MULTI-DRUG RESISTANT STAPHYLOCOCCUS AUREUS, ESCHERICHIA COLI AND

PROTEUS SPP FROM MEAT AND BUTCHER EQUIPMENT AROUND DUTSIN-MA

TOWN, KATSINA STATE

A PROJECT WORK

BY

YAKUBU EZRA

LSC/2020/15198

SUBMITTED TO THE DEPARTMENT OF MICROBIOLOGY, FEDERAL

UNIVERSITY DUTSIN-MA, IN PARTIAL FULFILMENT OF THE
REQUIREMENTS FOR THE AWARD OF THE DEGREE OF BACHELOR
OF SCIENCE (MICROBIOLOGY)

JULY, 2024
 DECLARATION

I declare that the work in this project titled: MULTI-DRUG RESISTANT

STAPHYLOCOCCUS AUREUS, E.COLI AND PROTEUS SPP FROM MEAT AND

BUTCHER EQUIPMENT AROUND DUTSIN-MA TOWN, KATSINA STATE, has been

carried out by YAKUBU EZRA from department of Microbiogy, faculty of life science, Federal

University dutsin-ma, Katsina state. The information derived from the literature is duly

acknowledge in further research and a list of references was provided. No part of this project was

previously presented for another degree or diploma at th or any other institution.

_____________________ __________________

Student signature Date

 CERTIFICATION

This project titled: MULTI-DRUG RESISTANT STAPHYLOCOCCUS AUREUS, E.COLI

AND PROTEUS SPP FROM MEAT AND BUTCHER EQUIPMENT AROUND DUTSIN-

MA TOWN, KATSINA STATE was carried out by YAKUBU EZRA meet the requirement

governing the award of bachelor degree of science in microbiology

Mal. Khalifa Jamil Saleh __________________________

Project supervisor Sign/Date

Mr. Wata Innocent John

-----------------------------------------

Project coordinator Sign/Date

Head of department. ________________________

Mal. Umaru AbdullMalik Sign/Dtae

________________________ ________________________

External Examiner Sign/Dtae

 ACKNOWLEDGEMENT

I wish to register my sincere and heartfelt gratitude to my supervisor Mal. KHALIFA JAMIL

SALEH, for his inspiring guidance, patience, encouragement, keen interest, scholarly comments,

and constructive suggestions throughout the course of the study. Special thanks to Mr. Alabi

Emmanuel Dayo from the Department of Microbiology for facilitating the laboratory work. I

also wish to cherish my parents Dr.Yakubu Jambo Makama and Mrs. Dinatu Yakubu Jambo, for

their support, prayers and mentorship. I also acknowledged the tremendous efforts of my

beloved brother Lucky Yakubu jambo, Florence Yakubu, Enoch Yakubu, Namadi Yakubu and

Jethro Yakubu for their prayer, support and encouragement in the course of my study.

I am also indebted to the Head of Department, the project coordinator, Mal. Umaru

AbdullMalik, Mr. Innocent Wata John, for their tireless assistance during the course of my study.

I acknowledge the contributions of my friends Nasara Bulus, Mary musa, Melody Kyument,

Godiya Haruna Lawal, Dorcas Bako, Kosarachi Nwankwo and so many that cannot be

mentioned. I’m grateful to my fellow students for their constructive criticism and

encouragement.
 DEDICATION

I dedicate this project to GOD THE FATHER, GOD THE SON AND GOD THE HOLYSPIRIT.

He has been the source of my strength throughout this program. I also dedicate this to my parent

Dr. Yakubu Jambo Makama and Mrs.Dinatu Yakubu Makama whose encouragement have made

sure that I give it all it takes to finish that which I have started. May Elohim be praise. "Amen".
 Table of Contents
DECLARATION.............................................................................................................................ii
CERTIFICATION..........................................................................................................................iii
ACKNOWLEDGEMENT..............................................................................................................iv
DEDICATION.................................................................................................................................v
ABSTRACT.................................................................................................................................viii
CHAPTER ONE................................................................................................................................
1.0 Introduction..................................................................................................................................
1.1 Background of The Study............................................................................................................
1.2 Statement of Problem..................................................................................................................
1.3 Justification of the study..............................................................................................................
1.4 Aim and Objectives of The Study...............................................................................................
CHAPTER TWO...............................................................................................................................
2.0 Literature Review........................................................................................................................
2.1 Meat.............................................................................................................................................
2.2 Meat micribiology.......................................................................................................................
2.3 Meat industry and its significance...............................................................................................
2.4 Mear seller pailm.........................................................................................................................
2.5 Meat seller knive..........................................................................................................................
2.6 Bucher table.................................................................................................................................
2.7 Escgerichia Coli..................................................................................................................
2.8 Staphylococcus aureus.................................................................................................................
2.9 Proteus spp...................................................................................................................................
2.9 .1 Antimicrobial Resistance-A Worldwide Threats.....................................................................
CHAPTER THREE...........................................................................................................................
MATERIALS AND METHODS......................................................................................................
3.1 Study Area...................................................................................................................................
3.2 Sampling size and Sample techniques.........................................................................................
3.3 Sample Collection........................................................................................................................
3.4 : Determination of Bacteria in samples, isolation and storage of bacteria...................................
3.5 Bacterial identification and characterization..........................................................................
3.6 Gram-Stainning.........................................................................................................................
3.7 Determination of antibiotic resistance profile of isolates.......................................................
CHAPTER FOUR ............................................................................................................................
4.O RESULTS………………………………………………………………………………………………

4.1 Table 1: Occurrences of Escherichia coli, Staphylococcus aureus and Proteus spp from
meat and butcher equipment from various locations………………………………………...

4.2 Antibiotic resistance pattern of Escherichia coli, Proteus spp and Staphylococcus
aureus isolate....................................................................................................................................
4.3 : Percentage of total selected multidrug resistant bacteria from meat and butcher
equipments from various locations…………………………………………………………….

4.4 Phenotypes of multidrug resistant bacteria isolates……………………………………….

CHAPTER FIVE………………………………………………………………………………………………………………………………………….

5.0 Discussion, Conclusion and Recommendation………………………………………………

5.1 Discussion……………………………………………………………………………………..

5.2 Conclusion…………………………………………………………………………………….

5.2CONCLUSION………………………………………………………………………….

5.3 RECOMMENDATIONS.............................................................................................................
REFERENCE....................................................................................................................................
APENDIX………………………………………………………………………………………………………………………………………………
 ABSTRACT
This study was conducted to determine the multidrug resistant E.coli, Proteus spp and S.aureus
from meat and butcher equipment around Dutsin-ma metropolis, Katsina state. A total of 40
samples were collected from 4 locations, while a total of 10 samples was collected from each
location, 2 swab samples from table, 2 knife sharpener, 2 swab samples from the palm, 2 swab
samples from the knife and 2 from the meat. A stratified random sampling Collecting were
employ to ensure representation from various socio-economic group within the local
government area. A total of 77 isolates were obtained and Antimicrobial susceptibility testing
was performed. Amongst this 77 isolated, Staphylococcus aureus was the frequently (37/77,
92.5%) isolated bacteria, Proteus spp (29/77, 72.5%) followed and Escherichia coli (28/77,
70%). The highest percentage of multidrug resistant bacteria isolated from this study was
observed to be Staphylococcus aureus (50%), Escherichia coli was observed to follow (41.3%)
and Proteus spp was observed to be the least (8.7%) amongst the total selected multidrug
resistant bacteria. The phenotypes of multidrug resistant bacteria isolates from meat and
butcher equipment around Dutsin-Ma town. A total of (46/77, 100%) MRD bacteria were
isolated from meat and butcher equipment collected from this study. Among these MDR
bacteria, Staphylococcus aureus was observed to have the most common (23/46, 50%) resistant
phenotype, which were resistant to ampicillin, gentamycin, erythromycin, ampiclox and Septrin
while Escherichia coli was observed to followed (19/46, 41.3%) and Proteus spp was observed
to have the least (4/46, 8.7%) resistant phenotype. This poses a serious public health concern to
the consumers of these meats and is an indicator of the level of hygienic practices at the various
meat stalls in Dutsin-Ma metropolis. The microbiological safety of food is achieved by as far as
possibly ensuring the absence of pathogenic microorganisms and by all means preventing their
multiplication.
 CHAPTER ONE

INTRODUCTION

1.0 Backgroung of the study

Animals are a great source of protein. When eaten as food, they can make up over 60% of the
dietary protein intake by adults, especially in rural areas (Adeleye OA, 2008).In Nigeria, "meat
form a much-cherished delicacy that cuts across socio-economic, age, religious and educational
barriers (Adebayo-Tayo BC, Onilude AA, 2008). Meat is a most cherished food delicacies in
Nigeria, but can be a source of dissemination of Multi-drug Resistant (MDR) bacteria (2013 ).
The presence of antibiotic-resistant bacteria in food-related environments is a growing concern
TOPIC: Multi-Drug Resi tant Staphylococ us aureus, E.coli and Proteus sp From Meat
worldwide. These bacteria, such as Escherichia coli, Proteus spp and Staphylococcus aureus,
pose a significant threat to public health due to their potential transmission through contaminated
food products (Martinez et al., 2009). Butchers equipment like knive and utensils, where raw
meat is processed bacteria led, can serve as potential reservoirs for antibiotic-resistant bacteria,
the meat and butcher equipment increasing the risk of foodborne infections. Understanding the
antibiotics profiles and multi-drug resistance patterns of these bacteria on butcher equipment is
crucial for effective public health interventions and the development of appropriate antimicrobial
stewardship strategies. Antibiotic resistance is a global public health crisis, diminishing the
efficacy of antibiotics and rendering previously treatable infections difficult to manage. Bacteria
such as Escherichia coli, Proteus spp and Staphylococcus aureus have demonstrated remarkable
adaptability, acquiring resistance genes through horizontal gene transfer, mutations, and selective
pressure from antimicrobial usage. Contaminated meat may be responsible for several meat-
borne diseases in humans (Umoh, 2002). Meat contamination may originate from animal
sources, food contact surfaces such as tables, utensils and other equipment (Bello and Son, 2009)
and environmental conditions where animals are processed (Olanike, 2002). In beef carcass
processing, bacterial organisms such as Escherichia coli increases or decreases during processing
depending on level of fecal contamination of live cattle, efficiency of evisceration (Rigobelo et
al., 2006), and hygienic practices in the abattoir (Rigobelo et al., 2006). Other organisms
commonly encountered in meat contamination are Salmonella spp., Escherichia coli O157:H7
and Listeria spp. (Mc Envoy et al., 2003; Declan et al., 2004). Public health concerns in meat
safety with regards to consumer health have resulted in recall of contaminated products from
market-places associated with microbial and especially bacterial pathogens (Sofos, 2003). To
ensure safe and wholesome meat to consumer at all the stages of processing, good hygiene
practice and standard operating procedures must be instituted up to the methods of packaging,
distribution and marketing of the meat and meat products. Meat must be free from bacterial cross
contamination and extrinsic contaminants such as toxin, chemical residues that will be injurious
to human (Olugasa et al., 2000).

Raw beef sold at retail outlets in Nigeria undergo a considerable amount of handling and contact
with microbes of different sources (Umoh et al., 2002). Keeping meat at ambient temperature for
a longtime which is commonly done by meat retailers in the market may allow multiplication of
pathogenic bacteria (Umoh et al., 2002). Detection of total coliform and Escherichia coli as bio-
marker organisms in food is widely applied in many food control laboratories. Meat can be
assessed with respect to microbiological safety by estimation of the growth of Escherichia coli.
For instance, research by Johnson et al. (2019) examined the prevalence of antibiotics resistant
Escherichia coli in retail meat samples, providing valuable insights into the potential sources and
transmission routes of antibiotic resistance. Similarly, Smith et al. (2020) conducted a study on
Staphylococcus aureus isolated from raw meat products, highlighting the presence of multi-drug
resistant strains and emphasizing the need for improved hygiene practices. And Dammy. (2023)
conducted a research on determination multi-drug resistant Escherichia coli and Staphylococcus
aureus associated with hands and equipment of meat seller. The research showed that multidrug
resistant bacteria can be isolated from meat seller benches, palm and knife collected. This calls
for the urgent implementation of integrated surveillance system within the entire food chain and
a critical reexamination of food sanitary quality to improve the keeping quality of meat.

1.1 Research Problem

The presence of Multi drug resistant bacteria associated with meat and butcher equipment pose a
significant public health concern like salmonellosis, cholera, dysentery and so on. This study will
help in identifying environmental sources; Environmental reservoirs for pathogenic bacteria can
potentially contaminate meat and butcher equipment include various sources; Soil and Water
bacteria such as Salmonella, Escherichia coli, Proteus spp, and Listeria can persist in soil and
water, especially in agricultural areas where livestock are raised, residual bacteria from animal
carcasses can contaminate equipment during slaughter and processing. Improperly cleaned
equipment and surfaces in butcher shops can harbor pathogens like Campylobacter, Proteus, E.
Coli and Staphylococcus aureus. Some bacteria can be airborne and settle on surfaces,
potentially contaminating meat and equipment. Transfer of bacteria from one surface or food to
another can occur through improper handling practices. Effective sanitation practices, including
regular cleaning and disinfection of equipment and surfaces, are crucial to mitigate these risks
and ensure food safety in meat processing environments and reservoir of multidrug resistant of
E.coli, S.aureus and proteus spp which is critical for understanding the spread of antibiotic
resistant infection and propose effective strategies for prevention and control the risk of
foodborne infection
1.2 Justification of study

The microbial analysis of bacteria isolates from meat and butcher equipment is a topic of
significant relevance to the society. The safety of meat as a perishable food product is a major
concern, considering that it can harbor various microorganisms that may pose health risks to
consumers.

The significance of this work is to enable those producers to improve the hygiene of the food (meat and
butcher equipment} and a good knowledge of safe food handling practice, suitable protective clothing to
be worn end it will also enable us to be aware of various pathogenic bacteria infections transmitted
through meat processing utensils like knives, table, knife sharpener, etc.

1.3 Aim and Objectives

1.3.1 Aim
The aim of this research is to determine the multidrug resistant Bacteria from meat and butcher
equipment around Dutsin-ma metropolis, Katsina state

1.3.2 Objectives:

1: To Isolate and identify Bacteria pathogens from meat and butcher equipment sold at different meat
selling points in Dutsin-ma metropolis.

2: To determine the antimicrobial susceptibility profile of the isolates.

3: To determine multi-drug resistance bacteria among the isolates

 CHAPTER TWO

2.0 LITERATURE REVIEW

Animals are a great source of protein. When eaten as food, they can make up over 60% of the
dietary protein intake by adults, especially in rural areas (Adeleye OA, 2008).In Nigeria, "meat
form a much-cherished delicacy that cuts across socio-economic, age, religious and educational
barriers (Adebayo-Tayo BC, Onilude AA, 2008). Meat cherished a food delicacies in Nigeria,
but can be a source of dissemination of Multi-drug Resistant (MDR) bacteria (2013).The
presence of antibiotic-resistant bacteria in food-related environments is a growing concern
worldwide. These bacteria, such as Escherichia coli, Staphylococcus aureus and proteus spp
pose a significant threat to public health due to their potential transmission through contaminated
food products. Food safety is an important concern that has increased in international trade.
Outbreaks of food-borne pathogens are one of the important things that leads to illness and death,
about 24-81 million of foodborne disease associated with meat every year were recorded
(Ahmed et al., 2013). The eating of contamination foods with pathogenic bacteria and their
products such as toxins and enzymes leading to serious diseases (Gdoura et al., 2018). Food
spoilage mean any change in food such as taste, smell, and appearance, but it remains safe for the
consumer until the number of microorganisms present in the food reaches a certain limit that
causes food-borne diseases (Anwer et al.,2017). Meat and meat products are considered a fertile
environment for microorganisms and their toxins, including bacteria (Yousuf et al., 2008). One
of the most important bacteria isolated from meat is Escherichia coli, Staphylococcus aureus,
Streptococcus spp., Shigella spp., Salmonella spp., and Clostridium perfringens (Karoki et al.,
2018). Food-borne pathogens are colonizing the gastro intestinal tracts of consumed domestic
animals by human (Onilude et al., 2016). The slaughtered animals are sterile but despite this,
meat gets easily contaminated with microorganisms by several processes (Abd et al., 2015).
Most of these organisms transmitted into meat and meat products through the hands and clothes
of workers and contaminated environment, devices, and knives (Mathew et al., 2016).
Foodborne microorganisms are a major source of disease and death, leading to significant
spending on healthcare. Children are more susceptible to food poisoning because of their weak
immune system (Nagarajan et al., 2018). There are two types of microbial contaminants,
microorganisms that capable to produce disease and the second type that spoils the meat products
and makes it unfit for human consumption. The increase consumption of meat especially the
contaminated foodborne bacteria because including proteins, vitamins, lipids, minerals and other
nutrients resulted outbreak of food-borne infections (Sayyadifar et al., 2012), possible Several
studies have investigated antibiotic resistance patterns in various food-related settings, including
retail meat environments. For instance, research by Johnson et al. (2019) examined the
prevalence of antibiotic-resistant Escherichia coli in retail meat samples, providing valuable
insights into the potential sources and transmission routes of antibiotic resistance. Similarly,
Smith et al. (2020) conducted a study on Staphylococcus aureus isolated from raw meat
products, highlighting the presence of multi-drug resistant strains and emphasizing the need for
improved hygiene practices. Understanding the antibiotic profile and multi-drug resistance of
Escherichia coli and Staphylococcus on meat seller's benches is crucial for implementing
effective control measures. By identifying the prevalent resistance patterns and risk factors
associated with contamination, interventions can be tailored to minimize the dissemination of
antibiotic-resistant bacteria. Furthermore, this knowledge can inform the development of
antimicrobial stewardship programs aimed at promoting responsible antibiotic use and reducing
the emergence of resistance.

The main reservoir of S. aureus is the hand from where it is introduced into food during handling
and preparation (Hui et al., 2001). Hand serves as a major vehicle of transmission of various
microbes including the Enterobacteriaceae group (Kapdi et al., 2008). They stressed that the
microbial population of the hand is extremely complex and variable, consisting of gram positive
organisms like S. aureus and gram-negative organism like Pseudomonas aeruginosa, which may
survive for sufficient period of time on the hand and may thus serve either as a reservoir or
shelter of infection.

2.2 Meat

Originally meat was a term used to describe any solid food, but has now come to be applied
almost to animal flesh. As such it has played a significant role in the human diet since the days of
hunting and gathering, and animals (sheep) were first domesticated at the beginning of the
Neolithic revolution around 9000 BC. Though abjured by some moral or religious grounds, meat
eating remains widely popular today. (Adams and Moss, 2000). Meat consumption is often
something of a status symbol and is generally for greater and wealthy societies. This is because
large- scale meat production is a relatively inefficient means of obtaining proteins. It requires
agriculture to produce a surplus of plant protein which can be fed to animals. With modern
production techniques it takes two kilograms of grain to obtain one kilogram of chicken, four for
one kilogram of pork and eight for one kilogram of beef. Red meat is derived from a number of
animal species (e.g. cattle, sheep, goats, camels and pigs). Total world production of red meat
was estimated to be approximately 120 million tones carcass weight in 1991. Red meats are
important in international trade with about 11 million tons per year being exported. Red meat
has the potential to carry food – poisoning organisms to consumers. The food – poisoning
organisms which can constitute a hazard in at least some meat products are Salmonella spp,
Entero-haemorrhagic Escherichia coli, (e.g. Serotype 0157), some serovars of Yersinia
enterocolitica, Campylobacter jejuni, Staphylococcus aureus, Listeria monocytogenes,
Clostridium perfringens, Clostridium botulinum and Bacillus cereus. Meats are also subject to
microbial spoilage by a range of microorganisms including Pseudomonas spp, Shewanella,
Enterobacteriaceae, Brochothrix thermosphacta, lactic acid bacteria, Yeasts and Molds
(Forsythe 2010)

2.3 Meat Microbiology

The internal tissues of a healthy animal are known to be sterile and free of microorganisms (Jay,
2005). Contamination at and after slaughter seems to be the only source of microorganisms,
their primary sources and roots are; The stick knives, Animal hides or skins, gastrointestinal
tract, hands of handlers, containers and utensils, storage environment, and Lymph nodes of the
animal itself.

2.4 Meat Industry and its Significance

The meat industry is a powerhouse in the global economy. It contributes significantly to

agricultural GDP and plays a vital role in many national economies. In the United States, for
instance, the livestock sector, including meat production, is a substantial part of the agricultural
economy (ERS, 2020). Similarly, in Australia, the meat and livestock sector has a prominent
position in the country's agricultural exports, making it a cornerstone of the Australian economy
(ABARES, 2019). Meat is a primary source of high-quality protein and is rich in essential amino
acids, vitamins, and minerals. This nutritional richness makes meat an essential component of
human diets, providing critical nutrients necessary for growth, development, and overall health.
Meat is particularly important for addressing protein-energy malnutrition in developing countries
(Drewnowski and Almiron-Roig, 2010). The meat industry sustains the livelihoods of millions of
people globally. It provides employment for farmers, ranchers, butchers, and workers in meat
processing plants. Furthermore, it promotes sustainable land use practices, as efficient animal
husbandry and pasture management are essential components of meat production (Thornton,
2010). The meat industry plays a vital role in ensuring food security by providing a reliable
source of protein and essential nutrients. It contributes to meeting the dietary requirements of
populations, particularly in regions with limited dietary diversity. For many, meat is a primary
source of protein, especially in low and middle-income countries (Delgado, 2003). The meat
industry is a significant player in international trade, with the export of meat products
contributing to economic growth and fostering international relations. Countries often engage in
the export and import of meat to meet domestic demands and address market fluctuations (Haley
& Seale, 2013). Meat holds a central place in culinary traditions and cultural practices around the
world. It is frequently associated with special occasions and traditional celebrations, making it a
symbol of cultural heritage and identity (Sobal, 2005).

2.5 Meat seller palm

Meat sellers serve as a crucial link in the food supply chain, responsible for providing consumers
with safe and wholesome meat products. The hygiene of their hands, particularly their palms, is a
critical factor in ensuring food safety and public health.

Studies have consistently shown that the palms of meat sellers can harbor a wide range of
microorganisms, including pathogenic bacteria like Escherichia coli and Staphylococcus aureus
(Ahmed et al., 2017). Contamination often results from direct contact with raw meat and cross-
contamination during food handling. Emerging research indicates the presence of antibiotic-
resistant strains of bacteria on meat seller palms, raising concerns about the potential for
antibiotic resistance transmission to consumers (Gonzalez et al., 2019).
Several factors contribute to the level of microbial contamination on meat seller palms. These
include;

1. The meat's source and quality

2. Personal hygiene practices
3. The cleanliness of the selling environment
Contaminated palms represent a significant public health risk, as pathogens can be transferred to
the meat products and, subsequently, to consumers. Proper food handling, thorough cooking, and
awareness of potential risks are crucial for mitigating these health hazards (Lues et al., 2006;
Powell et al., 2013). To address these concerns, regulatory agencies often implement guidelines
and standards for meat handling and hygiene practices among sellers. Regular inspections,
educational programs, and enforcement of these standards are essential (Codex Alimentarius,
2010; FDA, 2020). The hygiene of meat seller palms is pivotal in maintaining food safety. While
many meat sellers adhere to good hygiene practices, ongoing efforts are necessary to ensure
compliance, raise awareness, and reduce microbial contamination risks. Consumers also play a
crucial role in food safety by practicing proper food handling and cooking techniques.

2.6 Meat seller knife

The knife is a fundamental tool in the meat-selling, essential for various tasks such as cutting,
trimming, and portioning meat products. Meat sellers commonly utilize a range of knives,
including boning knives, butcher knives, and cleavers. Each type serves a specific purpose, such
as deboning, slicing, or chopping (Hui et al., 2016). Proper knife maintenance and sharpening
are essential to ensure efficient and safe meat cutting. Regular sharpening not only improves
precision but also reduces the chances of accident due to dull blades (Smith et al., 2018). The
knives used by meat sellers can become potential sources of microbial contamination. Inadequate
cleaning and sanitation practices may lead to the buildup of bacteria, including Salmonella and
Listeria, on knife surfaces (Sagoo et al., 2013; Jay, 2012). Cross-contamination can occur when
knives come into contact with raw meat and then contact ready-to-eat products or surfaces,
increasing the risk of foodborne illnesses (Sivertsvik et al., 2003; Rhee et al., 2003). To mitigate
contamination risks, meat sellers should follow strict hygiene protocols, including regular
cleaning and sanitization of knives and surfaces, as well as implementing color-coded knives to
prevent cross-contamination (Feng et al., 2008; Food Standards Agency, 2019). Knives are
indispensable tools for meat sellers, but their proper use and maintenance are vital to prevent
microbial contamination and foodborne illness risks. Following hygiene protocols and regulatory
standards can help minimize these risks and contribute to safer meat handling practices.

2.7 Butcher table

The butcher table, also known as a meat preparation table, is a surface specifically designed for Cutting,
trimming, and preparing meat products in butcher shops or meat processing facilities. These tables are
typically made of stainless steel or other easy-to-clean materials to maintain Hygiene standards. It's
important to keep butcher tables clean and sanitized to prevent cross-Contamination and ensure food
safety. Regular cleaning, disinfection, and proper handling practices are essential to maintain a sanitary
environment and minimize the risk of microbial contamination. Various microorganisms can be found,
including bacteria such as Salmonella, E. coli, and Listeria. These organisms can come from raw meat,
surfaces, and handling practices. Proper cleaning and sanitation protocols are crucial to minimize the risk
of contamination. Regular cleaning of butcher tables with disinfectants and ensuring proper food safety
practices essential to maintain a hygienic environment.

2.8 Escherichia coli

Scientists all over the world have studied Escherichia coli and it appears to be the most
thoroughly investigated and best understood of all model microorganisms (Hill et al., 2019,
Lovley et al., 2020, Ranganathan et al., and 2020 Macklin et al., 2020). We already know that it
is one of the first bacteria that colonies the human gut immediately after birth (Micenkova et al.,
2020, Bittinger et al., 2020, and Secher et al., 2016). On the other hand, E. coli is often the main
culprit of infections in the gastrointestinal tract (Rossi et al., 2018), as well as other parts of
human and animal organisms (Zhang et al., 2020 and Abebe et al., 2020). In more precise terms,
E. coli typically causes urinary infections (Isla et al., 2017 and Rodrigues et al., 2016), but it can
also lead to many other serious infections and conditions, such as: appendicitis (Song et al.,
2018), pneumonia (Park et al., 2019), meningitis (Zhao et al., 2018), endocarditis (Akuzawa et
al., 2018), gastrointestinal infections (Sarowska et al., 2019), etc. Research findings have shown
us that E. coli can cause infections in all age groups and those infections can be acquired in the
general population, i.e., community-acquired, as well as related to healthcare institutions
(Poolman et al., 2018, Kubone et al., 2020 and Djordjevic et al., 2016). After Alexander Fleming
had discovered penicillin in 1928, the whole course of medicine changed (Gaynes et al., 2017
and Hutchings et al., 2019). The revolutionary discovery of antibiotics made it possible for
doctors to treat extremely severe cases of infectious diseases, which had previously been a very
common cause of death (Dodds et al., 2017 and Aminov et al., 2017). That completely changed
after antibiotics had been introduced and soon penicillin became the most widely used antibiotic
in the world, saving millions of lives (de Opitz et al., 2020, Gajdacs et al., 2019 and Coates et
al., 2020). E. coli is an important zoonotic pathogen. Escherichia coli O157:H7 was first
recognized in 1982 as a human pathogen and cattle have been identified as a major source of
Escherichia coli O157:H7 infection of human but it is not pathogenic in cattle and present in the
feces of healthy cattle (Elder et al., 2000).

Escherichia coli are part of the mammalian and human intestinal microbiota. However, they are
also widely distributed in the environment. Among the Escherichia coli strains, not just
commensal, but also pathogenic, intestinal pathogenic Escherichia coli strains (IPEC) and extra
intestinal pathogenic E. coli (ExPEC), can be found. It is well documented that contamination of
meat with pathogens constitutes a major public health concern (Cohen et al., 2007)

Antimicrobial resistance take place when bacteria adjust or adapt in a way that permits them to
stay alive in the presence of antibiotics designed to kill them., bacteria evolve resistance to these
drugs, typically by acquiring chromosomal mutations and multidrug resistant plasmid &
transposon etc. (Finch et al., 2003; Levine et al., 2002; Nichol et al., 2003; Sheng, 2002). Meat
has long been regarded as a valuable source of protein, and many people's appetites for it are
growing every year (R. Ekli et al., 2020)

Most meats have a high-water content, with a water activity of around 0.99, which is ideal for
microbial growth (Rao et al., 2009). Food spoilage and foodborne infections in humans are both
caused by microbial growth, resulting in nancial and health losses (Komba et al., 2012)]. Some
strains of Escherichia coli have been linked to foodborne infections in humans.

The emergence of multi-drug resistance in bacterial human pathogens is one of the most serious
challenges for healthcare globally. Pathogens that earlier were sensitive to antibiotics are
becoming resistant by mutations in their pre-existing DNA or by acquisition of DNA containing
resistance genes (Martinez et al., 2009).
Recent studies have shown that antibiotic resistance genes are ancient and were present in the
environment long before the antibiotic era (D’Costa et al., 2011; Bhullar et al., 2012). The abuse
of antibiotics has led to increased selection pressures even in the environment. Resistance genes
may radically increase in abundance within the populations in bacterial communities exposed to
sufficient selection pressure from exposure to antibiotics, (Barbosa and Levy, 2000; Dethlefsen
et al., 2008; Kristiansson et al., 2011). Such increases may also be accompanied by increased
frequencies in genetic elements facilitating their mobility (Jernberg et al., 2010; Kristiansson et
al., 2011). Thus, exposure to antibiotics is expected to increase the risk for the transfer of
resistance between bacterial species.

2.9 Staphylococcus

The family of Staphylococcaceae encompasses 51 species and 27 sub-species of ubiquitous

bacteria occurring in grapelike clusters of Gram-positive cocci. They are grouped into coagulase-
positive (CoPS) and negative (CoNS) Staphylococci due to their ability to produce the coagulase
enzyme (Kluytmans, 2010; Osman et al., 2016). Staphylococcus species appear worldwide as
commensal colonizers of skin of animals and humans. Additionally, they are found on mucous
membrane of the upper respiratory and lower urogenital tracts, and transiently in the digestive
tract (Kluytmans, 2010).

S. aureus is the most important human pathogenic specie of this bacterial family and constitutes
together to be the main pathogenic coagulase-positive (CoPS) in animals. Globally, S. aureus
asymptomatically colonizes mucous membranes of the respiratory and intestinal tracts as well as
other body surfaces, but it is also frequently involved in a broad range of diseases ranging from
mild skin infections to life-threatening invasive infections in humans and animals. S. aureus
became a worldwide health problem due to the emergence of methicillin resistant S. aureus
(MRSA) (Njoungang et al., 2015). MRSA was first reported from animals in 1972 following its
isolation in milk from mastitic cows (Kluytmans, 2010). Subsequently, it was isolated from
various animal species including pets, horses, pigs, poultry, sheep, veal calves and dairy cows.
Although Staphylococcus aureus is the most important pathogen among all Staphylococcal
species, CoNS have recently gained importance with increasing concerns in human and animal
health due to the emergence and implications of resistant strains in several human and animal
infections (Bhargava and Zhang, 2012; Njoungang et al., 2015). More significantly, methicillin-
resistant CoNS (MRCoNS) have been detected worldwide from food animals including pigs,
poultry, calves and cows as well as food and food products (Huber et al., 2011). There is a dearth
of information about MRS in food animals and food products in Africa.

Staphylococcal food poisoning is one of the most economically important foodborne diseases
and produces gastrointestinal illness through a wide variety of toxins, including Staphylococcal
enterotoxins characterized by vomiting and diarrhea within 2 to 6 h after the consumption of
contaminated food in the United States (Cenci et al., 2003, Kim et al., 2018, Hachemi et al.,
2019). Staphylococcus aureus and other pathogens in meat, as a result of improper hygienic
practices at the point of handling by

2.9.1 Proteus spp

Proteus spp. are Gram-negative bacteria belonging the Enterobacteriaceae family and are
common commensals of the gastrointestinal microbiota. The family Enterobacteriaecae includes
the genus Proteus. P. mirabilis, P. vulgaris, P. penneri, and P. myxofaciens are the four known
species of Proteus. P. vulgaris is the most commonly isolated species from clinical illnesses and
opportunistic infections (Zappa et al., 2017). Proteus species are different from most of other
genera by their ability to swarm through a blood agar surface (Mohammed et al., 2016). These
bacteria are Gram negative bacillus, non-capsulated , non-spore forming, Lactose non-
fermenting , catalase positive ,oxidase negative , facultative anaerobic, hydrolyze urea quickly,
chemo-organotrophic having together a respiratory and a fermentative form of metabolism
(Ahmed, 2015).The Enterobacteriaceae bacterium family is the most difficult to eradicate from
meat and meat products around the world. In all cases of food poisoning linked to meat products,
E. coli, Salmonella, Proteus, and Klebsiella are the most common bacteria (Drzewiecka, 2016
and Nahar et al., 2014). Many food poisoning outbreaks have been linked to Proteus group
organisms, and with the increased incidence of Proteus spp. Related food borne diseases, there is
an urgent need for control and/or prophylactic for food poisoning outbreaks linked to meat
products. To ensure food safety and protect public health from microbial contamination of food,
it is critical to investigate such agents in foods and eliminate them (Cooper et al., 2005).The
Proteus bacteria found as normal flora in the intestinal tract being as saprophytes, whereas a
number of them may present as parasitic status and might be an opportunistic pathogens, which
producing numerous types of infections when they leave their natural inhabitation (Drzewiecka,
2016 ). Proteus species, like several other members of the Enterobacteriaceae family, are short
(1.5- to 2-μm) straight rods that exhibit dimorphism as "swimming" and "swarming" forms
(Armbruster and Mobley 2012).

The best temperature for the growing of Proteus spp. is 37 °C with a pH 7.4 many culture
media have been developed for diagnosis of Enterobacteriaceae, which in turn can discriminate
between genera in this family, including Proteus spp. (Chen et al., 2015).The bacteria seemed as
distinct pale colonies on MacConky agar (Mohammed et al ., 2016 ) . Non-ferment lactose,
moderate in size and entire edges colony as well as the odor of bacterial growth which is similar
to odor of fish decaying, which is the formula primary diagnostic for this bacteria and performed
swarming movement on the blood agar Swarming behavior makes it hard to isolate as the
distinct colony , this phenomena can restrained by increased concentration of agar (2.5-3.0%) or
P- nitrophenylglycerol considerably restrained swarming but its influence on growth amount was
not important (Kadhim et al., 2019).

2.9.2 Antimicrobial resistance—a worldwide threat

AMR is the ability of pathogens to grow despite the presence of antimicrobial agents usually
effective for treatment of infections caused by these pathogens (WHO, 2014a, 2014b). It is a
general term including resistance to antibiotics that treat infections caused by bacteria (e.g.
Escherichia coli), parasites (e.g. Plasmodium spp.), viruses (e.g. HIV), and fungi (e.g. Candida
spp.) while antibiotic resistance (ABR) is the specific term used to describe resistance to
antibiotics that occurs in commensal, pathogenic and zoonotic bacteria (WHO, 2014a, 2014b).
These resistance mechanisms are not a new finding but rather a natural evolutionary process that
affects humans, animals, and the environment (plants, soil and water). Indeed, during his Nobel
Prize allocution in 1945, Alexander Fleming, who discovered the first antimicrobial drug, had
already mentioned the possible acquisition of resistance by bacteria (WHO, 2014a, 2014b).
Microorganisms are becoming increasingly resistant to antibiotics with worldwide evolution as a
result of the selective pressure induced through extensive consumption of antibiotics whether
appropriate or inappropriate use (Founou et al., 2016). Bacteria have the remarkable ability to
develop resistance to antibiotics, rendering these drugs less effective in treating infections. The
misuse and overuse of antibiotics have accelerated the emergence of antibiotic-resistant bacteria,
posing a significant global health challenge. (Ventola., et al., 2015)
The occurrence and spread of AMR differ greatly with geographical distribution between
continent and within countries (Laxminarayan et al., 2013). In developed countries, multi-drug
resistant (MDR) bacteria cause infections which result in increasing cost of therapy whereas in
developing ones, the same pathogens considerably enhance mortality. Antimicrobial usage is
strictly controlled in human as well as in animal health in developed countries through several
initiatives and the strategies established for the prevention and containment of AMR illustrate the
significance of this threat in the developed world (Laxminarayan et al., 2013). The risk of
emergence and spread of AMR is worsened in developing countries because treatment of
bacterial infections is empirical and there is usually sub-optimal policy, monitoring and
surveillance systems, diagnosis capability and control of infectious diseases, antibiotic
consumption and AMR detection as well as numerous issues related to the quality and
accessibility of antibiotics (Vila, 2010; Laxminarayan et al., 2013; WHO, 2014b). Indeed,
several factors have been incriminated in the speedy emergence and spread of AMR. These
include poverty, overcrowding, hygiene status, education level, self-medication, poor and limited
supply chain, lack of knowledge about antibiotics, inappropriate use misuse of antibiotics and
chronic diseases –such as tuberculosis, hepatitis, asthma, HIV/AIDS which enhance the
proportion of immune-compromised population who are more prone to infection, requiring
frequent antibiotic treatment resulting in selection pressure for the development or exacerbation
of antibiotic resistance (Vila, 2010; Laxminarayan et al., 2013; WHO, 2014b). AMR is of
paramount importance because many common infectious diseases in the developing world,
especially malaria, HIV/AIDS, respiratory infections (influenza, asthma, tuberculosis, etc.) and
sexual transmissible infections raise the risk of acquisition of resistant pathogens (Vila, 2010;
Laxminarayan et al., 2013; WHO, 2014a, 2014b). Given the important ability of microorganisms
to adapt to their environment, a comprehensive strategy involving key stakeholders promoting
the “One Health” approach is likely to succeed in containing AMR.
 CHAPTER THREE

3.0 MATERIALS AND METHODS

3.1: Study area

Dutsin-Ma is located in Katsina State, which is situated in the Northern part of Nigeria. More
specifically, Dutsin-Ma is located in the northwestern region of Katsina State. It lies
approximately 80 kilometers (50 miles) south of Katsina, the capital city of the state. The
geographic coordinates of Dutsin-Ma are approximately 12.4522° N latitude and 7.4913° E
longitude. The LGA has an area of 527 km 2 and population of 169,671 at as of 2006 census
(NIPOST, 2009). The town is surrounded by a savanna landscape characterized by flat plains,
scattered hills, and the nearby Dutsin-Ma Dam, which provides irrigation for agricultural
activities in the area.

3.2: Sample size and Sampling Technique

A total of 40 samples were collected from 4 locations, while a total of 10 samples was collected
from each location, 2 swab samples from table, 2 knife sharpener, 2 swab samples from the
palm, 2 swab samples from the knife and 2 from the meat. A stratified random sampling
Collecting were employ to ensure representation from various socio-economic group within the
local government area.

3.3: Sample Collection

The surfaces of the meat and butcher equipment was swab using a sterile swab sticks that was
socked in normal saline from different spots in Dutsin-Ma metropolis. One sample per each meat
bench of four meat seller of the same location was collected, 2 swabs from the palm and knife
was also collected for four weeks, The locations include Wednesday market, Hayin gada, , meat
spots opposite school (FUDMA) and Darawa meat spots. The swabs were placed in a medium
(normal saline) in order to proliferate the bacteria on the swab samples, and to maintain the
viability of the bacteria during transportation to the laboratory.

3.4: Determination of Bacteria in samples, isolation and storage of bacteria.

After media preparation, media was pour in each sterile petri dishes. Afterward, a sterile eosin
methylene blue agar, manitol salt agar and Salmonella Shigella agar was streaked in each
inoculated plates. Plates were incubate at 37 0C for 18 to 24 hours. Count on mannitol salt agar is
described as total bacteria count while that on eosin methylene blue agar will be describe as total
Enterobactericeae count. Colonies were picked, and re-streaked on nutrient agar for further
purification, in-order to obtain distinct culture based on their colonial morphologies. (Buchanan
and Gibbons, 1994). Pure Colonies isolates is thereafter store on nutrient agar slants at 4 0C or
further identification and characterization.
3.5: Bacterial identification and characterization.

Isolates stored on nutrients agar slant were identify and characterize using culture media with
biochemical test described below. Afterwards, bacteria is identified by Bergey ,s manual of
systematic bacteriology (Bergey, s et al., 1974) using the biochemical test. Which are listed
below:

3.6: Gram-Stainning: Single isolated pure colony will be smear and heat-fixed on a glass slide
and later flooded with crystal violet for about 60 seconds thereafter rinsed with water and drain.
Afterwards, glass slide will be flood with iodine solution for about 60 seconds and thereafter
rinse with water. Alcohol will be added as a decolorizer and later rinse with water. Finally, slide
is flooded with safranin for about 60 seconds, rinse with water and blot dry. The glass slide was
dried with filter paper, mounted on a microscope and will be observe at 100X oil immersion
objective lenses, bacteria that retained the crystal violet and will appearing blue is considered to
be gram positive while bacteria that retained the counter stain and appeared red is regarded as
gram negative. The cell shape were viewed under the microscope.

Methyl – red Test: Pure overnight grown bacterial culture was inoculated into a sterile tube
containing nutrient broth. The test tube were incubated at 370C for 24hours. After incubation 3-5
drops of the methyl red indicator will be added to this tube and gently roll between the palms to
disperse the methyl red, positive test result turns red and negative test results turns yellow.
Vogues proskauer Test: This test was performed by adding vogues proskauer reagent to the
vogues prokauer broth which has been inoculated with test overnight grown bacterium. A cherry
red color indicates a positive result while a yellow brown color indicates a negative result.

Indole Test: This is used to determine the ability of an organism to split indole from the amino
acid tryptophan using the enzyme tryptophanase. Tryptophan broth will be inoculated with test
overnight grown organism and incubate for 24 hours at 37 0C. Drops of Kovacs reagent is
thereafter added to the broth. Formation of a red ring at the surface of the broth signified a
positive result.

Citrate Test: Pure overnight grown bacteria colonies will be picked with straight wire loop and
inoculate into slope of simmons citrate agar and incubate overnight at 37 0C, if the organism has
the ability to use citrate, the medium changes its color from green to blue

Catalase Test: This test is used to differentiate those bacteria that produces the enzyme catalase
such as Staphylococci from non- catalase producing bacteria such as Streptococci. Two to three
drops of hydrogen peroxide solution is added into a test tube. Then using a sterile glass rod, test
organism is remove from the petri dish and immersed in the hydrogen peroxide solution. An
immediate bubbling indicates a positive result and negative vice versa.

Coagulase Test: This is used to identify Staphylococcus aureus which produce the enzyme
coagulase. Three small test tubes will be labelled i.e. first test tube was labelled T (18-24 hours’
broth culture), the second test tube was labelled Pos (18-24 hours’ broth culture) and the third
tube was labelled T. Thereafter, 0.8ml of the S. aureus was added to the test tube labelled Pos
and also, about 0.8ml of sterile broth was added to the tube labelled Neg. After mixing gently,
the three test tubes were incubated at 35-37 0C. The test tubes were examined and observed for a
clotting reaction after few hours at room temperature.

TSI (triple sugar iron) agar: TSI agar was dispensed in test tubes to give slants and butt, the
butt was stabbed with the inoculation pin and then dragged out through the slant because bacteria
below the agar surface under oxygen limiting conditions that encourage fermentation, then
aerobic growth in the slant principle. It was incubated for 24hours at 370C.
3.7: Determination of antibiotic resistance profile of isolates
Antibiotic-resistant profiles of bacteria were determine by disc diffusion method with antibiotic
sensitivity disc using Antibiotics sensitivity test (AST) (Abtek Biologicals Ltd). Two sets of disc
was used in this study i.e. gram positive and gram negative bacteria sensitivity discs. Antibiotics
present on gram negative disc with their various concentrations. Antibiotics present on gram
negative disc with their various concentrations include: septrin (30ug), sparfloxacin (10ug),
gentamicin (30ug), ciprofloxacin (30ug), chloramphenicol (30ug), tarivid (10ug), streptomycin
(30ug), pefloxacin (30ug), augmentin (10ug) and amoxicillin (30ug) while that for gram positive
disc include: pefloxacin (10ug), ampiclox (30ug), gentamycin (10ug), erythromycin (10ug),
zinnacef (20ug), amoxicillin (30ug), rocephin (25ug), ciprofloxacin (10ug), streptomycin (30ug)
and septrin (30ug). The procedure includes the inoculation of the stored bacteria cultures stored
on slant at 40C into different 9ml of sterile nutrient broths in separate test tubes and thereafter
incubated at 370C in an incubator overnight. Thereafter, the organism was standardized using
MacFarland standard then sterile Mueller Hinton agar that has been previously prepared and
cooled to 550C in water bath was poured into each petri plate and allowed to solidify. Then, the
standardized organism will be spread evenly on the solidified Mueller Hinton agar using a sterile
swab stick then the antibiotic sensitivity disc was later aseptically placed on each of the
solidified plates and incubated at 370C in an incubator overnight.

Zones of inhibition observed around the antibiotics sensitivity discs after 24 hours of incubation
was measured and the length is categorize as resistant, intermediate and sensitive after
comparing with Clinical Laboratory Standard Institute (CLSI) for each bacterium isolate (CLSI,
2013). Thereafter, bacteria isolates which were resistant to three or more classes of antibiotics
were selected as multi drug resistant bacteria.
 CHAPTER FOUR

RESULTS

Occurrences of Escherichia coli, Staphylococcus aureus and Proteus spp from meat and
butcher equipment swab samples collected from various locations in Dutsin-Ma.

A total of 40 samples were examined for this study and 77 isolates were obtained. Amongst this
77 isolated, Staphylococcus aureus was the frequently (37/77, 92.5%), Proteus spp (29/77,
72.5%) followed and E.coli was the least (28/77, 70%).

Table 1: Occurrences of Escherichia coli, Staphylococcus aureus and Proteus spp from
meat and butcher equipment from various locations

Locations E.coli S. aureus Proteus

spp

Total Total Total Total Total Total

positive negative positive negative positive negative

DR 6 (60.0) 4 (40) 10 (100) 0 (0) 8 (80) 2 (20)

WM 10 (100) 0 (0) 10 (100) 0 (0) 10 (100) 0 (100)

HG 7 (70) 3 (30) 10 (100) 0 (0) 7 (70) 3 (30)

OF 5 (40) 5 (50) 7 (70) 3 (30) 4 (40) 6(60)

Total 28 (70) 12 (30) 37(92.5) 3 (7.5) 29 (72.5) 11(27.5)

KEY: DR=DARAWA, WM=WEDNESDAY MARKET, HG=HAYIN GADA, OPPOSITE

FUDMA GATE.
Table 2: Antibiotic resistance pattern of Escherichia coli, Proteus spp and Staphylococcus
aureus isolates from meat and butchers equipments samples from various locations

Antibiotics Escherichia Antibioti Proteus spp Antibioti Staphylococc

coli cs cs us aureus
Total Total Tota Tota Total Total
R S lR lS R S
n(%) n(%) n(%) n(%) n(%) n(%)
Septrin 30(10 0 (0) Ciproflox 6(75 2(25 Pefloxaci 35(89 4(10.
0) acin ) ) n .7) 3)

Chloramph 24 6 (20) Ampicilli 5(62 3(37 Gentami 21(53 18

enicol (80) n .5) .5) cin .8) (46.2)
Sparfloxaci28(93 2 Erythro 2(25 6(75 Ampilox 35(89 4(10)
n .3) (6.7) mycin ) ) .7)
Ciprofloxaci
4 26(86 Amoxicili 7(87 1(12 Zinnacef 39(10 0(0)
n (13.3) .7) n .5) .5) 0)
Amoxicillin30(10 0 (0) Gentami _ _ Amoxacil 35(89 4(10.
0) cin lin .7) 3)
Augmentin 12(40 18(60 Gentami _ _ Rocephin 27(69 12(30
) ) cin .2) .8)
Gentamicin 16(53 14 Tarivid _ _ Ciproflox 20(51 19(48
.3) (46.7) acin .2) .8)
Pefloxacin 19(63 11(36 Strepto _ _ Strepto 22(56 17
.3) .7) mycin mycin .4) (43.6)
Tarivid 22(73 8 Pefloxaci _ _ Septrin 38(97 1(2.6)
.3) (27.7) n .4)

15(50 15(50 Augmen _ _ Erythro 33(84 6(15.

Streptomyc ) ) tin mycin .6) 4)
in
Keys: AMP (ampicillin), GEN (gentamycin), PEF (pefloxacin), E (erythromycin), SXT
(septrin), APX (ampiclox), CPR (ciprofloxacin), CH (chloramphenicol), – (Antibiotics not
tested)
Table 3: Percentage of total selected multidrug resistant isolates from meat and butcher
equipments from various locations

Isolates Knive Palm meat sharpener Table Total

MDR(%)

E.coli 5 7 1 2 6 21(41.3)

S.aureus 4 7 3 1 8 23(50)

Proteus spp 0 1 1 0 2 4(8.7)

Grand 9 (15.2) 15(32.6) 5(10.9) 3(6.5) 16(34.8) 46(100)

total(%)

Key: MDR (Multidrug resistant).

4.5: Phenotypes of multidrug resistant bacteria isolates from meat seller benches, palm and
knife in Dutsin-Ma metropolis.

A total of (46/77, 100%) MRD bacteria were isolated from meat and butcher equipment
collected from this study. Among these MDR bacteria, Staphylococcus aureus was observed to
have the most common (23/46, 50%) resistant phenotype, which were resistant to each of
ampicillin, gentamicin, erythromycin, ampiclox and septrin while Escherichia coli was observed
to followed with (19/46, 41.3%) resistant phenotype and Proteus spp was observed to be the
least (4/46, 8.7%) which were resistant to each of ampicillin, chloramphenicol, gentamicin and
ciprofloxacin respectively.

Phenotypes of multidrug resistant bacteria isolates from meat and butcher equipment
samples from various locations

Bacteria Number (%) Resistant Antibiotics

Escherichia coli 2 AMP, CPR

6 AMP, GEN, PEFX, CPR, SXT

4 AMP, GEN, CPR, SXT, CH

2 AMP, SXT

5 AMP, GEN, AUG, CH

Total 19 (41.3)

Staphylococcus 5 AMP, GEN, ERY, SXT, APX

aureus

6 AMP, GEN, ERY, STR, SXT, APX

4 AMP, GEN, SXT

3 AMP, ERY, STR

5 AMP, GEN, ERY, STR, SXT, APX

 Total 23 (50)

Proteus spp AMP,CIP,ERY,AMC,AMO

4

TOTAL 4(8.7)

Grand total 46 (100)

Keys: AMP (ampicillin), GEN (gentamycin), PEFX (pefloxacin), ERY (erythromycin), SXT
(septrin), APX (ampiclox), CPR (ciprofloxacin), CH (chloramphenicol)
 CHAPTER FIVE

5.0 Discussion, Conclusion and Recommendation

5.1 Discussion

Animals are a great source of protein. When eaten as food, they can make up over 60% of the
dietary protein intake by adults, especially in rural areas (Adeleye OA, 2008).In Nigeria, "meat is
a much-cherished delicacy that cuts across socio-economic, age, religious and educational
barriers (Adebayo-Tayo BC, Onilude AA, 2008). Meat is a most cherished food delicacies in
Nigeria, but can be a source of dissemination of Multi-drug Resistant (MDR) bacteria (2013 ).
The organisms such as S. aureus, E. coli and Proteus spp were isolated and characterized based
on biochemical test. The present study revealed that all of the isolates of E. coli, Proteus spp and
Staphylococcus aureus from commercial meat and butcher equipment were resistance to multiple
antibiotics which coincided with the findings of Zhao et al. (2005), Guerra et al. (2003) and
Islam et al. (2008). Multiple antimicrobial resistance might have happened due to indiscriminate
use of antibiotics, chemotherapeutics and or disperse of drug resistant microorganism in the
environment (Van de Boogard and Stobberingh, 2000).

The presence of Escherichia coli, Proteus spp, Staphylococcus aureus reported in this study is
also in agreement with the findings of Adesokan et al. (2005) who reported the presence of
Escherichia coli among other organisms in selected brands of beer in Nigeria. The occurrences of
Escherichia coli. Proteus spp and Staphylococcus aureus from meat and butcher equipment from
various location. A total of 40 samples were examined for this study and 77 isolates were
obtained. Amongst this 77 isolated, Staphylococcus aureus was the frequently (37/77, 92.5%)
isolated bacteria, Proteus spp (29/77, 72.5%) followed and Escherichia coli (28/77, 70%) was the
least isolated bacteria this is not in agreement with the finding of (Drzewiecka, 2016 and Nahar
et al., 2014) due to sampling location, sampling method, types of meat and laboratory techniques
as well as analytical procedures due to Air pollution, water quality and noise pollution; the
environment was very open (market area).

The antibiotic resistance profiles of Escherichia coli, Proteus spp and Staphylococcus aureus
isolate from meat and butcher equipment sample among the isolated bacteria, Staphylococcus
aureus was observed highest (100%) resistance to each of Septrin and amoxicillin and least
(11.8%) resistance to augmentin, Escherichia coli was observed to show highest (100%)
resistance to zinnacef and least (51.2%) resistance to gentamicin and Proteus spp was observed
to show least resistance this is in agreement with the finding of ( Cardenas et al., 2015, Ghafur et
al., 2014 and Silva et al., 2017) whose finding investigated antibiotic resistance in E.coli and
enterococcus spp isolated in pork meat in india, antibiotic resistance pattern of Enterococcus spp
and S.aureus isolated from chicken meat, poultry, beef and chicken in Colombia, emphasizing
the need the need for strick antibiotic usage guidelines in poultry farming and implication for
food safety and public health and (Dammy Annibi 2021) who isolate S.aureus and E.coli in
chicken meat in B.U.K gate Kano, Nigeria and (Godiya et al., 2024) who isolates bacteria in fish
sold at Dutsin-ma market.

The highest percentage of multidrug resistant bacteria isolated from this study was observed to
be Staphylococcus aureus (50%), Escherichia coli was observed to follow (41.3%) and Proteus
spp was observed to be the least (8.7%) amongst the total selected multidrug resistant bacteria
This is in agreement with the finding of ( Eze et al., 2015) who isolates salmonella,shigella s and
E.coli assesse from suya and dried fish for multi-drug resistant bacteria sold at kaduna central
market.

The phenotypes of multidrug resistant bacteria isolates from meat and butcher equipment around
Dutsin-Ma town. A total of (46/77, 100%) MRD bacteria were isolated from meat and butcher
equipment collected from this study. Among these MDR bacteria, Staphylococcus aureus was
observed to have the most common (23/46, 50%) resistant phenotype, which were resistant to
ampicillin, gentamycin, erythromycin, ampiclox and Septrin while Escherichia coli was observed
to followed (19/46, 41.3%) and Proteus spp was observed to have the least (4/46, 8.7%) resistant
phenotype, which were resistant to ampicillin, chloramphenicol, gentamicin and ciprofloxacin
respectively. This poses a serious public health concern to the consumers of these meats and is
an indicator of the level of hygienic practices at the various meat stalls in Dutsin-Ma metropolis.
The microbiological safety of food is achieved by as far as possibly ensuring the absence of
pathogenic microorganisms and by all means preventing their multiplication (Okonko et al.,
2008).
The Hazard Analysis Critical Control Point (HACCP) concept is used to identify microbiological
vulnerable points in the food production process and processing, and to determine the most
appropriate methods of control to be applied.

The environmental reservoirs (surfaces, equipment, water and air in these facilities) and hygienic
practice; proper hygiene practices are essential to minimize contamination risks. This can be
thorough cleaning and disinfection of surface and equipment as well as water sanitation and
hygiene; access to safe potable water used for cleaning the meat,
good personal and environmental hygiene practice to prevent the spread of pathogens and
sanitization tools can be a major cause of this. Also, the indiscriminate use of these tools in the
cutting of the intestines constitute another possible source of meat contamination as the gut
contents can easily be spread to the table and the entire meat to be sold
5.2 CONCLUSION

The study provides valuable insights into the prevalence, antimicrobial resistance profile and
implication of multidtug-resistant staphylococcus aureus, E. coli and proteus spp in dutsin-ma,
katsina state.

The study revealed a significant presence of pathogenic bacteria including Escherichia coli,
Proteus spp, and Staphylococcus aureus in meat and butcher equipment samples, indicating
potential health risks associated with consuming contaminated meat products.

Antimicrobial susceptibility testing demonstrated widespread resistance among the isolated

bacteria, particularly Staphylococcus aureus, which showed resistance to multiple antibiotics.
This poses a serious challenge for effective treatment of infections and suggests a need for
improved antibiotic stewardship in agricultural practices. A substantial proportion of the bacteria
exhibited multidrug resistance, with Staphylococcus aureus being the most prevalent multidrug-
resistant pathogen. This highlights the urgency of addressing antibiotic misuse and implementing
rigorous hygiene protocols to curb the spread of resistant strains addressing these findings
require collaborative effort from public health authorities, veterinary service and meat industries
to ensure food safety and mitigate antimicrobial resistance risks. The presence of these bacteria
on the meat, table, palm, sharpener and knife is of great health concern and call for proper
sanitization and hygiene of the environment.

5.3 RECOMMEDATION
It is recommended that there should be adequate enforcement of existing:

1. Enhance hygiene practice: Implement stringent hygiene protocol in meat processing facilities
and butcher shops minimize bacterial contamination and also regular cleaning and
disinfection of equipment and surface should be enforce to reduce the spread of pathogens.
2. Antimicrobial stewardship: Promote responsible use of antibiotics in livestock and poultry
farming to mitigate the development and the spread of antimicrobial resistance.
3. Surveillance and monitoring: Establish routine surveillance programs to monitor
antimicrobial resistance patterns in foodborne pathogens from meat products. Enhance
collaboration between public health authorities, veterinary service, and food regulatory
agencies to track trend and implement timely interventions.
4. Research and policy development: support further research on antimicrobial resistance in
meat production system to inform evidence-based policies and intervention. Advocate for
policy initiatives that promotes sustainable agriculture practice and enhance food safety
standard.
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 APENDIX

Morphological features of isolate

Media used Samples Colony characteristics

EMB T Small, smooth edges, round, greenish

metallic sheen

P Small, smooth edges, round, greenish

metallic sheen

K Small, smooth edges, round, greenish

metallic sheen

Small, smooth edges, Round, greenish,

S
metallic sheen

Small, smooth edges, Round, greenish,

M metallic sheen

MSA T Round, small, yellowish, smooth edges,

P Round, small, yellowish, smooth edges,

K Round, small, yellowish, smooth edges,

S Round, small, yellowish, smooth edges,

M Round, small, yellowish, smooth edges,

SSA T

Swarmed pink, irregular, with a black

 P center

Swarmed pink, irregular, with a black

center
S
Swarmed pink, irregular, with a black
M
center

KEY: EMB=EOSIN METHYLENE BLUE, MSA=MANNITOL SALT AGAR, and

SSA=SALMONELLA SHIGELLA
AGAR.

T=TABLE, P=PALM, K=KNIFE, S=SHARPENER, M=MEAT

APENDIX 2

Gram’s reaction on the isolates

Samples Gram staining reaction

D(M,K,T,P,S) Gram negative, pink, short rods, arranged in singles and paired,
gram positive, purple, cocci, arranged in clusters, Gram negative,
pink, rod in shape, arranged in bacillus.

W(M,K,T,P,S) Gram negative, pink, short rods, arranged in singles and paired,
gram positive, purple, cocci, arranged in clusters, Gram negative,
pink, rod in shape, arranged in bacillus.

H(M,K,T,P,S) Gram negative, pink, short rods, arranged in singles and paired,
gram positive, purple, cocci, arranged in clusters, Gram negative,
pink, rod in shape, arranged in bacillus.

F(M,K,T,P,S) Gram negative, pink, short rods, arranged in singles and paired,
gram positive, purple, cocci, arranged in clusters, Gram negative,
pink, rod in shape, arranged in bacillus.

KEY: D=Darawa, W=Wednesday market, H=Hayin gada, F=FUDMA gate

M=MEAT, K=KNIVES, T=TABLE, P=PALM, S=SHARPENER

APENDIX 3

Sample Grm Catalase coagulase citrate Indole MR VP TSI Presumptive

RXN organism

D1T + + + - - - - S. aureus

D2M + + + - - - - S. aureus

W1H + + + - - - - S. aureus

W2K + + + - - - - S. aureus

W2T + + + - - - - S. aureus

F1M + + + - - - - S. aureus

F1K + + + - - - - S. aureus

F1H + + + - - - - S. aureus

F1S + + + - - - - S. aureus

F2T + + + - - - - S. aureus

F2H + + + - - - - S. aureus

F2H + + + - - - - S. aureus
 Escherichia
coli

D2T - - + + - A/A+G E. coli

D2S - - + + - A/A+G E. coli

W1S - - + + - A/A+G E. coli

W2M - - + + - A/A+G E. coli

F1M - - + + - A/A+G E. coli

H2H - - + + - A/A+G E. coli

H2K - - + + - A/A+G E. coli

Proteus spp

D1K - + + + - A/A,GAS Proteus spp

D1M - + + + - A/A,GAS Proteus spp

W1M - + + + - A/A,GAS Proteus spp

F1S - + + + - A/A,GAS Proteus spp

F1T - + + + - A/A,GAS Proteus spp

H1H - + + + - A/A,GAS Proteus spp

F1S - + + + - A/A,GAS Proteus spp