enterotoxins, which affect the
gastrointestinal tract and cause symptoms like diarrhea, vomiting, and abdominal cramps;
neurotoxins, which impact the nervous system and can lead to paralysis, muscle weakness, and
respiratory failure; and cytotoxins, which damage or destroy cells, resulting in tissue damage and
inflammation (Balter 2006, Sadiku et al. 2020).

Certain pathogenic bacteria are particularly significant as food contaminants due to their
association with foodborne illnesses and the resulting public health concerns. These include
Salmonella, Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus,
Shigella, and Clostridium perfringens, Listeria monocytogenes (Schatzmatr and Streit 2013).

2.3 Factors contributing to Bacterial food contamination

2.3.1 Poor personal hygiene of food handlers

Poor personal hygiene is a major contributor to bacterial food contamination, especially in the
setting of street-vendor foods. Several studies have found a correlation between poor personal
hygiene and food contamination (Abdu 2019).

In uncontrolled street food venues, vendors struggle to maintain acceptable hygiene practices due
to a lack of access to proper sanitation facilities. Ineffective hand washing has been identified as
a key risk factor for food contamination and foodborne infections, particularly due to a lack of
clean water and hand washing facilities. This contributes to the spread of dangerous bacteria
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from vendors' hands to the food they handle. Furthermore, vendors frequently reuse utensils
without properly cleaning and sanitizing them, raising the danger of cross-contamination
between raw and cooked items (Malavi and Muzhingi 2021).

Wearing unclean or infected clothing while handling food may also introduce bacteria into the
food preparation process. Food workers who wear unclean aprons or reuse soiled aprons increase
the danger of cross-contamination. Furthermore, the absence of protective equipment, such as
gloves, directly transfers bacteria from the hands to the meal (Abdu 2019, Lim et al. 2019). Poor
respiratory hygiene is another issue, as coughing or sneezing directly over meals spread airborne
microorganisms. Food workers who have respiratory infections or who do not adequately cover
their lips and noses risk contaminating the food with bacteria that can cause illness. Finally,
certain personal habits, like touching the face, scratching the head, or handling money without
proper hand washing afterward, can also introduce bacteria into the food (Clayton et al. 2003).

2.3.2 Inadequate cleaning and sanitation practices

Inadequate cleaning and sanitation practices contribute significantly to bacterial food

contamination. Inadequate and ineffective cleaning methods can leave food residues in the
kitchen, allowing bacteria to adhere and thrive. In residence kitchens, improper cleaning and
disinfection of chicken carcasses can contribute to cross-contamination and the spread of
pathogens such as Campylobacter and Salmonella. Similarly, inadequate cleaning of food
contact surfaces may allow bacteria such as Staphylococcus aureus to survive and persist (Todd
et al. 2010).

Inadequate cleaning methods extend beyond food preparation areas. Bacterial contamination can
arise on a variety of surfaces, including stethoscopes and mobile phones, as a result of irregular
cleaning techniques. Improper cleaning of retail equipment, such as deli slicers, can potentially
contribute to the risk of foodborne illness (Asfaw and Genetu 2021, Azanaw et al. 2021).

It is critical to promote proper hygiene and food safety education among food handlers and
customers in order to solve inadequate cleaning and sanitation practices. Food safety training
programs have been found to be helpful in improving attitudes among individuals and improving
food safety practices. Clear cleaning and disinfection guidelines and protocols, as well as regular

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inspections and monitoring, can help guarantee that proper cleaning techniques are followed
(Clayton et al. 2003).

2.3.3 Improper storage temperature

Improper food storage temperature is a crucial factor that contributes significantly to bacterial
food contamination, particularly in the setting of street-vendor foods. Because street vendors
often have no access to suitable refrigerated facilities, they are especially vulnerable to the risks
connected with temperature abuse (Abdu 2019, Lim et al. 2019).

When street-vendor foods are not preserved at proper temperatures, harmful microorganisms can
multiply rapidly resulting in foodborne diseases. Bacteria such as Salmonella, Escherichia coli,
and Staphylococcus aureus grow in inadequately controlled temperatures. These bacteria have
the potential to cause serious gastrointestinal infections and are a major public health concern
(Asfaw and Genetu 2021, Azanaw et al. 2021).

Street vendors frequently rely on ambient temperatures to preserve their food supplies,
particularly perishable items such as meats, dairy products, as well as cooked foods. However,
ambient temperatures can vary greatly, particularly in warmer climates or during the summer
months. Bacteria can multiply at a rapid rate when these foods are exposed to temperatures over
the acceptable range, raising the risk of infection (Clayton et al. 2003).

Furthermore, poor storage procedures, such as leaving food out in the open or incorrectly
packaging it, may further exacerbate the problem. Street vendors frequently suffer resource and
time constraints, which might lead to shortcuts in food handling processes. For instance, leaving
food out in the open exposes it to airborne contaminants, insects, and other bacterial
contamination sources (Abdu 2019, Lim et al. 2019).

Food sold on the street is frequently cooked in small amounts to fulfil instant demand, which can
result in leftovers. If these leftovers are not rapidly cooled and stored at the proper temperature,
they become bacteria breeding grounds. Inadequate refrigeration allows bacteria to proliferate
before food is reheated and served again, increasing the risk of foodborne illness (Asfaw and
Genetu 2021, Azanaw et al. 2021).

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2.3.4 Inadequate cooking

Inadequate cooking is a major contributor to bacterial food contamination. Due to limited

resources, time constraints, and varying levels of culinary proficiency, street vendors frequently
face challenges that can lead to insufficient cooking practices and an increased risk of foodborne
illnesses (Asfaw and Genetu 2021, Azanaw et al. 2021, Todd et al. 2010).

When street-vendor foods are not thoroughly cooked, harmful bacteria present in raw or partially
cooked ingredients can survive and multiply, posing a health risk to customers. Bacteria such as
Salmonella, Campylobacter, and Listeria monocytogenes are frequently associated with raw or
undercooked foods and can cause severe gastrointestinal infections (Abdu 2019, Lim et al.
2019).

Due to the lack of precise temperature controls and inadequate cooking equipment, street
vendors may have difficulty achieving and maintaining the necessary cooking temperatures.
Inadequate heating or uneven heat distribution can cause uneven cooking, leaving certain areas
of the food undercooked and vulnerable to bacterial contamination (Asfaw and Genetu 2021,
Azanaw et al. 2021).

Furthermore, the time constraints that street vendors face can result in rushed cooking processes
in which foods are not allowed to reach the required internal temperature for bacterial
elimination. Inadequate cooking times, particularly for meat and poultry products, can result in
partially cooked products that may be contaminated with harmful bacteria (Todd et al. 2010).

2.3.5 Cross contamination

On account of limited workspace, inadequate sanitation facilities, and time constraints, street
vendors regularly encounter challenges that increase the risk of cross contamination and the
spread of harmful bacteria (Abdu 2019, Lim et al. 2019).

Cross contamination occurs when bacteria from one source, such as raw or contaminated food,
are inadvertently transferred to another, such as cooked or ready-to-eat foods. Street vendors
often handle multiple food items at the same time, using the same utensils, cutting boards, and
surfaces without proper cleaning and sanitation procedures. This increases the risk of bacterial
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contamination by transferring bacteria from raw or contaminated ingredients to cooked foods
(Abdu 2019, Lim et al. 2019).

Separating raw and cooked foods can be difficult in street-vendor settings where resources and
space are limited. Without proper segregation, raw or contaminated ingredient juices, drippings,
or residues can come into contact with ready-to-eat foods, resulting in bacterial cross
contamination. For example, if a cutting board is used to prepare raw chicken and then used to
chop vegetables for a salad, any bacteria present on the chicken can be transferred to the salad
ingredients (Abdu 2019, Lim et al. 2019).

Cross contamination can also be caused by poor hand hygiene practices among street vendors.
Hand washing is critical in preventing bacteria transfer from hands to food. However, access to
clean water and appropriate hand washing facilities may be difficult for street vendors. This can
lead to insufficient hand washing, which allows bacteria on hands to contaminate the food being
prepared or served (Asfaw and Genetu 2021, Azanaw et al. 2021).

2.4 Health impacts of bacterial food contamination

2.4.1 Food poisoning

Bacterial food poisoning is a significant threat to public health around the world. It occurs when
individuals consume food that has been contaminated with harmful bacteria or their toxins,
which may trigger symptoms ranging from mild gastrointestinal discomfort to severe illness
(Balter 2006, Kadariya et al. 2014).

Pathogenic bacteria can contaminate food at various stages, including production, processing,
storage, and handling. When contaminated food is consumed, these bacteria can multiply and
produce toxins that cause illness under certain conditions. Due to poor hygiene practices and
uncontrolled storage conditions, street-vended foods, which are sold in open-air markets or by
mobile vendors, are especially prone to bacterial contamination (Abdu 2019, Lim et al. 2019).

There are three types of bacteriological food poisoning: toxicoinfections, intoxications, and
infections. Toxicoinfection occurs when pathogenic bacteria in food are consumed, allowing the
bacteria to grow and multiply in the intestines. This leads to the production of toxins within the
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body, resulting in symptoms such as diarrhoea, abdominal cramps, fever, and sometimes
vomiting. Salmonella and Shigella infections constitute examples of toxicoinfection (Clayton et
al. 2003).

Intoxications, on the other hand, are caused by ingesting pre-formed toxins produced by bacteria
in food prior to consumption. These toxins are heat stable and can withstand cooking
temperatures. Staphylococcus aureus and Bacillus cereus are two common pathogens associated
with street food. Intoxication symptoms typically include nausea, vomiting, abdominal pain, and,
on rare occasions, diarrhoea (Asfaw and Genetu 2021, Azanaw et al. 2021, Todd et al. 2010).

Infections occur when pathogenic bacteria are ingested, and they subsequently colonize and
invade the gastrointestinal tract, leading to illness. These bacteria can cause direct damage to the
digestive system's lining or produce toxins that cause inflammation and other symptoms.
Campylobacter, Escherichia coli, and Listeria monocytogenes are examples of bacterial
infections. Bacterial infections can cause a variety of symptoms, including diarrhoea, abdominal
pain, fever, and, in some cases, vomiting (Asfaw and Genetu 2021, Azanaw et al. 2021).

In Africa, several bacteria are commonly implicated in food poisoning, causing significant health
concerns. These bacteria can contaminate various food sources, including street-vended foods,
leading to widespread illnesses. While the prevalence of specific bacteria may vary across
different regions and countries in Africa, several common pathogens are frequently associated
with foodborne illnesses, including salmonella, staphylococcus aureus, [Link], Vibrio cholera,
Campylobacter, Bacillus cereus, shigella and Listeria monocytogenes (Clayton et al. 2003).

Salmonella food poisoning

Salmonella, a type of bacteria that causes salmonellosis, is a genus that includes several
serotypes, the most common of which is Salmonella enterica. These rod-shaped bacteria are
members of the Enterobacteriaceae family and can survive in environments with or without
oxygen, making them facultative anaerobes. Salmonella has a wide temperature range for
growth, with an optimal temperature of 37-42°C. Certain serotypes, however, can thrive at lower
temperatures, including those found in refrigerated environments. Salmonella can also tolerate a

14
wide pH range, ranging from acidic to alkaline, and can survive in low moisture environments
(Abdu 2019, Lim et al. 2019).

Salmonella infection is a common type of foodborne illness that typically causes a brief
gastrointestinal illness. The infection is usually caused by consuming contaminated food or
water, but other factors can also contribute to its spread. The time it takes for symptoms to
appear can range from 6 to 48 hours, with most cases occurring within 12 to 36 hours. Variations
in incubation time can be influenced by factors such as the amount of bacteria consumed, the
virulence of the specific Salmonella strain, the individual's susceptibility, and the characteristics
of the contaminated food. Even a small dose of around 15 bacterial cells can cause illness.
Diarrhoea, abdominal cramps, vomiting, and fever are common symptoms that last one to seven
days. The bacteria, however, can remain in the faeces for several weeks after the symptoms have
subsided. While most cases resolve on their own, the mortality rate in older adults and those with
weakened immune systems have been observed (Clayton et al. 2003).

Salmonella can contaminate the food supply and cause illness in three different ways. Salmonella
can be found in animals, making meat, poultry, eggs, and milk common sources of infection.
Bacteria can also enter the environment, possibly through manure or litter, and contaminate fruits
and vegetables grown on farms. Cross-contamination in food service or at home, particularly
between raw poultry and ready-to-eat products such as raw vegetables, can also result in
salmonellosis. Although any strain of Salmonella can be harmful to humans, non-host-adapted
serotypes cause the majority of foodborne illnesses. Human contact with domestic and wild
animals is a significant risk factor for salmonellosis (Asfaw and Genetu 2021, Azanaw et al.
2021).

Staphylococcus aureus food poisoning

Staphylococcus aureus is a spherical bacterium commonly found on human skin and in the nose.
While it normally exists harmlessly as part of the body's microbial community, it can become
pathogenic and cause various infections, including those transmitted through food.
Staphylococcus aureus belongs to the Staphylococcaceae family and can thrive in temperatures
between 7 to 48°C, with optimal growth occurring at 30 to 37°C. It is a facultative anaerobe,

15
capable of growing with or without oxygen, and can tolerate high salt levels, enabling survival in
salty environments (Abdu 2019, Lim et al. 2019).

Under favourable conditions, Staphylococcus aureus can cause foodborne illnesses. When the
bacteria multiply, they produce heat-stable toxins known as enterotoxins, which cause illness
when consumed. Toxin production typically requires a population of around 105 cells per gram
of food and occurs under favourable conditions of time, temperature, pH, water activity, and
atmosphere. Foods rich in proteins, such as meat, poultry, fish, dairy products, and cream-filled
bakery items, facilitate the growth of Staphylococcus aureus (Clayton et al. 2003).

Food contamination can happen in various ways. The bacterium can be easily transferred to food
during preparation through the hands, fingertips, hair, or skin of food handlers. Contamination
can also occur when food handlers with infected wounds or boils handle food. Additionally,
coughing and sneezing by individuals with respiratory infections can contribute to the spread of
Staphylococcus aureus. Livestock can carry the bacterium on their skin and hides, leading to
food contamination during the slaughter process (Abdu 2019, Lim et al. 2019).

Compared to other foodborne illnesses, staphylococcal foodborne illness has a relatively short
incubation period. Symptoms typically appear within 30 minutes to 8 hours after consuming
toxin-contaminated food, with the majority of cases occurring within 2 to 4 hours. Symptoms of
staphylococcal food poisoning include nausea, vomiting, abdominal cramping, sweating, chills,
weakness, shock, shallow breathing, and low body temperature. Recovery from this illness,
which is rarely fatal, usually takes 24 to 48 hours. Determining the true incidence of
staphylococcal foodborne illness is challenging due to various factors, such as inaccurate
information provided by victims during interviews, misdiagnosis resembling other types of food
poisoning, inadequate sample collection for laboratory analysis, and improper laboratory
examination. Enterotoxin A is the most potent and commonly associated with staphylococcal
food poisoning outbreaks, with even a small amount of less than 1µg capable of causing illness
in humans (Abdu 2019, Lim et al. 2019).

Staphylococcus aureus intoxications are often linked to institutions like schools and prisons,
where large quantities of food are prepared and stored before consumption. Poor sanitation
practices and improper handling of time-temperature combinations by workers can lead to
16
product contamination and the growth of Staphylococcus aureus to toxin-producing levels
(Clayton et al. 2003).

Escherichia coli food poisoning

Escherichia coli is a bacterium that naturally inhabits the intestines of humans and warm-
blooded animals. While most types of E. coli are harmless, certain harmful strains can lead to
foodborne illnesses. One well-known pathogenic strain is Escherichia coli O157:H7, which
produces a toxin called Shiga toxin. E. coli bacteria have a rod-shaped structure and belong to
the Enterobacteriaceae family (Asfaw and Genetu 2021, Azanaw et al. 2021).

E. coli thrives within a temperature range similar to the human body temperature, between 37-
42°C. It is a facultative anaerobe, meaning it can grow in both oxygen-rich and oxygen-poor
environments. E. coli can tolerate various pH levels but prefers a neutral pH. It can also survive
in low-moisture environments.

Contamination of food by E. coli can occur through different pathways. The primary source is
faecal contamination, which can happen through contaminated water, soil, or improper food
handling. Common routes of transmission include consuming undercooked ground beef, raw
vegetables, unpasteurized milk, and contaminated water. Cross-contamination during food
preparation and poor hygiene practices can also contribute to the spread of E. coli. Although E.
coli O157:H7 is commonly associated with undercooked ground beef, it can also be found in
other meats, raw milk, raw vegetables, and contaminated water. Other pathogenic strains, such as
Shiga toxin-producing E. coli (STEC), have been linked to outbreaks associated with leafy
greens, sprouts, and unpasteurized juices (Clayton et al. 2003).

Escherichia coli (E. coli) are a diverse bacterium, and certain strains can cause severe foodborne
illnesses. Enterohemorrhagic E. coli (EHEC) is of particular concern. It can be transmitted
through contaminated food, such as undercooked ground beef, contaminated vegetables, and
unpasteurized dairy products. The infective dose of EHEC is relatively low, requiring the
ingestion of only a small number of bacteria (10 to 100 cells) to cause illness. The incubation
period typically ranges from 2 to 10 days. Symptoms of E. coli food poisoning include severe

17
diarrhoea (often with blood), abdominal pain, and, in some cases, kidney failure (Asfaw and
Genetu 2021, Azanaw et al. 2021).

Clostridium perfringens food poisoning

The gram-positive, anaerobic bacterium Clostridium perfringens causes food poisoning. This
bacterium is commonly found in the environment as well as in human and animal intestines. It
produces a variety of toxins that contribute to the pathogenesis of food poisoning. Clostridium
perfringens can produce heat-resistant spores, allowing it to survive at cooking temperatures
(Asfaw and Genetu 2021, Azanaw et al. 2021, Todd et al. 2010).

In comparison to other foodborne pathogens, the infectious dose required to cause Clostridium
perfringens food poisoning is relatively high. In order to develop the illness, a large number of
bacterial cells, typically 106 to 108 cells must be consumed. Toxins produced by bacteria, rather
than bacteria themselves, play a key role in causing symptoms. Clostridium perfringens food
poisoning has a relatively short incubation period, typically ranging from 6 to 24 hours after
consuming contaminated food. The rapid onset of symptoms is caused by the rapid growth of
bacteria in the gastrointestinal tract, which results in the production of toxins (Asfaw and Genetu
2021, Azanaw et al. 2021, Todd et al. 2010).

Clostridium perfringens food poisoning symptoms primarily affect the gastrointestinal system.
Severe abdominal pain with cramping, localized pain, and bloating constitutes some of them.
Watery diarrhoea is a common symptom that may or may not contain blood. Nausea and
vomiting are less common than abdominal pain and diarrhoea. Without medical treatment, the
illness usually resolves within 24 to 48 hours, but dehydration can occur, especially in the elderly
or those with weakened immune systems (Clayton et al. 2003).

Proper food handling and storage techniques are essential for avoiding Clostridium perfringens
food poisoning. Cooking food, particularly meat and poultry, to the proper internal temperature
kills any bacteria present. It is critical to avoid leaving cooked food at room temperature for
extended periods of time and to rapidly cool and refrigerate leftovers. Additionally, maintaining
good hand hygiene, practicing proper sanitation when handling food, and preventing cross-
contamination between cooked and raw foods by using separate utensils, cutting boards and

18
storage containers can significantly reduce the risk of contamination (Asfaw and Genetu 2021,
Azanaw et al. 2021).

Bacillus cereus food poisoning

Bacillus cereus has been linked to two types of foodborne illnesses: emetic and diarrheal
responses each caused by different toxins produced by the microorganism. A large protein causes
the diarrheal response, while a small peptide, cereulide, causes the emetic response and remains
stable even at high temperatures (Clayton et al. 2003).

The diarrheal response symptoms, which are similar to those caused by C. perfringens, include
diarrhoea, abdominal cramps, and tenesmus, and typically appear 6 to 15 hours after ingestion.
The emetic response, on the other hand, causes nausea and vomiting one to six hours after
ingestion, similar to symptoms caused by staphylococcal enterotoxin. Both types of illness
usually clear up within 24 hours, and complications are uncommon (Asfaw and Genetu 2021,
Azanaw et al. 2021, Todd et al. 2010).

B. cereus is commonly found in soil, vegetation, and a variety of food sources, including dairy
products, meats, spices, dried goods, and cereals, particularly rice. Because of its widespread
distribution in the environment, it is occasionally ingested as part of the transient human
intestinal microflora. B. cereus is commonly found in soil, vegetation, and a variety of food
sources, including dairy products, meats, spices, dried goods, and cereals, particularly rice.
Because of its widespread distribution in the environment, it is occasionally ingested as part of
the transient human intestinal microflora. Studies have shown that low numbers of B. cereus are
present in the intestinal tract of over 10% of healthy adults (Asfaw and Genetu 2021, Azanaw et
al. 2021).

Dishes containing corn, corn starch, mashed potatoes, vegetables, meat products, puddings,
soups, and sauces have been linked to outbreaks of B. cereus diarrheal food poisoning. Although
fried or boiled rice dishes are commonly associated with B. cereus emetic-type food poisoning,
other starchy foods such as macaroni and cheese have also been linked to this syndrome. B.
cereus can produce spores that are resistant to cooking. From harvest to primary processing,
these spores are naturally present on many foods (Asfaw and Genetu 2021, Azanaw et al. 2021).

19
While the microorganism is not considered a hazard at the low levels found in food, problems
arise when food is stored at temperatures ranging from 10°C to 55°C for extended periods of
time. B. cereus multiplies rapidly under these conditions, releasing toxins as it grows in food and
the intestinal tract after ingestion. Although more than 105 cells/g are needed to cause illness,
consuming large amounts of B. cereus does not always result in illness. Outbreaks of both types
of B. cereus illness have occurred as a result of poor temperature control or slow cooling of large
amounts of food. These illnesses are intoxications, not infections, and are not contagious
(Clayton et al. 2003) (Asfaw and Genetu 2021).

Shigella food poisoning

Shigellosis, also known as bacillary dysentery, is a bacterial infection caused by Shigella bacteria
such as Shigella dysenteriae, Shigella flexneri, Shigella boydii, and Shigella sonnei. These
bacteria are typically found in the intestines of humans and primates.

While person-to-person transmission via the faecal-oral route is the most common mode of
infection transmission, there have been reports of shigellosis caused by contaminated food.
During outbreaks, the main sources of Shigella are asymptomatic carriers or people recovering
from the disease. Bacteria can live in the digestive tract for months (Asfaw and Genetu 2021,
Azanaw et al. 2021, Todd et al. 2010).

Shigellosis symptoms include diarrhoea, abdominal pain, fever, and vomiting. With bloody
stools, mucus secretion, and dehydration, the severity can range from mild to severe.
Complications of Shigellosis include reactive arthritis, haemolytic uremic syndrome, and Reiter's
disease. Foodborne shigellosis is distinguished by a high rate of infection in a group, a common
source of contamination, and short incubation periods of 12-50 hours. Symptoms typically last 3-
14 days. Certain Shigella strains can be fatal in some cases, with a mortality rate of 10-15%.
Individuals frequently become asymptomatic carriers during the recovery phase, which can last
at any point from a few days to several months. Ingestion of as few as 10-100 Shigella
microorganisms has been shown to cause illness. The disease develops when bacteria invade the
intestinal lining. Shigella can produce enterotoxins known as Shiga toxins, which can inhibit
mRNA transcription and cause endothelial cell death. Shigella infections are frequently
accompanied by secondary infections with other pathogens (Abdu 2019, Lim et al. 2019).
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Listeria monocytogenes

Listeria monocytogenes is a bacterium that causes listeriosis, a severe infection. It can be found
in a variety of environments, including soil, plants, and water. It has a rod-shaped structure and is
a member of the Listeriaceae family. Listeria, unlike many other foodborne bacteria, can survive
and multiply in cold temperatures, as well as tolerate a wide range of pH levels and high salt
concentrations. It can also produce biofilms, which aid in its survival in food processing
environments (Asfaw and Genetu 2021, Azanaw et al. 2021).

People with compromised immune systems are more likely to develop severe listeriosis from
virulent Listeria. Among Listeria species, only L. monocytogenes is consistently pathogenic.
Listeriosis is a potentially fatal infection that primarily affects the elderly and newborns. Listeria,
on the other hand, has recently been discovered to cause gastrointestinal illness in healthy adults,
typically more than 12 hours after ingestion. Large amounts of L. monocytogenes are usually
associated with this milder form of illness. In humans, just 1,000 bacteria cells can cause flu-like
symptoms (fatigue, diarrhoea, and mild fever). Some people may have no or only mild
symptoms. A carrier state can also form, with 1-10% of asymptomatic people becoming L.
monocytogenes excretors (Abdu 2019, Lim et al. 2019).

2.4.2 Antibiotic resistance

Antibiotic resistance is a growing public health concern around the world, and bacterial food
contamination is a major contributor to its development and spread. Antibiotic resistance
happens when bacteria evolve and become resistant to the drugs that are designed to kill them.
Resistance can develop through a variety of mechanisms, such as genetic mutations or the
acquisition of resistance genes from other bacteria. Antibiotic resistance has serious implications
for public health because it reduces antibiotic effectiveness, making infections more difficult to
treat and increasing the risk of complications and mortality (Banik et al. 2019, Muyanja et al.
2011).

A variety of factors contribute to the emergence and spread of antibiotic resistance in foodborne
bacteria. Antibiotic overuse and misuse in both human and veterinary medicine is a significant
factor. Antibiotic use in livestock farming, for example, promotes the emergence and spread of

21
resistant bacteria in animals and their products, including meat and dairy products. When
humans consume these contaminated food products, the resistant bacteria can colonize the
human gut and potentially transfer their resistance genes to other bacteria (Banik et al. 2019,
Eromo et al. 2016).

In addition to antibiotic misuse, poor hygiene practices and inadequate food safety measures can
contribute to the contamination of food with antibiotic-resistant bacteria. Inadequate sanitation
during food processing, poor storage conditions, and cross-contamination in food preparation
and handling can all introduce resistant bacteria into the food chain. Once introduced, these
bacteria can multiply and spread, posing a risk to consumers (Eromo et al. 2016).

Several studies have been conducted to investigate the antibiotic susceptibility patterns of
foodborne bacterial pathogens. These studies provide important information about the prevalence
and distribution of antibiotic resistance in foodborne bacteria. For example, studies have revealed
that strains of Salmonella and Campylobacter, both common foodborne pathogens, frequently
exhibit resistance to multiple antibiotics, including those commonly used in human medicine.
This discovery emphasizes the risk of antibiotic-resistant foodborne infections and the limited
treatment options available (Banik et al. 2019, Muyanja et al. 2011, Rane 2011, Wer et al. 2020).

Street food, which is popular in many countries, has also been linked to the spread of antibiotic
resistance among foodborne bacteria. These foods are frequently prepared and sold under
unregulated conditions, with poor sanitation and little supervision. As a result, they can act as
reservoirs for antibiotic-resistant bacteria. Various resistant strains have been identified in street-
vendor foods, including those resistant to critical antibiotics used in human medicine.
Consumption of such contaminated foods may result in the transmission of these resistant
bacteria to consumers, further fuelling the spread of antibiotic resistance (Banik et al. 2019,
Muyanja et al. 2011).

2.5 Microbiological safety of street-vended ready-to-eat foods

2.5.1 Street-vended ready-to-eat foods and their popularity

22
Street-vended ready-to-eat foods have gained significant popularity in developing countries due
to their convenience, affordability, and availability. These foods, which are sold by vendors in
open-air markets or mobile carts, provide a variety of advantages to consumers on the go. They
are frequently less expensive than dining out or shopping at supermarkets, making them
accessible to people with limited financial resources. Street foods offer a diverse culinary
experience as well, reflecting the region's cultural heritage and offering distinct flavors and
cooking techniques. Their accessibility and convenience make them a quick and easy option for
busy people, while also serving as social spaces for community interaction. However, there are
food safety and hygiene issues that must be addressed in order to ensure the safety of street-
vendor foods (Abdu 2019, Lim et al. 2019).

2.5.2 Potential microbiological hazards associated with street-vended foods

Street-vended foods are susceptible to microbiological hazards due to various factors such as
preparation, handling, and storage in unregulated environments. These hazards can lead to the
contamination of street-vended foods with harmful microorganisms, posing a risk of foodborne
illnesses for consumers (Asfaw and Genetu 2021, Azanaw et al. 2021).

One significant hazard is the poor personal hygiene practices among street food vendors.
Insufficient hand washing facilities, limited access to clean water, and inadequate knowledge of
proper hygiene techniques can result in bacteria transferring from the vendors' hands to the food
they handle. This can introduce pathogens like Salmonella, E. coli, and Staphylococcus aureus
into the food, posing health risks (Asfaw and Genetu 2021, Azanaw et al. 2021, Todd et al.
2010).

Contaminated water used during food preparation is another hazard. Street vendors may rely on
untreated or poorly treated water sources, which can introduce pathogens into the food. Cross-
contamination is also a concern due to limited space and equipment, increasing the likelihood of
transferring pathogens between raw and cooked foods. Improper storage temperatures further
contribute to microbiological hazards. Without refrigeration, street vendors may store perishable
ingredients and cooked foods at temperatures that promote bacterial growth. This can lead to
rapid multiplication of pathogens and an increased risk of foodborne illness (Abdu 2019, Lim et
al. 2019).
23
The quality and safety of raw ingredients used in street-vended foods can vary, as vendors may
source them from unregulated suppliers. Contaminated or spoiled ingredients can introduce
pathogens into the food, compromising its safety. Additionally, the lack of effective regulatory
oversight and inspections in street food environments can exacerbate microbiological hazards.
Without proper enforcement of food safety standards, vendors may not adhere to hygienic
practices, increasing the chances of microbial contamination (Abdu 2019, Lim et al. 2019).

2.5.3 Health risks posed by these pathogens to consumers

Because of the presence of various pathogens, street-vended ready-to-eat foods pose a significant
risk to consumer health. Salmonella and Campylobacter, for example, are commonly found in
such foods and can cause foodborne infections, which can result in symptoms such as fever,
abdominal pain, and diarrhoea. Staphylococcus aureus is another pathogen that can produce
toxins that cause food poisoning, resulting in symptoms such as vomiting, diarrhoea, and nausea.
Furthermore, Escherichia coli have been linked to severe gastrointestinal illnesses such as
kidney failure and bloody diarrhoea. These examples highlight the critical importance of
ensuring the microbiological safety of street-vendor ready-to-eat foods (Asfaw and Genetu 2021,
Azanaw et al. 2021).

2.5.4 Review of previous studies on the microbiological contamination of street-vended

foods

Numerous research studies have investigated the presence of harmful bacteria in street-
vended ready-to-eat foods. For example, a study in Ethiopia discovered bacteria such as
Salmonella and Staphylococcus aureus in street-vended foods such as local bread, raw fish, chili,
avocado, and cooked potato (Clayton et al. 2003). Similarly, a study in Kenya identified Bacillus
cereus, Clostridium perfringens, Staphylococcus aureus, and Salmonella spp. in street-vended
foods (Eromo et al. 2016). Comparable findings have been reported in studies conducted in other
countries such as Uganda, Malaysia, and Bangladesh, emphasizing the importance of improving
food safety practices and regulations in street food vending (Asfaw and Genetu 2021, Azanaw et
al. 2021, Todd et al. 2010).

24
Measures such as promoting proper hygiene practices such as hand washing, appropriate storage,
and cooking temperatures can help reduce the risk of microbiological contamination. Public
awareness campaigns and educational initiatives can also help promote safe food handling
practices among street food vendors (Eromo et al. 2016). Furthermore, regular monitoring and
inspection of street food vending facilities can aid in identifying and addressing potential food
safety hazards (Asfaw and Genetu 2021, Azanaw et al. 2021, Todd et al. 2010).

2.6 Strategies to improve microbiological quality of street-vended foods

2.6.1 Interventions and strategies to ensure the safety of street-vended foods

Improving the microbiological quality of street-vended foods necessitates the implementation of

a variety of interventions and strategies to address the challenges associated with microbial
contamination (Boels 2014). The development and enforcement of food safety regulations and
standards specific to street food vendors is a crucial first step. Personal hygiene, food handling
practices, equipment sanitation, and proper food storage and display should all be addressed in
these regulations. Governments and regulatory bodies must collaborate closely with street
vendors to ensure compliance with these standards through regular inspections, training
programs, and public awareness campaigns (Nani 2020).

Education and training initiatives aimed at street food vendors are critical to improving food
safety practices. Educating vendors on proper food handling, hygiene, and storage techniques
can significantly reduce the risk of microbial contamination. Training programs should cover
topics like hand washing, safe water usage, temperature control, cross-contamination prevention,
and the importance of sourcing safe ingredients. Additionally, encouraging the use of hazard
analysis and critical control points (HACCP) principles can assist vendors in identifying and
mitigating potential microbial hazards (Adhikani 2012, Boels 2014).

Access to clean water and adequate sanitation facilities is critical for maintaining the
microbiological quality of street-vendor foods. Governments and local governments thus should
invest in improving water supply systems and sanitation infrastructure in street food areas.
Provision of clean water for food preparation, hand washing stations with soap and drying

25
facilities, and proper waste management systems are essential to minimize the risk of microbial
contamination (Nani 2020).

Technology can also help improve food safety in street vendors. Mobile applications and digital
platforms can help to monitor vendors' compliance with food safety standards in real time. These
platforms can allow vendors to report on their daily activities, such as food handling practices,
ingredient sourcing, and temperature control. Authorities can use this information to identify
potential sources of contamination and provide targeted interventions (Nani 2020).

Collaboration among stakeholders is critical in addressing the issues associated with microbial
contamination in street-vendor foods. To develop comprehensive strategies, governments,
regulatory bodies, public health agencies, local communities, and vendors should work
collaboratively. This collaboration may involve sharing best practices, conducting joint training
programs, and establishing communication channels for reporting food safety concerns or
outbreaks (Adhikani 2012, Boels 2014).

2.6.2 Gaps and challenges in addressing microbial contamination of street-vended foods

Despite efforts, there are a number of gaps and challenges in addressing microbial contamination
in street-vended foods. Limited resources and capacity are significant impediments, particularly
in low-income communities. Many street vendors operate in unregulated or informal
environments, making it difficult to implement and enforce food safety standards. Furthermore,
cultural practices and traditions may have an impact on food handling practices, necessitating
context-specific interventions that are sensitive to local norms and customs (Adhikani 2012,
Boels 2014).

In order to tackle the gaps and challenges in improving the microbiological quality of street-
vended foods, a multifaceted approach is required. It entails a combination of regulatory
measures, education and training programs, infrastructure development, technological
innovations, and stakeholder collaboration. By implementing these strategies, it is possible to
improve food safety practices, reduce microbial contamination, and ensure that consumers
receive safe and nutritious street-vended foods (Adhikani 2012, Boels 2014).

26
2.6.3 The Food and Food Standards Act and regulation of street-vended foods in
Zimbabwe

The Zimbabwe Food and Food Standards Act is essential for regulating the sale of street-vended
foods and protecting consumers from health risks. It sets standards for food safety, hygiene, and
vending site cleanliness that street food vendors must follow. Regulatory authorities, such as the
Zimbabwe Food Standards Authority (ZFDA), are empowered to enforce these regulations and
conduct inspections to ensure compliance (Nani 2020, Madziba 2017). However, despite the
existence of this act, the illegal sale of street-vended foods in Zimbabwe has been on the rise.
One reason for this increase is the high rate of company closures, pushing many people towards
street vending as a means of livelihood. Additionally, street vending offers income opportunities
for the urban poor and allows vendors to bypass the costs and regulations associated with formal
businesses (Madziba 2017).

The rise in illegal street-vended foods poses risks to consumers, including food safety hazards
and microbial contamination. Unregulated vendors may not adhere to proper hygiene practices,
increasing the chances of cross-contamination, improper food storage, and inadequate cooking
temperatures. This raises the risk of foodborne illnesses and endangers consumer health.
Moreover, the lack of standardization and quality control in the illegal street food sector leads to
inconsistent food safety practices and a lack of information for consumers about the safety and
origin of the food they purchase (Madziba 2017).

To address the increase in illegal street-vended foods, a comprehensive approach is needed. This
involves raising awareness among vendors about compliance and assisting them in transitioning
to the formal sector. Simplifying licensing procedures, providing food safety training, and
creating economic opportunities can motivate vendors to operate within the regulated
framework. Strengthening enforcement mechanisms and enhancing the capacity of regulatory
authorities to monitor compliance through inspections are also crucial. Collaboration between
government agencies, local authorities, and stakeholders is necessary to effectively tackle the
challenges of illegal street vending and ensure the safety of street-vended foods. By addressing
the root causes and implementing robust regulatory measures, Zimbabwe can mitigate risks to
consumers and protect public health in the street food sector (Nani 2020, Madziba 2017).

27
2.7 Techniques for detecting foodborne pathogens

The detection of foodborne pathogens is crucial for ensuring food safety and protecting public
health. While modern techniques such as molecular-based methods and biosensors offer
advantages in sensitivity, speed and specificity, traditional methods continue to play an important
role in foodborne pathogen detection. Traditional methods remain commonly utilized alongside
modern techniques for several reasons. First, traditional methods tend to be more cost-effective
since they do not require specialized equipment, reagents or highly trained personnel, making
them accessible to laboratories with limited resources. Second, traditional methods have
standardized protocols that are well-established and internationally recognized, necessitating less
sophisticated infrastructure and expertise. This accessibility renders them suitable for
laboratories in remote or resource-constrained settings (Colacecchio et al. 2017).

Third, regulatory bodies and food safety agencies frequently have standardized protocols and
guidelines founded on traditional methods that have undergone extensive validation and are
widely accepted in the industry. Adherence to these established methods is necessary for
regulatory compliance. Finally, the extensive historical data generated by traditional methods
allows for comparisons and trend analysis over time, facilitating the identification of emerging
pathogens or alterations in pathogen prevalence. The ability to assess long-term data trends is
valuable for monitoring and responding to food safety issues (Dark 2013, Liao et al. 2023).

2.7.1 Culture-based methods

For many years, traditional culture-based techniques such as agar plates, broth cultures, and
biochemical tests have been considered the benchmark for pathogen detection. These methods
enable the separation, recognition, and measurement of pathogens. They offer valuable insights
into the viability and attributes of the pathogens found in a food sample. Culture-based methods
have a long-standing history, are extensively employed, and possess standardized protocols,
rendering them accessible to laboratories with limited resources (Tao et al. 2022, Zhang et al.
2022).

2.7.2 Serological methods

28
Pathogen-specific antigens or antibodies can be detected using serological methods such as
enzyme-linked immunosorbent assays (ELISA). These methods are simple, inexpensive, and can
produce results quickly. Serological methods are especially helpful when screening large
numbers of samples. However, when compared to molecular-based techniques, they may have
limitations in terms of sensitivity and specificity (Zhang et al. 2022).

2.7.3 Polymerase Chain Reaction (PCR)

PCR and its variants, such as real-time PCR (qPCR), are molecular techniques used to amplify
and detect pathogen DNA sequences. PCR produces results with high sensitivity, specificity, and
speed. It can detect pathogens even at low concentrations and can identify multiple pathogens at
the same time. PCR, on the other hand, necessitates specialized equipment, trained personnel,
and the development of specific assays for each target pathogen, making it less accessible in
resource-constrained settings (Zhang et al. 2022).

2.7.4 Next-generation sequencing (NGS)

NGS technologies are capable of offering detailed genomic information about pathogens found
in food samples. They can detect both known and unknown pathogens, identify genetic variants,
and provide information about virulence factors and antimicrobial resistance. NGS has
transformed pathogen detection and surveillance, but its high cost, complex data analysis
requirements, and specialized expertise make it unsuitable for routine use in many laboratories
(Zhang et al. 2022).

2.8 Culture-based methods for assessing bacterial quality of street-vended foods

The evaluation of bacteriological quality in street-vended foods is crucial for ensuring food
safety. These assessment methods involve the identification and analysis of various
microorganisms, with a particular emphasis on hygiene indicator organisms and Staphylococcus,
to determine the bacteriological quality of the food (Dark 2013, Liao et al. 2023). The detection
of hygiene indicator organisms, such as coliforms and Staphylococcus, plays a significant role in
assessing the bacteriological quality of street-vended foods by identifying poor hygiene
practices, evaluating potential fecal contamination, and assessing the risk of pathogens. The

29
presence of these organisms indicates inadequate sanitation and poor hygiene during food
preparation, handling, or storage. Higher counts of these organisms suggest a greater risk of
potential pathogens and microbial contamination. Furthermore, the presence of hygiene indicator
organisms can indicate a higher likelihood of the presence of pathogenic bacteria, such as
Staphylococcus aureus strains that can produce toxins and cause food poisoning (Tao et al. 2022,
Zhang et al. 2022).

The detection of hygiene indicator organisms provides valuable information about the overall
bacteriological quality of street-vended foods. However, it does not directly measure the
presence of specific pathogens. Therefore, additional testing for specific pathogens, such as
Salmonella or Listeria monocytogenes, may be necessary to comprehensively assess the safety of
these foods. Regular monitoring and assessment of hygiene indicator organisms, including
coliforms and Staphylococcus, help identify potential food safety hazards, evaluate the
effectiveness of hygiene practices, and take appropriate corrective actions to ensure the
bacteriological safety of street-vended foods (Zhang et al. 2022).

2.8.1 Detection of Enteric bacteria

Enteric bacteria, which include pathogenic strains such as Salmonella, Shigella, and
Campylobacter, live primarily in the intestines of humans and animals. Their presence in food
indicates faecal contamination and raises the possibility of foodborne illness. Assessing the
presence and levels of enteric bacteria aids in the identification of potential sources of
contamination as well as the evaluation of overall hygiene practices during food preparation and
handling (Tao et al. 2022, Zhang et al. 2022).

Samples are typically collected from street-vendor foods and subjected to culture-based methods
to detect enteric bacteria. In these methods, food samples are inoculated onto selective agar
media for enteric bacteria, such as Salmonella-Shigella agar or Campylobacter blood-free
selective agar. The plates are then incubated under specific conditions to observe the growth of
distinct colonies. Individual colonies are typically confirmed and identified using biochemical
tests or molecular techniques such as polymerase chain reaction (PCR) (Zhang et al. 2022).

2.8.2 Detection of Coliform bacteria

30
Coliform bacteria, a subset of enteric bacteria, are frequently used as faecal contamination
indicators. They include the genera Escherichia, Klebsiella, and Enterobacter, which ferment
lactose to produce gas and acid. While not all coliforms are pathogenic, their presence in food
indicates a possible lapse in hygiene practices and raises concerns about the possible existence of
pathogenic microorganisms (Tao et al. 2022, Zhang et al. 2022).

Coliforms in street-vendor foods are detected using similar culture-based methods as enteric
bacteria. Food samples are plated on selective agar media that promote the growth of coliforms,
such as MacConkey agar or Violet Red Bile agar. The plates are then incubated at specific
temperatures, typically around 35-37°C, and the characteristic colonies are counted. Biochemical
tests, such as the lactose fermentation test, can be used to confirm coliform identity further. This
test determines the bacteria's ability to produce gas and acid from lactose (Dark 2013, Liao et al.
2023).

2.8.3 Detection of Escherichia coli

Escherichia coli (E. coli), a member of the coliform group, is particularly concerning due to
pathogenic strains that can cause severe foodborne illnesses. Enterohemorrhagic E. coli (EHEC)
and enterotoxigenic E. coli (ETEC) strains can cause symptoms ranging from mild diarrhoea to
severe complications such as haemolytic uremic syndrome (HUS). The presence of E. coli in
street-vended foods indicates faecal contamination and highlights potential consumer risks. The
same culture-based methods used for coliforms are used to detect E. coli in street-vendor foods.
Selective agar media such as MacConkey agar or Eosin Methylene Blue agar are used, and the
plates are incubated at 35-37°C. The presence of E. coli is confirmed through biochemical tests
such as the lactose fermentation test and additional tests specific to pathogenic strains, such as
the detection of specific virulence factors or toxins (Tao et al. 2022, Zhang et al. 2022).

2.8.4 Detection of Staphylococcus bacteria

Another important group of hygiene indicator organisms is Staphylococcus bacteria, particularly

Staphylococcus aureus. Because Staphylococcus can be found in human nasal passages, skin,
and hair, its presence in food samples indicates poor personal hygiene. Heat-stable toxins
produced by Staphylococcus aureus can cause foodborne illnesses. As a result, Staphylococcus

31
detection and enumeration aid in assessing the potential risks associated with poor hygiene and
the presence of pathogenic strains (Dark 2013, Liao et al. 2023).

Culture-based methods are used to detect S. aureus in street-vended foods. Food samples are
plated on selective media such as Baird-Parker agar or Mannitol Salt agar, which promote
Staphylococcus growth while inhibiting the growth of other bacteria. The plates are incubated at
appropriate temperatures, typically 35-37°C, and colonies with distinctive morphology are
chosen for further confirmation. Confirmation can include biochemical tests like coagulase
testing or immunological methods like enzyme-linked immunosorbent assay (ELISA) to detect
staphylococcal enterotoxins (Tao et al. 2022, Zhang et al. 2022).

2.9 Methods for analysis of antibiotic resistance among foodborne bacteria

The analysis of antibiotic resistance among foodborne bacteria entails the use of various methods
to assess these bacteria's susceptibility to various antibiotics. These approaches include both
phenotypic and genotypic approaches, and they provide valuable insights into the resistance
mechanisms and patterns displayed by foodborne bacteria (Dark 2013, Liao et al. 2023).

2.9.1 Phenotypic methods

Phenotypic methods are commonly used to determine the antibiotic resistance profile of
foodborne bacteria. The disk diffusion method, also known as the Kirby-Bauer method, is the
most widely used. In this method, paper disks impregnated with different antibiotics are placed
on an agar plate inoculated with bacteria. After that, the plates are incubated, and the growth
inhibition zones around each disk are measured. The size of the inhibition zone indicates whether
the bacteria are susceptible or resistant to the specific antibiotic. Another phenotypic method is
broth microdilution, which involves exposing bacteria to a series of antibiotic dilutions in liquid
media and determining the minimum inhibitory concentration (MIC) that prevents bacterial
growth (Dark 2013, Liao et al. 2023).

2.9.2 Genotypic methods

Specific resistance genes or mutations associated with antibiotic resistance in foodborne bacteria
are detected using genotypic methods (Dark 2013, Liao et al. 2023). Polymerase chain reaction
32
(PCR) is a common genotyping technique. It entails using DNA primers and a DNA polymerase
enzyme to amplify specific target genes. The amplified products can then be tested for the
presence or absence of resistance genes using gel electrophoresis. Real-time PCR, also known as
quantitative PCR, allows for the quantification of target genes and the abundance of resistance
genes in the bacterial population. Furthermore, DNA sequencing techniques like Sanger
sequencing and next-generation sequencing (NGS) can be used to identify known and novel
resistance genes, as well as mutations within these genes (Zhang et al. 2022).

Furthermore, molecular techniques such as DNA microarrays and whole-genome sequencing

(WGS) have changed the way antibiotic resistance in foodborne bacteria is studied. By
hybridizing labelled DNA fragments from bacteria to specific probes on a microarray chip, DNA
microarrays enable the simultaneous detection of multiple resistance genes (Dark 2013, Liao et
al. 2023). This method provides an in-depth look at the resistance genes found in the bacterial
genome. WGS entails sequencing the bacteria's entire genome, allowing for the identification of
resistance genes, mutations, and the analysis of the genetic context surrounding these genes.
WGS data can also be used to investigate the dynamics of antibiotic-resistant bacteria
transmission and to identify potential sources of contamination (Zhang et al. 2022).

33
 CHAPTER 3: METHODS AND MATERIALS

3.1 Study Area

The study was carried out at the University of Zimbabwe is located in Harare, Zimbabwe, at
coordinates 17.78537° South latitude and 31.05311° East longitude. The main campus of the
University of Zimbabwe is situated in the Mount Pleasant neighbourhood in northern Harare, and
it spans 299 hectares in the southern part of Mount Pleasant. On the main campus, there are five
residences for women: Swinton Hall, Complexes 1, 4, and 5, and Carr-Saunders, and four
residences for men: Manfred Hodson Hall, Complex 2, Complex 3, and Manfred Hodson Annex
(Wikipedia)

3.2 Sampling

A cross-sectional study was conducted to assess and evaluate the bacteriological contamination
of selected street-vended ready-to-eat food: cooked chicken, beef, and sausage and vegetable
salad, at the University of Zimbabwe campus, and to determine the antibiotic resistance profiles
of the bacterial isolates. This study was carried between September and December 2023.

3.2.1 Selection of study sites

34
Several street vendors sell ready-to-eat food at the University of Zimbabwe campus. The
numbers of these vendors reach peak during the night when food demand by on campus students,
especially the illegal residents, is high. Ready-to-eat foods are available as takeaways from the
car boot, just outside the main gate of the college which is a paved area. The vendors prepare
their food home and transport them to the campus where they serve them cold from the car boots.

Three vendors were chosen for sample collection, based on the volume of sales and the
frequency at which they visit the campus for their business; the vendors were designated vendors
A, B and C. Those sellers that sell their food everyday were chosen.

3.2.2 Sample collection

The study identified the main food choices among students during the night and gathered
samples from three selected vendors (A, B, and C). Four food items: cooked beef, chicken and
sausage, and vegetable salads, were purchased and served in foam takeout containers, from each
vendor. The sampling was independently done on three different days (day 1, 2 and 3) from 16 to
18 October, and the bacteriological quality analysis was independently done for each day, to
assess variations over time. The samples were procured between 6:00 PM and 8:00 PM.

A total of 36 food samples (12 for each day) were purchased and the foam takeout containers
were placed in sterile polythene container bags and labelled. The labelled samples were
immediately placed in a cooler box with ice packs to maintain their freshness and transported to
the Central Veterinary Laboratory in Harare for further processing. To prevent the growth of
bacteria, the samples were stored in a deep freezer at a temperature of -4°C before analysis
(Quinn et al. 1999).

During the sampling process, strict hygiene practices were monitored, including the use of
sanitary clothing such as gloves and aprons and use of food covers. Additionally, observations
were made and recorded regarding the temperature at which the food was served (hot or cold)
and the overall appearance of the car boot from where the food was being served (Table 3.1).
35
Table 3.1: Observations on food hygiene practices by street vendors A, B and C

Hygiene component Sampling Vendor A Vendor B Vendor C

day

Use of sanitary clothing Day 1 Absent Absent Absent

Day 2 Absent Absent Absent

Day 3 Absent Absent Absent

Temperature at which the Day 1 Cold Cold Cold

food was served
Day 2 Cold Cold Cold

Day 3 Cold Cold Cold

Appearance of the car boot Day 1 Moderately Moderately Moderately

clean clean clean

Day 2 Moderately Moderately Moderately

clean clean clean

Day 3 Moderately Moderately Moderately

clean clean clean

Use of food covers Day 1 Absent Mesh food Absent

covers

Day 2 Absent Mesh food Absent

covers

Day 3 Absent Mesh food Absent

covers

36
The street vendors were also interviewed on the source of their poultry and beef, whether it is
home-reared or purchased and the source of their vegetables, whether they are home-grown, or
purchased (table 3.2).

Table 3.2: Source of meat, sausage and vegetables prepared by street vendors A, B and C

Commodity Vendor A Vendor B Vendor C

Poultry Home-reared Home-reared Supermarket

Beef Local butchery Local butchery Local butchery

Sausage Local butchery Local butchery Supermarket

Vegetables Home-grown Home-grown Supermarket

3.3 Bacteriological Aanalysis

The laboratory procedures on the bacteriological analysis of the food samples were carried out
using the standard operating procedures at the Central Veterinary Laboratory in Harare. All the
protocols were followed without modification.

3.3.1 Isolation and identification of enteric bacteria

Isolation and identification for bacteria was done using the Central Veterinary Laboratory
standard operating procedure version 10 (SOP/BA/005-V10). The isolation was independently
done for each of the three batch samples. Ten (10) grams of each food sample was weighed and
placed into 90mls of peptone water for bacteria enrichment (Quinn et al. 1999). The mixture was
then homogenized and incubated at 37ºC and 44°C for 18-24 hours. After incubation, a loopful
of the incubated samples was cultured onto blood ager and MacConkey for selective isolation of
most enteric bacteria and Rappaport Vasiliadis (RV) for the selective isolation of salmonella

37
(Bratislave et al. 2016).For precision, the agar plates were cultured in replicates of two per
sample. The cultured agar plates and the RV were subsequently incubated aerobically for 24
hours.

After incubation, the plates were examined for the presence of colonies showing characteristic
morphology of enteric bacteria. Representative colonies were then selected based on their
appearance such as different morphology, colour, or texture, and transferred onto fresh agar
plates using the streaking method, to obtain pure cultures (Bratislave et al. 2016). Concurrently, a
loopful of inoculated selenite broth or RV was subcultured onto XLD. The streaked plates were
then incubated aerobically at 37ºC for 18-24 hours.

Enteric bacteria were identified using colony morphology, gram staining, and standard
biochemical tests (figure 1). The sterilized food samples were used as a negative control and
during biochemical testing different ATCC organisms (Escherichia coli ATCC 25922 and
Klebsiella pneumoniae ATCC 13883) were used as positive controls; in cases where positive
controls were unavailable reference text books were used (Quinn et al. 1999).

Biochemical tests

Gram staining

The Gram staining procedure was carried out as per SOP/BA/025 for the isolation and
identification of enteric bacteria in three batch samples. For each sample, a single colony was
picked up from a pure culture using a sterile wire loop and smeared on a slide with a few drops
of water. The colonies were heat fixed on the slides by gently heating them for 20 seconds.

The slides were then flooded with crystal violet stain for 1 minute, and the stains were rinsed off
with sterile water. A few drops of iodine solution were added to each slide, and they were left for
another minute before being washed off with sterile water. Next, a few drops of 95% ethanol
were applied to each smear to decolorize the slides, which were then washed again with sterile
water. Finally, a few drops of safranin solution were placed on each slide as a counterstain for 1
minute. After washing the slides with sterile water, they were blotted, air-dried, and examined
under a light microscope at 400x magnification using immersion oil.

38
The Gram-stained slides were analyzed for the identification of enteric bacteria based on their
staining characteristics. Gram-positive bacteria, appearing purple or blue, were observed, as well
as Gram-negative bacteria, appearing pink or red. The morphological features of the bacteria,
such as their shape (for example, cocci, bacilli) and arrangement (for example, pairs, chains,
clusters), were noted (Quinn et al. 1999).

Indole-motility test

The indole motility test was carried out using the SOP/BA/048 and the sulfur indole motility
(SIM) media. For each bacterial isolate, a single colony was picked up from a pure culture using
a sterile inoculating loop. The inoculating loop was then smeared onto the SIM medium,
ensuring that it reached the bottom of the tube to create a straight stab line (Bratislave et al.
2016). This process was repeated for each bacterial isolate, and the tubes were labeled
accordingly.

The SIM tubes were placed in an incubator set to the appropriate temperature and incubation
time as specified in SOP/BA/048. Typically, the enteric bacteria were incubated at 37°C for 24-
48 hours. After the incubation period, the SIM tubes were carefully observed for the presence of
indole production and motility. A few drops of Kovac's reagent were added to the SIM medium
to react with indole and produce a red or pink color in the presence of indole-positive organisms.
The development of a red or pink color in the SIM medium indicated a positive reaction for
indole production, while the absence of a color change indicated a negative reaction (Tshipamba
et al. 2018).

The SIM medium was also observed for any spreading or diffuse growth radiating from the stab
line to determine motility. The presence of radiating growth indicated positive motility, while the
absence of radiating growth suggested negative motility (Quinn et al. 1999).

The results of the indole and motility tests were recorded for each bacterial isolate according to
the guidelines in SOP/BA/048. These results were later combined with other biochemical and
cultural characteristics to aid in the identification of the enteric bacteria.

Citrate test

39
The citrate test was carried out using the SOP/BA/049 to identify enteric bacteria. A sterile
inoculating loop was used to pick a single colony from a pure culture of each bacterial isolate.
The inoculating loop was then stabbed and streaked onto the surface of the Simmon's citrate
agar, ensuring proper aseptic techniques. The agar plates were incubated at the specified
temperature and duration mentioned in SOP/BA/049. Typically, enteric bacteria were incubated
at 37°C for 24-48 hours.

During the incubation period, the plates were observed for any changes in the colour of the agar
and the growth of the bacterial isolates. The citrate utilization by bacteria was indicated by a
change in the color of the medium from green to blue, indicating a positive citrate test. The
absence of color change indicated a negative citrate test (Tshipamba et al. 2018).

The growth of the bacterial isolates on the agar plates was also monitored. Enteric bacteria
capable of utilizing citrate as a sole carbon source would display growth radiating from the stab
line, whereas those unable to utilize citrate would show limited or no growth (Quinn et al. 1999).

The results of the citrate test were recorded for each bacterial isolate following the guidelines
provided in SOP/BA/049. These results were later analyzed alongside other biochemical and
cultural characteristics to aid in the identification of the enteric bacteria.

Oxidase test

The oxidase test was carried out using the SOP/BA/051 to identify enteric bacteria based on their
ability to produce the enzyme oxidase (Quinn et al. 1999). A sterile inoculating loop was used to
pick a small amount of bacterial growth from a pure culture of each isolate. The inoculating loop
was then touched to an oxidase test strip or a filter paper soaked with the oxidase reagent as
specified in SOP/BA/051.

The test strip or filter paper was observed for any color change. The presence of a purple or dark
blue color within a specified time indicated a positive oxidase test, suggesting the presence of the
oxidase enzyme. A lack of color change within the given time indicated a negative result
(Bratislave et al. 2016, Tshipamba et al. 2018, Quinn et al. 1999).

40
The results of the oxidase test were recorded for each bacterial isolate according to the guidelines
provided in SOP/BA/051. These results were later analyzed alongside other biochemical and
cultural characteristics to aid in the identification of the bacteria.

Urease test

The urease test was carried out using the SOP/BA/056 to identify enteric bacteria based on their
ability to produce the enzyme urease (Quinn et al. 1999). A sterile inoculating loop was used to
pick a small colony from a pure culture of each isolate. The inoculating loop was then streaked
onto a urease agar medium as specified in SOP/BA/056. The agar plates were subsequently
incubated at 37°C for 24-48 hours.

During the incubation period, the plates were being observed for any changes in the color of the
agar surrounding the bacterial growth. Urease-positive bacteria produced the enzyme urease,
which hydrolyzed urea in the medium, leading to the production of ammonia. This resulted in a
color change of the agar from its original color (usually yellow) to a pink or magenta color
(Quinn et al. 1999). Urease-negative bacteria did not produce urease, and the agar color
remained unchanged.

The results of the urease test were recorded for each bacterial isolate according to the guidelines
provided in SOP/BA/056. These results were later analyzed alongside other biochemical and
cultural characteristics to aid in the identification of the bacteria.

Triple sugar iron test

The triple sugar iron (TSI) test was conducted using SOP/BA/054 to differentiate and identify
enteric bacteria based on their ability to ferment sugars and produce hydrogen sulfide gas (Quinn
et al. 1999).

During the procedure, a sterilized inoculating loop was used to pick a single colony from a pure
culture of each bacterial isolate. The loop was then used to streak the surface of a TSI agar slant,
following the specific instructions outlined in SOP/BA/054. Subsequently, the TSI agar slants
were incubated at the recommended temperature and duration as mentioned in the SOP/BA/054,
typically at 37°C for 18-24 hours.
41
After the incubation period, the TSI agar slants were carefully examined for various
characteristics, including colour changes and gas production. The TSI medium contained three
sugars: glucose, lactose, and sucrose, along with a pH indicator and ferrous sulfate. The
appearance of the slant provided insights into the bacteria's sugar fermentation ability and gas
production (Tshipamba et al. 2018).

The observations included colour changes in the medium, such as a shift to yellow (acidic) or red
(alkaline), as well as the presence or absence of cracks or lifting of the agar due to gas
production. The formation of a black precipitate (ferrous sulfide) indicated the production of
hydrogen sulfide gas. The results were accepted when there was hydrogen sulfide (H₂S)
production from the Salmonella spp of the quality control procedure and no H₂S production
from the control of [Link].

Lysine decarboxylase test

Lysine decarboxylase (LDC) test was performed following the established protocol in
SOP/BA/052 to differentiate salmonella and other members of the enterobacteriaceae (Quinn et
al. 1999). Using a sterile inoculating loop, a single colony from each bacterial isolate was picked
from a pure culture. The loop was then used to inoculate a tube containing LDC broth, as
specified in the standard operating procedure (SOP/BA/052).

The inoculated tubes were incubated at 37°C for 18-24 hours. After the incubation period, the
tubes were observed for any visible changes. A purple color development indicated a positive
result for lysine decarboxylation, while the absence of color change indicated a negative result.
To confirm the results, the tubes were gently agitated to mix the contents and observe for any gas
production, indicated by the formation of bubbles or turbidity in the medium.

Results were accepted when a purple colour was observed in both suspended with Salmonella
typhimurium ATCC 14028 controls and if decarboxylase turns yellow in the tube inoculated with
Proteus control in the control procedure.

Data interpretation

42
 The results of the biochemical tests were analyzed using the identification key as per the Central
veterinary laboratory SOP/BA005-V10 (figure 3.1), to determine the main enteric pathogens
from the food samples.

Gram staining SOP/BA/025

+ve: cocci -ve: rods

Catalase SOP/BA/05 Lactose fermenters Non-lactose fermenter

Indole Mortility Citrate

SOP/BA/0 SOP/BA/0 SOP/B
48 51 A/049
-ve: +ve:
Streptococci Staphylococci [Link] + (+) -
Klebsiella sp - - +

Citrobacter sp + + +
-ve: CNS Coagulase SOP/BA/030
Enterobacter - + +
sp

+ve Oxidase

MSA

+ve

-ve
-ve: [Link] +ve: [Link] Sugars tests

SOP [Link] Proteus Salmonella

Urea SOP/BA/056 + -
43 TSI SOP/BA/054 + +
Lys/Dec SOP/BA/052 - +
Figure 3.1: Biochemical tests species identification key for main pathogenic bacteria (Source:
CVL SOP/BA 005-V10)

3.3.2 Isolation of staphylococcal aureus

Staphylococcal aureus was isolated for three batch samples using the SOP/BA/043. The isolation
was independently done for each of the three batch samples. Ten (10) grams of each food sample
was weighed and placed into 90mls of peptone water for bacteria enrichment (Quinn et al. 1999).
The mixture was then homogenized and incubated at 37ºC and 44°C for 18-24 hours. After
incubation, a loopful of the incubated samples was cultured onto mannitol salt agar (MSA)
plates. The inoculated plates were then inverted and placed in an incubator set at 35-37°C for 24-
48 hours (Bratislave et al. 2016, Tshipamba et al. 2018).

After the incubation period [Link] colonies were identified as small colonies surrounded by
yellow zones. Suspected colonies of staphylococcus aureus were confirmed using gram staining
and biochemical tests, catalase and coagulase tests, (figure 1). The results were accepted as
[Link] when they conformed to the quality control biochemical tests. Quality control for
[Link] was done to show that mannitol salt agar (MSA) support the growth of the bacterium,
and is sterile. For positive control and fertility test, each sample was spiked with [Link]
(ATCC 33862) and cultured on MSA. After incubation, the observed colonies were confirmed
using biochemical tests (figure 3.1). For negative control, sterile PBS was used as a sample, and
forb sterility check an uninoculated agar plate was incubated along with the cultured plates.

3.3.3 Estimation of total coliforms and [Link] counts

The total coliforms and [Link] counts were determined for three batch samples using the
SOP/BA/002. One gram of each sample was transferred into universal bottles containing 9 ml of
buffered phosphate saline (PBS) and homogenized. Serial dilutions were prepared by transferring
1 mL from the previous dilution tube into the next tube containing sterile PBS, up to the dilution
factor 10⁻⁴.
44
Using a sterile pipette, 0.1 mL of each dilution was aseptically spread onto the surface of the
MacConkey agar plates; two plates were included per each dilution. Care was taken to ensure
even distribution of the sample over the agar surface. The inoculated plates were then inverted
and placed in an incubator at 37°C and 44°C for 18-24 hours (Bratislave et al. 2016, Tshipamba
et al. 2018).

After the incubation period, coliforms colonies were identified as pink colonies in agar plates
incubated at 37°C and pink colonies in plates incubated at 44°C were attributed to the growth of
[Link]. Plates that yielded between 30 and 300 colonies were read, for greater accuracy. The
colony count was obtained as an average of the colonies of the two plates inoculated with the
selected dilution. The colony counts were expressed as colony-forming units per milliliter
(CFU/mL) of the original sample by multiplying the number of colonies counted by the
respective dilution factor and the conversion factor (Quinn et al. 1999).

Suspected colonies of coliforms and [Link] were counted and were confirmed using gram
staining and biochemical tests. The results were accepted as coliforms when they conformed to
the quality control biochemical tests for [Link], Klebsiella, Enterobacter and Citrobacter (Figure
1). Quality control was done to show that MacConkey agar support the growth of the coliforms,
and is sterile. For positive control and fertility test, each sample was spiked with [Link] (ATCC
25922) and Klebsiella pneumonia (ATCC 13883) and cultured on MacConkey agar plates. After
incubation, the observed colonies were confirmed using biochemical tests (figure 1). For
negative control, sterile PBS was used as a sample, and forb sterility check an uninoculated agar
plate was incubated along with the cultured plates.

3.4 Antibiotic susceptibility test

The antimicrobial susceptibility testing (AST) was conducted for each bacteria isolate from the
three batch samples, using the Kirby-Bauer disc diffusion method, to determine the susceptibility
patterns of the isolated bacterial species to antibiotics following the procedures outlined in
SOP/BA/102 as described by Koeth et al. 2015.

45
Using a sterile inoculating loop, a few colonies of the bacterial isolates were picked and
transferred into a tube containing sterile saline solution to prepare a turbidity equivalent to a 0.5
McFarland standard.

Sterile petri plates containing Mueller Hinton agar (MHA) were labeled with the appropriate
sample identification and antibiotic disks to be tested. Using a sterile swab, the prepared bacterial
suspension was evenly spread onto the surface of the agar plates, ensuring complete coverage.

Following the guidelines provided in SOP/BA/102 antibiotic impregnated discs (Oxoid) were
applied onto the MHA plates using an antibiotic dispenser. The antibiotics penicillin 30µg,
Ampicillin 10µg, Gentamycin 30µg, Erythromycin 15 µg and Tetracycline 30µg (Oxoid,
Germany) were used.

The inoculated plates with antibiotic disks were inverted and placed in an incubator and
incubated for 24 hours at 37ºC specified in SOP/BA/102. After the incubation period, the plates
were carefully examined for the presence of inhibition zones around the antibiotic disks. Using a
ruler, the diameters of the inhibition zones were measured according to the guidelines specified
in SOP/BA/102, and the results were interpreted as resistant, intermediate, or sensitive to the
different antibiotics used according to reference interpretive criteria (Quinn et al. 1999).

3.6 Data Analysis

The results of the biochemical tests were analyzed using the identification key as per the Central
veterinary laboratory SOP/BA005-V10 (figure 3.1), to determine the main enteric pathogens
from the food samples. Percentage prevalence of isolated bacteria was calculated as the number
of times the bacteria detected over the total number of bacterial isolates detected. Pie charts were
then used to represent the overall prevalence rates of enteric bacteria isolated. A bar chart was
used to compare the mean, total coliform counts and E. coli counts between the food samples.

46
 CHAPTER 4: RESULTS

4.1 Isolation and Identification of enteric bacteria

The results of the biochemical tests were analyzed using the identification key as per the Central
veterinary laboratory SOP/BA005-V10 (figure 3.1), to determine the main enteric pathogens
from the food samples.

4.1.1 Isolation and Identification of enteric bacteria on day 1

Vendor A's cooked beef sample tested positive for E. coli, which appeared as negative rod-
shaped bacteria in the Gram stain. The biochemical tests indicated positive results for indole and
motility, but negative results for citrate utilization. Similarly, another sample from vendor A, the
cooked chicken, showed the presence of Salmonella. The bacteria displayed negative rod-shaped
morphology and did not produce oxidase or urease. However, it tested positive for the lysine
decarboxylase test and showed positive results in the TSI test. The sausage from vendor A also
contained E. coli, similar to the cooked beef sample. The bacteria displayed negative rod-shaped
morphology and stained negatively in the Gram stain. The biochemical tests showed positive
results for indole and motility, while citrate utilization was negative (table 4.1).

In the case of the vegetable salad from vendor A, both E. coli and Salmonella were found. The
bacteria exhibited negative rod-shaped morphology and stained negatively in the Gram stain.
The biochemical tests showed positive results for indole and motility, while citrate utilization
was negative. Additionally, the bacteria tested negative for oxidase and urease but showed
positive results in the TSI test and lysine decarboxylase test (table 4.1).

Vendor B's samples were contaminated with Staphylococcus aureus. The cooked beef, cooked
chicken, and vegetable salad samples all contained positive cocci-shaped bacteria. The
biochemical tests indicated positive results for catalase and coagulase. No bacteria were isolated
from the sausage sample obtained from vendor B. Similarly there was no bacterial growth

47
 observed in the cooked beef, cooked chicken, sausage, or vegetable salad sample from the
vendor C (table 4.1).
Table 4.1: Colony morphology, gram stain and biochemical tests characterization of
isolated bacteria from vendors A, B and C on day 1

Vendor Food Morphology of Gram staining Biochemical tests Tentative

sample bacteria bacteria
Vendor Cooked Blood agar: Grey Negative rods Indole (+); Motility (+); E. coli
A beef round large non- Citrate (-)
haemolytic colonies;
MacConkey agar:
Round large pink
colonies
Cooked Xylose lysine dextrose Negative rods Oxidase (-); Urea (-); Salmonella
chicken agar: pink to red TSI (+); Lysine
colonies with a black decarboxylase test (+)
center
Sausage Blood agar: Grey Negative rods Indole (+); Motility (+); E. coli
round large non- Citrate (-)
haemolytic colonies;
MacConkey agar:
Round large pink
colonies
Vegetable Blood agar: Xylose Negative Negative Indole Oxidase (-); E. coli;
salad Grey round lysine rods rods (+); Urea (-); TSI Salmonella
large non- dextrose Motility (+); Lysine
haemolytic agar: (+); decarboxylase
colonies; pink to Citrate test (+)
MacConkey red (-)
agar: colonies
Round with a
large pink black
48
 colonies center
Vendor Cooked Mannitol salt agar: Positive cocci Catalase (+); Coagulase S. aureus
B beef Small colonies (+)
surrounded by yellow
zones
Cooked Mannitol salt agar: Positive cocci Catalase (+); Coagulase S. aureus
chicken Small colonies (+)
surrounded by yellow
zones
Sausage No bacteria isolated No bacteria isolated No bacteria isolated No
bacteria
isolated
Vegetable Mannitol salt agar: Positive cocci Catalase (+); Coagulase S. aureus
salad Small colonies (+)
surrounded by yellow
zones
Vendor Cooked No bacteria isolated No bacteria isolated No bacteria isolated No
C beef bacteria
isolated
Cooked No bacteria isolated No bacteria isolated No bacteria isolated No
chicken bacteria
isolated
Sausage No bacteria isolated No bacteria isolated No bacteria isolated No
bacteria
isolated
Vegetable No bacteria isolated No bacteria isolated No bacteria isolated No
salad bacteria
isolated

49
4.1.2 Isolation and Identification of enteric bacteria on day 2

Vendor A's cooked beef sample showed the presence of E. coli and Proteus bacteria. The
appearance of the bacteria on blood agar and MacConkey agar indicated grey round large non-
haemolytic colonies and round large pink colonies, respectively. Gram staining revealed the
presence of negative rods. Indole and motility tests were positive, while citrate utilization test
was negative. Other specific tests provided further information about the bacteria. E. coli was
also identified in vendor A's cooked chicken and vegetable salad samples, with similar
morphological characteristics and biochemical test results. No bacteria were found in the sausage
sample of vendor A (table 4.2).

Vendor B's cooked beef sample contained Staphylococcus aureus, which appeared as small
colonies surrounded by yellow zones on Mannitol salt agar. Gram staining indicated positive
cocci, and biochemical tests confirmed the presence of catalase and coagulase. No bacteria were
isolated from the cooked chicken sample of vendor B. E. coli was identified in the sausage
sample, similar to the findings in vendor A's sausage sample. Vendor B's vegetable salad sample
showed the presence of Staphylococcus aureus, with morphological and biochemical
characteristics matching those of the cooked beef sample (table 4.2).

Vendor C's cooked beef sample exhibited Proteus and Klebsiella bacteria. The appearance on
blood agar and MacConkey agar was described as grey swarming non-haemolytic colonies and
round pale colonies, respectively. Gram staining indicated negative rods. Biochemical tests
provided further details, including negative results for indole and motility and positive result for
citrate utilization. Vendor C's cooked chicken sample contained E. coli, showing similar
morphological and biochemical characteristics to the other samples. No bacteria were found in
the sausage and vegetable salad samples of vendor C (table 4.2).

Table 4.2: Colony morphology, gram stain and biochemical tests characterization of
isolated bacteria from vendors A, B and C on day 2

50
Vendor Food Morphology of bacteria Gram Biochemical tests Tentative
sample staining bacteria

Vendor Cooked Blood agar: Blood agar: Negative Indole (+); Oxidase (-); E. coli;
A beef Grey round Grey rods Motility (+); Urease (+); Proteus
large non- swarming Citrate (-) TSI (+);
haemolytic non- Lysine
colonies; haemolytic decarboxylase
MacConkey colonies; test (-)
agar: MacConkey
Round agar:
large pink Round pale
colonies colonies

Cooked Blood agar: Grey round Negative Indole (+); Motility (+); E. coli
chicken large non-haemolytic rods Citrate (-)
colonies; MacConkey
agar: Round large pink
colonies

Sausage No bacteria isolated No No bacteria isolated No

bacteria bacteria
isolated isolated

Vegetable Blood agar: Grey round Negative Indole (+); Motility (+); E. coli
salad large non-haemolytic rods Citrate (-)
colonies; MacConkey
agar: Round large pink
colonies

Vendor Cooked Mannitol salt agar: Small Positive Catalase (+); Coagulase (+) S. aureus
B beef colonies surrounded by cocci

51
 yellow zones

Cooked No bacteria isolated No No bacteria isolated No

chicken bacteria bacteria
isolated isolated

Sausage Blood agar: Grey round Negative Indole (+); Motility (+); E. coli
large non-haemolytic rods Citrate (-)
colonies; MacConkey
agar: Round large pink
colonies

Vegetable Mannitol salt agar: Small Positive Catalase (+); Coagulase (+) S. aureus
salad colonies surrounded by cocci
yellow zones

Vendor Cooked Blood agar: Blood agar: Negative Oxidase (-); Indole (-); Proteus;
C beef Grey Grey rods Urease (+); Motility (-); Klebsiella
swarming mucoid TSI (+); Citrate (+)
non- non- Lysine
haemolytic haemolytic decarboxylase
colonies; colonies; test (-)
MacConkey MacConkey
agar: agar:
Round pale Round
colonies large pink
colonies

Cooked Blood agar: Grey round Negative Indole (+); Motility (+); E. coli
chicken large non-haemolytic rods Citrate (-)
colonies; MacConkey
agar: Round large pink
colonies

52
 Sausage No bacteria isolated No No bacteria isolated No
bacteria bacteria
isolated isolated

Vegetable No bacteria isolated No No bacteria isolated No

salad bacteria bacteria
isolated isolated

4.1.3 Isolation and Identification of enteric bacteria on day 3

Vendor A's cooked beef sample contained E. coli and Proteus bacteria. These bacteria formed
large, non-hemolytic grey round colonies on blood agar and large pink colonies on MacConkey
agar. They were gram-negative rod-shaped bacteria and tested positive for indole production and
motility. The bacteria did not utilize citrate but showed positive results for urease and triple sugar
iron (TSI) tests. The lysine decarboxylase test was negative. No bacteria were isolated from
Vendor A's cooked chicken and sausage samples (table 4.3).

In Vendor A's vegetable salad sample, E. coli and Salmonella bacteria were identified. They
formed large, non-hemolytic grey round colonies on blood agar, large pink colonies on
MacConkey agar, and pink to red colonies with a black center on Xylose lysine dextrose agar.
These gram-negative rod-shaped bacteria tested positive for indole production and motility. They
did not utilize citrate or urea but showed positive results for the TSI and lysine decarboxylase
tests. No bacterial isolates were obtained from Vendor B's cooked beef, cooked chicken, sausage,
or vegetable salad samples (table 4.3).

Vendor C's cooked beef sample yielded Proteus and Klebsiella bacteria. They formed swarming,
non-hemolytic grey colonies on blood agar and pale round colonies on MacConkey agar. These
gram-negative rod-shaped bacteria tested negative for indole production and motility. They
showed positive results for urease and TSI tests, while the lysine decarboxylase test was

53
 negative. They utilized citrate. No bacteria were isolated from Vendor C's cooked chicken,
sausage, and vegetable salad samples (table 4.3).

Table 4.3: Colony morphology, gram stain and biochemical tests characterization of
isolated bacteria from vendors A, B and C on day 3

Vendor Food Morphology of bacteria Gram Biochemical tests Tentative

sample staining bacteria

Vendor Cooked Blood agar: Blood agar: Negative Indole (+); Oxidase (-); E. coli;
A beef Grey round Grey rods Motility (+); Urease (+); Proteus
large non- swarming Citrate (-) TSI (+);
haemolytic non- Lysine
colonies; haemolytic decarboxylase
MacConkey colonies; test (-)
agar: MacConkey
Round agar:
large pink Round pale
colonies colonies

Cooked No bacteria isolated No No bacteria isolated No

chicken bacteria bacteria
isolated isolated

Sausage No bacteria isolated No No bacteria isolated No

bacteria bacteria
isolated isolated

Vegetable Blood agar: Xylose Negative Indole (+); Oxidase (-); E. coli;
salad Grey round lysine rods Motility (+); Urea (-); TSI Salmonella
large non- dextrose Citrate (-) (+); Lysine
haemolytic agar: pink decarboxylase
colonies; to red test (+)
MacConkey colonies
54
 agar: with a black
Round center
large pink
colonies

Vendor Cooked No bacteria isolated No No bacteria isolated No

B beef bacteria bacteria
isolated isolated

Cooked No bacteria isolated No No bacteria isolated No

chicken bacteria bacteria
isolated isolated

Sausage No bacteria isolated No No bacteria isolated No

bacteria bacteria
isolated isolated

Vegetable No bacteria isolated No No bacteria isolated No

salad bacteria bacteria
isolated isolated

Vendor Cooked Blood agar: Blood agar: Negative Oxidase (-); Indole (-); Proteus;
C beef Grey Grey rods Urease (+); Motility (-); Klebsiella
swarming mucoid TSI (+); Citrate (+)
non- non- Lysine
haemolytic haemolytic decarboxylase
colonies; colonies; test (-)
MacConkey MacConkey
agar: agar:
Round pale Round
colonies large pink
colonies

Cooked No bacteria isolated No No bacteria isolated No

55
 chicken bacteria bacteria
isolated isolated

Sausage No bacteria isolated No No bacteria isolated No

bacteria bacteria
isolated isolated

Vegetable No bacteria isolated No No bacteria isolated No

salad bacteria bacteria
isolated isolated

4.2 Prevalence of isolated bacteria

The overall prevalence rates of the bacterial species isolated were calculated and are shown in
Fig . Escherichia coli has the highest prevalence rate of 41.7% (10 out of 24 isolates) followed
by S. aureus 20.8% (5 out of 24 isolates) followed by Proteus bacteria 16.7% (4 out of 24
isolates), Salmonella 12.5% (3 out of 24 isolates) and Klebsiella had the least prevalence rate of
8.3% (2 out of 24 isolates).

Table: Overall prevalence rates of enteric bacteria isolated from isolated bacteria from
vendors A, B and C

Bacteria isolate Prevalence (%)

E. coli 41.7

Salmonella 12.5

S. aureus 20.8

Proteus 16.7

Klebsiella 8.3

56
4.3 Enumeration of coliforms and E. coli.

4.2.1 Enumeration of coliforms and E. coli on day 1

Enumeration of coliforms

The total coliform counts were measured for various food samples obtained from three vendors:
vendor A, vendor B, and vendor C, to assess bacterial contamination. Among the vendors,
Vendor A showed the highest total coliform counts across all food samples. The cooked beef had
a count of 85 (cfu/g), cooked chicken had a count of 80 (cfu/g), cooked sausage had a count of
75, and vegetable salad had a count of 95 (cfu/g) (table 4.4).

In comparison, vendor B had significantly lower total coliform counts for their food samples.
The cooked beef had a count of 10 (cfu/g), cooked chicken had a count of 15 (cfu/g), cooked
sausage had a count of 10 (cfu/g), and vegetable salad had a count of 20 (cfu/g) (table 4.4).

Vendor C fell between vendor A and vendor B in terms of total coliform counts. The cooked
beef and vegetable salad samples had counts of 25 (cfu/g), while the cooked chicken and sausage
samples had counts of 15. These values indicate a moderate level of bacterial contamination in
vendor C's food products (table 4.4).

Table 4.4: Total coliforms from food samples from vendors A, B and C on day 1

Vendor Food sample Total coliform count (cfu/g)

Vendor A Cooked beef 85
Cooked chicken 80
Cooked sausage 75
Vegetable salad 95
Vendor B Cooked beef 10
Cooked chicken 15
Cooked sausage 10
Vegetable salad 20
Vendor C Cooked beef 25
Cooked chicken 15

57
 Cooked sausage 15
Vegetable salad 25

Enumeration E. coli

Table 4.5 provides the E. coli counts in food samples collected from vendors A, B, and C on day
1. For vendor A, the cooked beef sample had 15 (cfu/g) E. coli bacteria, the cooked chicken
sample had none, the cooked sausage sample had 10 (cfu/g), and the vegetable salad sample had
8 (cfu/g). In contrast, vendor B's food samples showed no detectable E. coli counts. The cooked
beef, cooked chicken, cooked sausage, and vegetable salad samples all had counts of 0 (cfu/g),
indicating the absence of E. coli bacteria in these samples. Similarly, vendor C's food samples
also exhibited no detectable E. coli counts. The cooked beef, cooked chicken, cooked sausage,
and vegetable salad samples all had counts of 0 (cfu/g), suggesting the absence of E. coli bacteria
in these samples as well.

Table 4.5: E. coli counts from food samples from vendors A, B and C on day 1

Vendor Food sample E. coli counts (cfu/g)

Vendor A Cooked beef 15
Cooked chicken 0
Cooked sausage 10
Vegetable salad 8
Vendor B Cooked beef 0
Cooked chicken 0
Cooked sausage 0
Vegetable salad 0
Vendor C Cooked beef 0
Cooked chicken 0
Cooked sausage 0
Vegetable salad 0

58
4.2.2 Enumeration of coliforms and E. coli on day 2

Enumeration of coliforms
On day 2, Vendor A had relatively high total coliform counts in the cooked beef and cooked
chicken samples, with counts of 95 (cfu/g) and 90 (cfu/g), respectively. Vendor B had somewhat
lower count across all food samples, with the highest count of 90 observed in the cooked beef
sample. Vendor C had similar levels of total coliforms, with the cooked beef sample having the
highest count of 95 (cfu/g) (table 4.6).

Table 4.6: total coliforms from food samples from vendors A, B and C on day 2

Vendor Food sample Total coliform count (cfu/g)

Vendor A Cooked beef 95
Cooked chicken 90
Cooked sausage 30
Vegetable salad 85
Vendor B Cooked beef 90
Cooked chicken 40
Cooked sausage 85
Vegetable salad 55
Vendor C Cooked beef 95
Cooked chicken 70
Cooked sausage 34
Vegetable salad 45

Enumeration E. coli

Food samples from vendors A, B, and C were tested for E. coli counts on day 2. Vendor A's
samples revealed E. coli counts of 8 (cfu/g) for cooked beef, 5 (cfu/g) for cooked chicken, 0
59
(cfu/g) for cooked sausage, and 9 (cfu/g) for vegetable salad. Vendor B's cooked beef and
cooked chicken samples had no detectable E. coli, but the cooked sausage had a count of 7
(cfu/g). The vegetable salad from Vendor B did not contain any E. coli. Vendor C's cooked beef
and cooked sausage samples did not show any detectable E. coli, while the cooked chicken had a
count of 4 (cfu/g). The vegetable salad from Vendor C was also free from E. coli contamination
(table 4.7).

Table 4.7: E. coli counts from food samples from vendors A, B and C on day 2

Vendor Food sample E. coli counts (cfu/g)

Vendor A Cooked beef 8
Cooked chicken 5
Cooked sausage 0
Vegetable salad 9
Vendor B Cooked beef 0
Cooked chicken 0
Cooked sausage 7
Vegetable salad 0
Vendor C Cooked beef 0
Cooked chicken 4
Cooked sausage 0
Vegetable salad 0

4.2.3 Enumeration of coliforms and E. coli on day 3

Enumeration of coliforms
The total coliform counts from food samples collected on day 3 were examined for vendors A, B,
and C. Vendor A's food samples had varying total coliform counts: cooked beef had 98 (cfu/g),
cooked chicken had 70 (cfu/g), cooked sausage had 65, and vegetable salad had 95 (cfu/g). In
60
comparison, Vendor B's food samples had relatively lower counts: cooked beef had 35 (cfu/g),
cooked chicken had 45 (cfu/g), cooked sausage had 40 (cfu/g), and vegetable salad had 25
(cfu/g). Vendor C's food samples also showed differing counts: cooked beef had 85, cooked
chicken had 30, cooked sausage had 15 (cfu/g), and vegetable salad had 20 (cfu/g) (table 4.8).

Table 4.8: total coliforms from food samples from vendors A, B and C on day 3

Vendor Food sample Total coliform count (cfu/g)

Vendor A Cooked beef 98
Cooked chicken 70
Cooked sausage 65
Vegetable salad 95
Vendor B Cooked beef 35
Cooked chicken 45
Cooked sausage 40
Vegetable salad 25
Vendor C Cooked beef 85
Cooked chicken 30
Cooked sausage 15
Vegetable salad 20

Enumeration E. coli

On day 3, food samples from vendors A, B, and C were analysed for E. coli counts. Vendor A's
food samples showed E. coli counts of 4 (cfu/g) for cooked beef, 0 (cfu/g) for cooked chicken, 0
(cfu/g) for cooked sausage, and 5 for vegetable salad. Vendor B's food samples, including
cooked beef, cooked chicken, cooked sausage, and vegetable salad, did not show any detectable
presence of E. coli. Similarly, Vendor C's food samples, consisting of cooked beef, cooked
chicken, cooked sausage, and vegetable salad, also exhibited no detectable E. coli counts (table
4.9).

Table 4.9: E. coli counts from food samples from vendors A, B and C on day 3
61
Vendor Food sample E. coli counts (cfu/g)
Vendor A Cooked beef 4
Cooked chicken 0
Cooked sausage 0
Vegetable salad 5
Vendor B Cooked beef 0
Cooked chicken 0
Cooked sausage 0
Vegetable salad 0
Vendor C Cooked beef 0
Cooked chicken 0
Cooked sausage 0
Vegetable salad 0

4.3 Antibiotic Susceptibility profiling of bacteria isolated from street-vended foods

All tested bacterial strains demonstrated varying levels of resistance to different antibiotics.
Penicillin resistance was observed in all strains, with E. coli and Klebsiella exhibiting complete
resistance. Proteus showed no resistance to Penicillin, while S. aureus and Salmonella had
resistance rates of 80% (4 out of 5 isolates) and 66.7% (2 out 3 isolates) respectively. For
tetracycline, E. coli and S. aureus had the highest resistance rates at 80%, followed by Proteus at
75% (3 out of 4 isolates). Klebsiella exhibited a resistance rate of 50% (1 out of 2 isolates), while
Salmonella showed no resistance to Tetracycline (table 4.10).

Ampicillin resistance rates were similar to Tetracycline for E. coli and Klebsiella at 80% and
50% respectively. Proteus and S. aureus had no resistance to Ampicillin, while Salmonella
62
showed a resistance rate of 33.3%. Erythromycin resistance was observed in E. coli and S.
aureus, both displaying a resistance rate of 40%. Klebsiella, Proteus, and Salmonella showed no
resistance to Erythromycin. Gentamycin had the lowest resistance rates among the tested strains,
with none of the strains showing resistance except for Salmonella, which had a resistance rate of
33.3% (table 4.10).

Table 4.10: Antibiotic resistance rates of the isolated bacteria (%) from street-vended foods

Antibiotic E. coli Klebsiella Proteus S. aureus Salmonella

Penicillin 100 100 0 80 66.7
Tetracycline 80 50 75 80 0
Ampicillin 80 50 0 40 33.3
Erythromycin 40 0 0 40 0
Gentamycin 0 0 0 0 33.3

REFERENCES

Balter, S. (2006). Foodborne pathogens: microbiology and molecular biology. Emerging

Infectious Diseases, 12(12), 2003-2003. [Link]

Kadariya, J., Smith, T., & Thapaliya, D. (2014). Staphylococcus aureus and staphylococcal food-
borne disease: an ongoing challenge in public health. Biomed Research International, 2014, 1-9.
[Link]

Mwove, J., Imathiu, S., Orina, I., & Karanja, P. (2023). Environmental exposure assessment of
lead and cadmium in street vended foods sold in selected locations in kenya. Food Science &
Nutrition, 11(6), 2610-2619. [Link]

63
Sadiku, M., Ashaolu, T., & Musa, S. (2020). Food contamination: a primer. International Journal
of Advances in Scientific Research and Engineering, 06(03), 01-07.
[Link]

Schatzmayr, G. and Streit, E. (2013). Global occurrence of mycotoxins in the food and feed
chain: facts and figures. World Mycotoxin Journal, 6(3), 213-222.
[Link]

Abdu, A. (2019). Determinants of food safety practices among food handlers in selected food
establishments. International Journal of Public Health Science (Ijphs), 8(2), 229.
[Link]

Asfaw, T. and Genetu, D. (2021). High rate of bacterial contamination on healthcare worker’s
mobile phone and potential role in dissemination of healthcare-associated infection at debre
berhan referral hospital, north shoa zone, ethiopia. Risk Management and Healthcare Policy,
Volume 14, 2601-2608. [Link]

Azanaw, J., Andualem, Z., & Adane, T. (2021). Food safety knowledge, attitude, and practice of
college students, ethiopia, 2019: a cross-sectional study. Biomed Research International, 2021, 1-
10. [Link]

Clayton, D., Griffith, C., & Price, P. (2003). An investigation of the factors underlying
consumers’ implementation of specific food safety practices. British Food Journal, 105(7), 434-
453. [Link]

Lim, S., Lim, E., Kim, J., Paik, H., & Koo, O. (2019). Survival of foodborne pathogens on
stainless steel soiled with different food residues. Food Science and Biotechnology, 29(5), 729-
737. [Link]

Malavi, D., Abong, G., & Muzhingi, T. (2021). Effect of food safety training on behavior change
of food handlers: a case of orange-fleshed sweetpotato purée processing in kenya. Food Control,
119, 107500. [Link]

Banik, A., Abony, M., Datta, S., & Towhid, S. (2019). Microbiological quality of ready - to - eat
food from dhaka, bangladesh. Current Research in Nutrition and Food Science Journal, 7(1),
161-168. [Link]

Eromo, T., Tassew, H., Daka, D., & Kibru, G. (2016). Bacteriological quality of street foods and
antimicrobial resistance of isolates in hawassa, ethiopia. Ethiopian Journal of Health Sciences,
26(6), 533. [Link]

Muyanja, C., Nayiga, L., Brenda, N., & Nasinyama, G. (2011). Practices, knowledge and risk
factors of street food vendors in uganda. Food Control, 22(10), 1551-1558.
[Link]
64
Rane, S. (2011). Street vended food in developing world: hazard analyses. Indian Journal of
Microbiology, 51(1), 100-106. [Link]

Were, L., Were, G., & Aduol, K. (2020). Hygiene practices and microbial contamination of
street-vended foods in kenyatta university’s environs. European Journal of Agriculture and Food
Sciences, 2(5). [Link]

Boels, D. (2014). It's better than stealing: informal street selling in brussels. International Journal
of Sociology and Social Policy, 34(9/10), 670-693. [Link]

Madziba, E. (2017). Street vending in zimbabwe: an urban scourge or viable enterprise. Journal
of Corporate Governance Insurance and Risk Management, 4(1), 55-71.
[Link]

Nani, G. (2020). Trading space rivalry between street vendors and shop-based traders in
bulawayo metropolitan province, zimbabwe: a conceptual perspective for co-existence.
Academic Journal of Interdisciplinary Studies, 9(4), 168. [Link]
0071

Adhikari, D. (2012). Income generation in informal sector: a case study of the street vendors of
kathmandu metropolitan city. Economic Journal of Development Issues, 1-14.
[Link]

Colavecchio, A., Cadieux, B., Lo, A., & Goodridge, L. (2017). Bacteriophages contribute to the
spread of antibiotic resistance genes among foodborne pathogens of the enterobacteriaceae
family – a review. Frontiers in Microbiology, 8. [Link]

Dark, M. (2013). Whole-genome sequencing in bacteriology: state of the art. Infection and Drug
Resistance, 115. [Link]

Liao, X., Deng, R., Warriner, K., & Ding, T. (2023). Antibiotic resistance mechanism and
diagnosis of common foodborne pathogens based on genotypic and phenotypic biomarkers.
Comprehensive Reviews in Food Science and Food Safety, 22(4), 3212-3253.
[Link]

Tao, Q., Wu, Q., Zhang, Z., Liu, J., Tian, C., Huang, Z., & Yong, Z. (2022). Meta-analysis for
the global prevalence of foodborne pathogens exhibiting antibiotic resistance and biofilm
formation. Frontiers in Microbiology, 13. [Link]

Zhang, L., Gao, F., Ge, J., Li, H., Xia, F., Bai, H., & Shi, L. (2022). Potential of aromatic plant-
derived essential oils for the control of foodborne bacteria and antibiotic resistance in animal
production: a review. Antibiotics, 11(11), 1673. [Link]

65
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