Final Dissertation

MALAWI UNIVERSITY OF SCIENCE AND TECHNOLOGY
ACADEMY OF MEDICAL SCIENCES
DEPARTMENT OF BIOLOGICAL SCIENCES
UNDERGRADUATE RESEARCH DISSERTATION
ASSESSING THE EFFECTIVENESS OF ANTIBACTERIAL ACTIVITY OF HONEY AND

GINGER EXTRACTS MIXTURE AGAINST BACTERIAL ISOLATES CAUSING URINARY
TRACT INFECTIONS FROM BLANTYRE, MALAWI.
BY
ANTHONY NKHUNGUNI (MMB/026/19)
SOLIUM NGULUWE (MMB/044/16)
CHAWEZI CHIRWA (MMB/014/19)
SUBMITTED TO UNDERGRADUATE’S OFFICE
IN PARTIAL FULFILMENT FOR THE REQUIREMENTS FOR
THE AWARD OF BACHELOR OF SCIENCE DEGREE IN MEDICAL MICROBIOLOGY
FEBRUARY, 2024
DECLARATION
We, hereby, declare that this dissertation titled “Assessing The Effectiveness Of Antibacterial
Activity Of Honey And Ginger Extracts Mixture Against Bacterial Isolates Causing Urinary Tract
Infections From Blantyre, Malawi” is based on the original research work of Anthony Nkhunguni,
Chawezi Chirwa and Solium Ngulube is submitted for the award of Bachelor of Science in
Medical Microbiology at the Malawi University of Science and Technology. Indebtedness to other
research works used in this research has been duly acknowledged at the relevant places. It is also
declared that this work has not been submitted for the award of bachelor’s degree or any other
degree in this or any other university.
Anthony Nkhunguni 4th February 2024
Chawezi Chirwa 4th February 2024
Solium Nguluwe 4th February 2024
SUPERVISOR
Dr. Willard Mbewe 1st March 2024
I
DEDICATION
We dedicate this work to our parents, Mrs. Mary Nkhunguni, Miss Yvonnie Gondwe and Mr.
Nguluwe for the assistance rendered to us to complete our four-year study at MUST through
their financial, spiritual and moral support and to all our MUST lecturers, who have provided us
the knowledge which we have used in this research.
II
ACKNOWLEDGEMENTS
Firstly, we give all thanks and glory to Almighty God for keeping us in good health and making
everything possible so that the project was successful despite our numerous setbacks. We would
also like to express a very special thanks to our supervisor Dr. Willard Mbewe for his assistance
both financially and academically through-out the course of our research. Many thanks to Mr.
Madalitso Muhakeya for his guidance during the analytical stage of this research’s findings, we
really appreciate you. We also appreciate working stuff of Queen Elizabeth Central Hospital for
allowing us to conduct our research using samples from their facility, with honorary mentions of
Edward Phillipo, Thoko Noniwa, Mtendere Pensulo and Falesi Kayesa . Lastly, we thank the
laboratory technicians and assistants from the Malawi University of Science and Technology for
their assistance and support during the laboratory experiments.
III
ABSTRACT
The global increase in resistance to antimicrobial agents has emerged as a significant contributor
to morbidity and mortality, despite the initial belief in victory against microorganisms with the
introduction of antibiotics in the 1900s. It became evident that these microorganisms could
develop resistance to any drugs employed, prompting scientific research into the use of herbs and
plants as potential antimicrobials.
This study aimed to assess the effectiveness of antimicrobial activity from a mixture of honey and
ginger at different concentrations against bacterial isolates, namely Escherichia coli, Klebsiella
Pneumoniae, Methicillin-resistant Staphylococcus Aureus, and Enterococcus faecalis - causative
agents for most urinary tract infections.
A total of 16 bacterial samples were collected from Queen Elizabeth Central Hospital and Malawi
University of Science and Technology which were then cultured on nutrient agar for resuscitation
then later sub-cultured on Muller-Hinton agar for antimicrobial susceptibility testing where filter
papers impregnated with honey, ginger, the mixture of honey and ginger and clinically used
antibiotics as controls.
4 resistant bacterial isolates were collected from Queen elizabeth central hospital and were
cultured on NA for resuscitation and later subcultured on MHA for antimicrobial sensitivity
testing where filter papers impregnated with honey, ginger and mixture of honey and ginger and
clinically used antibiotics as controls.
The results in form of zones of inhibitions were read and recorded after 24 hours of incubation. It
was observed that ginger had mean antimicrobial effects ranging from 11mm to 20mm against all
the named bacterial isolates; whereas honey showed an effect range between 17mm to 34mm. The
mixture of honey and ginger exhibited an even greater effect range between 26mm to 37mm.
These findings suggest that both ginger and honey have antimicrobial properties but their
synergistic use as a mixture produced more effective inhibition zones when used at higher
concentrations.
This experiment demonstrated the potency and effectiveness of using mixtures made up of honey
and ginger as remedies for treating urinary tract infections caused by bacterial colonization.
Furthermore, it suggests the need to conduct more and deeper research on utilizing them both as
IV
raw materials for developing new drugs while taking advantage of their natural antibiotic
properties.
Key words: Escherichia coli, Klebsiella Pneumoniae, Methicillin-resistant Staphylococcus

Aureus, Enterococcus faecalis, honey, ginger and Antimicrobial susceptibility tests.
V
TABLE OF CONTENTS
Contents
DECLARATION................................................................................................................................I
DEDICATION...................................................................................................................................II
ACKNOWLEDGEMENTS.............................................................................................................III
ABSTRACT.....................................................................................................................................IV
LIST OF FIGURES.......................................................................................................................VIII
LIST OF TABLES...........................................................................................................................IX
ABBREVIATIONS............................................................................................................................1
1.0 INTRODUCTION........................................................................................................................2
1.1. Bacteria Species and Diversity................................................................................................2
1.1.2 Urinary tract infections (UTI) in humans..........................................................................3
1.1.3 Synthetic antibiotics against UTIs in clinical use.............................................................3
1.2 Exploring natural antibacterial remedies..................................................................................4
1.3 problem statement....................................................................................................................5
1.4 objectives..................................................................................................................................6
1.4.1 Main Objective..................................................................................................................6
1.4.2 Specific Objectives............................................................................................................7
1.5 Hypothesis of the Study...........................................................................................................7
1.5.1 Main Hypothesis...............................................................................................................7
1.5.2 Specific Hypotheses..........................................................................................................7
1.6 Significance of the study..........................................................................................................7
2.0 LITERATURE REVIEW.............................................................................................................9
2.1 Bacterial species associated with UTI......................................................................................9
2.2Antimicrobial properties of honey and ginger........................................................................12
VI
3.0 MATERIALS AND METHODS...............................................................................................18
3.1 Materials.................................................................................................................................18
3.2 Study Design..........................................................................................................................18
3.3 Study Setting..........................................................................................................................18
3.4 Sampling Techniques.............................................................................................................19
3.5 Preparation of Growth Media.................................................................................................20
3.5.1 Nutrient Agar (NA).........................................................................................................20
3.5.2 Mueller-Hinton Agar (MH).............................................................................................21
3.6 Sample Processing and Culture..............................................................................................22
3.7 Processing of Natural Remedy Mixture.................................................................................23
3.7.1 Honey..............................................................................................................................23
3.7.2 Ginger extracts................................................................................................................23
3.7.3 Combination of honey and ginger extracts.....................................................................24
3.8 Antimicrobial Susceptibility Testing.....................................................................................25
4.0 RESEARCH FINDINGS...........................................................................................................27
4.1 Control antibiotics..................................................................................................................27
4.2 Antimicrobial effects of honey, ginger and honey-ginger mixture........................................29
5.0 DISCUSSIONS..........................................................................................................................38
5.1 Control antibiotics..................................................................................................................38
6.0 CONCLUSIONS AND RECOMMENDATIONS.....................................................................41
6.1 Conclusions............................................................................................................................41
6.2 Recommendations..................................................................................................................41
REFERENCES.................................................................................................................................43
APPENDICES..................................................................................................................................50
Appendix 1: Calculations for different concentrations of honey and ginger...............................50
VII
Appendix 2: Certificates of approval from MUSTREC and QUECH Research Office..............52
Appendix 3: Growth media details...............................................................................................53
Appendix 4: Some of the equipment used...................................................................................55
LIST OF FIGURES
Figure 1: Map showing the study area.............................................................................................19

Figure 2: Bacterial isolate samples...................................................................................................20
Figure 3: Media preparation.............................................................................................................22
Figure 4: Sub-cultured bacterial isolates..........................................................................................23
VIII
Figure 5: Preparation of honey and ginger.......................................................................................24
Figure 6: Preparation of diffusion discs for AST.............................................................................25
Figure 7: Inhibition zones by honey and ginger on enterococcus spp.............................................29
Figure 8: Inhibition zones by hone and ginger on all bacterial species...........................................30
Figure 9: Figure 1: Average inhibition in mm for ginger powder water extracts, honey, mixture of
honey-ginger extracts and standard antibiotic discs (Ciproflaxicin, Co-trimaxole, Vancomycin and
Amoxicillin).....................................................................................................................................35
Figure 10: Approval of research by QUECH..................................................................................52
Figure 11: MUSTREC approval of research....................................................................................53
Figure 12: Nutrient agar...................................................................................................................54
Figure 13: Mueller-Hinton agar.......................................................................................................55
LIST OF TABLES
Table 1: shows Ciprofloxacin against Klebsiella Pneumoniae with EUCAST breakpoints

interpretation....................................................................................................................................27
Table 2: shows sxt-trimethoprim against Escherichia coli with EUCAST breakpoints interpretation
..........................................................................................................................................................27
Table 3: shows vancomycin against Methicillin-resistant Staphylococcus Aureus with EUCAST
breakpoints interpretation.................................................................................................................28
IX
Table 4: shows amoxicillin against Enterococcus faecalis with EUCAST breakpoints interpretation
..........................................................................................................................................................28
Table 5: showing zones of inhibition by 100% honey, ginger and their mixture in their natural
form against different bacterial isolates...........................................................................................31
Table 6: showing zones of inhibition by 75% honey, ginger and their mixture against different
bacterial isolates...............................................................................................................................32
Table 7: showing zones of inhibition by 50% honey, ginger and their mixture against different
bacterial isolates...............................................................................................................................33
Table 8: Results of one-way ANOVA and descriptive statistics of the inhibitions of honey, ginger
extracts and honey –ginger extracts mixtures on the bacteria isolates.............................................35
Table 9: Results of Turkeys Honestly Significant Difference (HSD) test of honey-ginger extract
mixtures on the test bacteria isolates................................................................................................36
X
ABBREVIATIONS
AST: Antimicrobial Susceptibility Testing
UTI: Urinary Tract Infection(s)
MRSA: Methicillin-Resistant Staphylococcus Aureus
AMR: Antimicrobial resistance
EUCAST: European Committee of Antimicrobial Susceptibility Tests

CHAPTER 1
1.0 INTRODUCTION
1.1. Bacteria Species and Diversity

Bacteria are a group of microscopic, single-celled organisms that inhabit virtually all environments, including
soil, water, organic matter, and the bodies of multicellular animals. Bacteria are distinguished in part by their
morphological and genetic features; for instance, they may have spherical, baccili, or spiral shapes. They also
can be divided into two main groups, gram-positive or gram-negative, based on the structure of their cell wall
and their reaction to the gram stain. Many bacteria swim by means of flagella. The DNA of most bacteria is
found in a single circular chromosome and is distributed throughout the cytoplasm rather than contained
within a membrane-enclosed nucleus. Though some bacteria can cause food poisoning and infectious diseases
in humans, most are harmless and many are beneficial. They are used in various industrial processes,
especially in the food industry for example the production of yogurt, cheeses, and alcohol (McCutcheon,
2021). As prokaryotic organisms (having no membrane-bound nucleus), they are distinct from eukaryotes.
Bacteria are linked to both illnesses and advantages, they have a dual function in human health. On the one
hand, a variety of human illnesses can be caused by harmful bacteria like Mycobacterium tuberculosis, which
causes tuberculosis, or Clostridium difficile, which causes severe gastrointestinal infections. Medical
attention is necessary since these infections have the potential to be fatal. Contrarily, probiotics—beneficial
bacteria—are essential for digestion and nutrition absorption in the human gut. Additionally, they aid in
vitamin production, immune system bolstering, and defense against dangerous microorganisms. For general
human health, it is crucial to maintain a balance between eradicating dangerous bacteria and encouraging the
growth of helpful ones. (Sundar et al., 2023). The most common bacteria species implicated in the etiology
of UTI include; Klebsiella pneumoniae., Escherichia coli, Enterococcus spp., Pseudomonas aeruginosa,
Proteus mirabilis and staphylococcus saprophyticus with over 80% of UTIs being caused by E. coli (Ewnetu,
et al., 2013)
1.1.2 Urinary tract infections (UTI) in humans

Urinary tract infections (UTIs) are one of the most common infectious complications, affecting approximately
150 million people worldwide each year (Snegarova-Toneva et al., 2023). Urinary Tract Infections are
common infections that happen when bacteria, often from the skin or rectum, enter the urethra, and infect the
urinary tract. The infections can affect several parts of the urinary tract, but the most common type is a
bladder infection (cystitis). Kidney infection (pyelonephritis) is another type of UTI (Reyes and Koyle, 2023).
Some people are at higher risk of getting a UTI such as those who are immunocompromised and also UTIs are
more common in females because their urethras are shorter and closer to the rectum. This makes it easier for
bacteria to enter the urinary tract. Other factors that can increase the risk of UTIs: A previous UTI, Sexual
activity, Changes in the bacteria that live inside the vagina, or vaginal flora (Snegarova-Toneva, et al 2023)
For example, menopause or the use of spermicides can cause these bacterial changes, Pregnancy, age (older
adults and young children are more likely to get UTIs), Structural problems in the urinary tract, such as
enlarged prostate, Poor hygiene, for example, in children who are potty-training. Symptoms of a bladder
infection can include: Pain or burning while urinating, Frequent urination, Feeling the need to urinate despite
having an empty bladder, bloody urine, pressure or cramping in the groin or lower abdomen. Symptoms of a
kidney infection can include: fever, chills, lower back pain or pain in the side of your back, nausea or
vomiting.
1.1.3 Synthetic antibiotics against UTIs in clinical use

Synthetic antibiotics are used to treat urinary tract infections (UTIs). E. coli bacteria are the main culprits
behind UTIs, which can be unpleasant and disruptive to everyday life. UTIs are frequently treated with
synthetic antibiotics including ciprofloxacin, nitrofurantoin, and trimethoprim-sulfamethoxazole. They
function by stopping the spread of germs and curing the ailment. In order to overcome resistance and
guarantee effective treatment results, effective UTI care entails prudent antibiotic administration, regular
monitoring, and continuing research into alternative therapies (Caneschi et al., 2023). Several works have
been documented on use of synthetic antibiotics against UTI related bacterial (Bondad-Reantaso et al., 2023).
Generally, therapeutic treatment given to patients with UTIs caused by bacteria is antibiotic-based, antibiotics
are powerful substances used to treat bacterial infections, these formulated antibiotics target bacteria to cause
death with different mechanisms (Allen et al., 2023). They work by targeting and killing or inhibiting the
growth of bacteria, helping the body's immune system to fight off the infection. These medications have been
a cornerstone of modern medicine, saving countless lives. The most common antibiotics used in Malawian
hospitals to treat UTIs are Ceftriaxone, amoxicillin, Ciprofloxacin and Levofloxacin (Malawi Standard
Treatment Guidelines 2023).
1.2 Exploring natural antibacterial remedies
Natural remedies, are compounds found in nature that possess antibacterial properties. These natural
alternatives have been used for centuries in traditional medicine. They include substances like garlic, ginger,
oregano oil, and turmeric, which have shown antibacterial effects and can support the body's defense against
infections. However, of most important to this research are two natural remedies namely Honey and ginger:
Honey which is the scientific name for honey. The scientific name Honey has a Latin origin whereby genus
name Apis is Latin for "bee", and mellifera is the Latin for "honey" (DeGrandi and Flenniken, 2022). The
western honey bee or European honey bee is the most common of the 7–12 species of honey bees worldwide
and hence being the most common honey maker in the tropics in actuality Malawi.
Honey is a natural substance produced and stored in the honeycombs by honeybees with carbohydrates
constituting about 95 to 97% of the dry weight of honey. Fructose and glucose are the most predominant
sugars present and responsible for most of the physical and nutritional characteristics of honey. The volatile
compound found in honey includes alcohols, ketones, aldehydes, acids, esters and terpenes. Phenolic acids
(benzoic and cinnamic acids) and flavonoids contribute significantly to the therapeutic capacity of honey
which varies greatly depending on the floral source. Different studies have shown honey to have
antimicrobial effect, anti-inflammatory effects, anti-oxidant effects and boosting of the immune system.
Common Apinae honey bees honey (Honey honey) and stingless honeybees honey are the two types of
honeys found in the world. The common honey bees and stingless bees (sub family Meliponinae) are grouped
in the family Apidae (Ewnetu et al., 2013). This chemical composition give honey its probable antimicrobial
activity against other previously tested isolates of bacteria.
Lately, it has been observed that beekeeping has been an overgrowing industry in Malawi as most farmers
have been switching from charcoal to beekeeping which is more valuable in terms of financial benefits thus
increasing the availability of natural honey in Malawi at even fairly affordable prices to both tourists and the
locals (Resilient Food Systems Newsletter Sept,2022). On the other hand, zingiber officinale is the scientific
name for ginger, its name originates from Greek zingiberis, which comes from the Sanskrit name of the spice,
singabera (Hottum and Kress, 2023). It is an herbaceous perennial plant of the family Zingiberaceae,
probably native to southeastern Asia, or its pungent aromatic rhizome used as a spice, flavoring, food, and as
a medicinal herb. Ginger is well-identified for a variety of health benefits, including pharmacological effects,
antioxidant,anti-bacterial, anti-inflammatory among others phenolics and terpene molecules are among the
main active ingredients of ginger. Therefore, ginger can be expected as a plant with antibacterial activity
against both gram positive and gram-negative bacteria due to the composition of its phytochemicals.
1.3 problem statement

Urinary tract infections (UTIs) represent a prevalent global health concern. UTIs are a major cause of
morbidity and mortality worldwide, affecting millions of individuals annually, 2022 estimates for the entire
world included 404.61 million cases, 236,790 fatalities, and 520,200 DALYs. In example, a 2.4-fold increase
in fatalities was seen between 1990 and 2021 (Yang et al., 2022), and the age-standardized mortality rate
(ASMR) increased from 2.77/100,000 to 3.13/100,000. According to epidemiological data (WHO, 2023),
UTIs are responsible for a substantial portion of the global disease burden, with approximately 150 million
UTIs reported each year. In Malawi, UTIs have become a significant public health issue, contributing to the
escalating burden of infectious diseases (Malawi Ministry of Health, 2020). Several risk factors contribute to
the high incidence of UTIs globally, these factors include socioeconomic disparities, inadequate sanitation
facilities, and limited access to clean water in certain areas exacerbate the vulnerability of the population to
UTIs (Storme et al., 2022). Additionally, immunocompromised individuals, such as those living with
HIV/AIDS, face an increased risk of UTIs, further compounding the problem which is a major risk to
populations in Malawi as there is a growing rate of immunosuppressed individuals (UNAIDS, 2021 and
Storme et al.,2022).
The primary approach to UTI management in Blantyre, as in many parts of the world, relies heavily on the
use of antibiotics. However, the widespread and indiscriminate use of antibiotics has led to the development
of antibiotic-resistant strains of bacteria, posing a grave threat to public health (McCallin et al., 2023).
Moreover, the accessibility and affordability of antibiotics remain significant challenges for many individuals
in developing countries such as Malawi (World Bank, 2018).
Despite the potential of natural herbs and remedies to offer alternative treatments for UTIs, there is limited
scientific research on their efficacy and proper usage according to previous studies (Kehkashan et al.,2022,
Budhi et al., 2022, Maner 2022). Local practices often involve traditional herbal medicine, such as a mixture
of Honey and ginger, which is believed to have antibacterial properties. However, there is a dearth of rigorous
studies that scientifically establish the effectiveness of such remedies against the bacterial isolates responsible
for UTIs and established pharmacological uses of such natural remedies such as the appropriate mixing ratios
for maximum antibacterial activity (Kehkashan et al.,2022)
By evaluating the efficiency of the Honey and ginger combo against bacterial isolates causing UTIs in a
Malawian environment, this research attempts to close this important information gap. With the goal of
contributing to the diversification of UTI management strategies and the decrease in antibiotic resistance, we
aim to provide evidence-based insights into the potential of this natural remedy as an alternative or adjunctive
treatment for UTIs through methodical investigation and evaluation.
1.4 objectives
This section seeks to provide an overview of the main and specific objectives of the study.
1.4.1 Main Objective

To assess the effectiveness of the mixture of Honey and ginger as an antibacterial agent against bacterial
isolates that cause UTIs namely; Methicillin-resistant Staphylococcus Aureus, Escherichia coli, K.
pneumoniae, E. coli, Enterococcus faecalis.
1.4.2 Specific Objectives

 To describe (Evaluate or assess can be a better word) the synergistic antibacterial activity of the
mixture of Honey and ginger as a combination therapy against bacterial isolates that cause Urinary
tract infections.
The above specific objective appear to be the main objective too.Can be divided into three ie.
1. Assess antibacterial activity of ginger on…
2. Assess antibacterial activity of honey on…
3. Assess antibacterial activity of a mixture of ginger and garlic on…
You also need to have a specific objective that will talk about comparison between ginger, honey and
mixture….Refer to the analysed data…This must appear here.
 To compare the effectiveness between use of synthetic antibiotics and natural remedy as
antibacterial agents against UTI-causing bacteria isolates.
 To determine the appropriate mixing ratios of Honey and ginger that produce best antibacterial
effect.
Remove this objective as your data lacked proper concentrations for the extracts
1.5 Hypothesis of the Study

Presented in this section are the null hypotheses to the objectives of this study in a respective order.
1.5.1 Main Hypothesis
It is anticipated that the mixture of Honey and ginger will be highly effective as an antibacterial agent against
bacteria isolates that cause UTIs.
1.5.2 Specific Hypotheses

 The mixture of Honey and ginger will have a high bactericidal/static effect as a combination
compared to the antibacterial activities of Honey and ginger as single formulations.
 It is expected that synthetic antibiotics for clinical use will have a high efficacy at killing UTI-
causing bacteria as compared to Honey and ginger both as a mixture or single dose formulations.
 It is expected that a mixture with high percentage of ginger to that of Honey will produce effective
antibacterial activity since honey is highly effective than ginger from previous studies. Remove this
hypothesis
1.6 Significance of the study

Studying the effectiveness of a combination therapy using natural remedies like Honey and ginger in the field
of microbiology in Malawi holds significant potential for addressing several pressing healthcare challenges.
This approach can help tackle issues such as antibiotic resistance, scarcity of antibiotics in some hospitals,
and the need to explore alternative and complementary therapies. The rise of antibiotic resistance is a global
health crisis, and Malawi is no exception. Overuse and misuse of antibiotics have led to the emergence of
drug-resistant pathogens, making once treatable infections increasingly difficult to manage. The effectiveness
of honey and ginger as natural remedies against a wide range of microorganisms, not only limited to bacteria,
has been documented in scientific literature. By studying the combination therapy of honey and ginger, we
can explore their synergistic antimicrobial properties, potentially providing an alternative or adjunct to
traditional antibiotics. The need to use natural remedies and combination therapies is underscored by the rich
traditional knowledge and practices in Malawi. Honey and ginger have been used for centuries in traditional
medicine for their healing properties. Incorporating these natural remedies into modern healthcare practices
through evidence-based research can bridge the gap between traditional and conventional medicine. It allows
us to tap into the wealth of indigenous knowledge and harness the therapeutic potential of natural
antimicrobials.
The study of combination therapy acknowledges the holistic nature of healthcare. Many infections and
diseases involve complex interactions between pathogens and the host's immune system. Combining honey
and ginger, each with its unique bioactive compounds, may provide a more comprehensive approach to
combat infections. Honey, for instance, possesses not only antimicrobial properties but also wound-healing
and anti-inflammatory effects. Ginger, on the other hand, offers antioxidant and anti-inflammatory benefits.
Hence this study may help to provide insights on the antimicrobial nature of honey and ginger from Malawi
and open new research areas such as phytochemistry for further exploration of their antimicrobial properties
and active ingredients.
CHAPTER 2
2.0 LITERATURE REVIEW
2.1 Bacterial species associated with UTI

Urinary tract infections have been described since ancient times with the first documented description in the
Ebers Papyrus dated to c. 1550 BC (Mutonyi et al., 2023). It was described by the Egyptians as "sending forth
heat from the bladder". Effective treatment did not occur until the development and availability of antibiotics
in the 1930s before which time herbs, bloodletting and rest were recommended (Mutonyi et al., 2023).
Urinary tract infections (UTIs) are among the most common bacterial infections worldwide and encompass a
broad spectrum of diseases, ranging from uncomplicated cystitis to life-threatening urosepsis. Studies using
data from the Global Burden of Disease (GBD) show that in 2019 the absolute number of UTI cases had
increased by 60.50% since 1990, to a total of 404.61 million patients (Zeng et al., 2022). The age-
standardized incidence of UTIs was 3.6 times higher for females, and UTI incidences increased with age in
teenagers, peaking around 35 years (Zeng et al., 2022).
The most frequent bacterial infections in women, particularly in pregnant women, are UTIs. For instance, a
UTI during pregnancy is thought to occur between 30 and 50 percent of the time in sub-Saharan Africa
(Warkulwiz et al., 2017). Due to the high incidence rate, frequent recurrence, and, in some cases, lack of
reporting, this has become an issue in the medical community (Warkulwiz et al., 2017).
Due to a lack of awareness, stigma, and accessibility to a clinic for diagnosis, few UTI infections are detected
quickly in developing nations with underdeveloped healthcare systems. According to World Health
Organization, UTI symptoms might include, but are not limited to, discomfort or burning when urinating,
frequent urination, lower abdominal pain, and murky, bloody, or foul urine. UTIs can harm the kidneys and
bladder, which can result in more severe issues or even death if they are not addressed. If untreated, UTIs in
pregnant women can have a severe impact on the pregnancy and increase the chance for premature labor
(Warkulwiz et al., 2017). Despite the fact that women are more likely to have UTIs than men, males can also
get them, but the likelihood is considerably lower: just 14% of men and 53% of women will get a UTI in their
lifetime, respectively (Warkulwiz et al., 2017). There are not many studies on UTIs in the poor world; the
literature on the topic is scarce; and there are not enough statistics to demonstrate how serious the problem is
(Warkulwiz et al., 2017).
The most prevalent bacteria associated with UTIs include:
1. Escherichia coli (E. coli): According to the National Kidney Foundation, this bacterium is responsible
for 80 to 90% of UTIs. E. coli resides in your stomach innocuously most of the time. However, it might be
problematic if it gets into your urinary system, which typically happens when stool migrates into the urethra.
2. Klebsiella pneumoniae: K. pneumoniae has emerged as a major cause of healthcare associated

opportunistic infections, such as bacteraemia, pneumonia, and UTIs (Effah et al.,
2020). They can be especially problematic for patients with underlying health conditions (Bobbadi et al.,
2023).
3. Enterococcus species: Enterococci, such as Enterococcus faecalis and Enterococcus faecium, are
responsible for a smaller percentage of UTIs but can still pose a threat, especially in healthcare-associated
UTIs (Govindarajan and Kandaswamy, 2022).
4. Staphylococcus saprophyticus: This Staphylococcus species is a less frequent cause of UTIs,

primarily affecting young women. It can adhere to the urinary tract walls and cause infections (Mancuso et
al., 2023).
5. Proteus mirabilis: Proteus mirabilis is a significant nosocomial UTI cause. This species' capacity to
produce virulence factors like biofilms, adhesion molecules, urease, proteases, siderophores, and toxins is
linked to its pathogenicity (Danilo de Oliveira et al., 2021).
6. Pseudomonas aeruginosa: Within the hospital setting, P. aeruginosa is the third most common cause
of urinary tract infection (7−10%) after E. coli and E. faecalis (Khalil et al., 2022). Patients with underlying
conditions, such as urinary tract abnormalities or indwelling urinary catheters, are more susceptible to UTI
caused by P. aeruginosa (Mancuso et al., 2023). The inherent multiple antibiotic resistance of this
microorganism, combined with its ability to develop new resistance to multiple classes of antibiotics and to
form biofilms, explains the high mortality and morbidity of UTIs caused by P. aeruginosa (Mancuso et al.,
2023).
7. Methicillin-resistant Staphylococcus aureus (MRSA): This particular strain of Staphylococcus aureus

can cause infections, including urinary tract infections (UTIs), primarily in ill people and can arise anywhere
that healthcare is provided (Brown et al., 2021). Since MRSA are resistant to a certain class of antibiotics
(penicillins), which are frequently used to treat S. aureus infections, treatment options for MRSA infections
can occasionally be limited. This indicates that penicillins have no effect on the bacteria and that the patient
is not expected to benefit from treatment with this class of antibiotics (Brown et al., 2021).
UTIs are mainly caused by Gram-negative bacteria that are becoming an increasing threat to public health
because of their ability to acquire genes located on transferable plasmids that code for extended-spectrum β-
lactamases (ESBLs) (Bedenic and Mestrovic, 2021). These enzymes are capable of hydrolysing third-
generation cephalosporins and monobactams but not carbapenems (Mancuso et al., 2021). In addition, ESBLs
pose a public health problem because they are encoded on plasmids that usually carry other resistance genes
against different classes of antibiotics (e.g., aminoglycosides, sulphonamides, and quinolones) (Mancuso et
al., 2023). As a result, bacteria that acquire these plasmids become multidrug resistant. Numerous antibiotic
prescriptions are written due to UTIs, which is recognized to be a significant factor in the spread of
antimicrobial resistance (Li et al., 2023) (Biondo 2023). Therefore, developing new medications to tackle
antimicrobial resistance and broadening the scope of research to discover new therapeutic choices have taken
the lead. Söderström et al. recently published a paper in which they for the first time explained how
uropathogenic Escherichia coli (UPEC) bacteria proliferate and spread (Soderstrom et al., 2022). The authors
discovered that during the infection cycle of UTIs, UPECs develop spaghetti-like strands that measure several
hundred times their original length before reverting to the rod shape (Soderstrom et al., 2022). They made this
discovery using a human bladder cell infection model. This study has prepared the path for the identification
of new therapeutic alternatives for the treatment of UTIs brought on by UPECs, even though more research is
required to determine why the bacteria carry out this metamorphosis. This is crucial given that uropathogenic
E. coli is responsible for the majority of UTIs (80%) (Shah et al., 2019). Although numerous uropathogens'
toxins and proteases have been investigated as prospective vaccine targets for UTI prevention, more research
is required to assess the effectiveness of these vaccines (Mancuso et al., 2023). Numerous siderophore
systems have been researched as potential vaccine targets since uropathogens need an iron supply to colonize.
According to this research, siderophore vaccinations can lessen the number of bacteria that mice's bladders
become colonized with during an illness, making them useful antigens to consider in the future (Mancuso et
al., 2023). There are currently no effective vaccines against urinary tract infections, despite the fact that a
number of vaccines have been researched for the prevention of these illnesses.
A recent study reported the use of a novel method. In order to boost the recruitment of bacterial elimination
cells and avoid further infections, the scientists delivered the vaccine along with an adjuvant directly into the
bladder (Wu et al., 2021). The most promising new antibiotics for the treatment of UTIs are gepotidacin and
the two oral carbapenems, tebipenem and sulopenem. Tebipenem and sulopenem are in various stages of
clinical development for the treatment of complicated and uncomplicated UTIs, while gepotidacin, a member
of the pyranopyridine class, selectively inhibits bacterial DNA replication (Scangarella-Oman et al., 2022,
Veeraraghavan et al., 2021).
2.2Antimicrobial properties of honey and ginger

Antibiotics used to treat bacterial infections must change due to bacteria's enduring antibiotic resistance
(Younas et al., 2019). Additionally, antibiotics have a variety of adverse effects on the gut flora of the host,
such as immunosuppression and hypersensitivity (Lin, 2021). For the treatment of bacterial strains that are
resistant to antibiotics, the pharmaceutical industry has created the most recent generations of antibiotics (Lin,
2021). The first reports of antibiotic-resistant bacteria date back to Brazil in the 1980s, and they are still
prevalent today. Natural bioactive antimicrobial compounds have demonstrated to be a good alternative for
the treatment of bacterial infections with very few limits despite the fact that numerous techniques, including
modified pharmaceuticals, have been created up to this point (Buckova et al., 2018). The herbal medicines
have greater antimicrobial activity due to the presence of different bioactive chemicals like allicin,
flavonoids, terpenoids, tannins, alkaloids, etc. (Fayyaz et al., 2019; Mahmood et al., 2019; Abbas et al.,
2019). Different plant materials have been reported to possess immunostimulant (Mahmood et al., 2014),
antiviral (Aslam et al., 2014), antibacterial (Arshad et al., 2017; Yasmin et al., 2020), and antiprotozoal
activity (Abbas et al., 2017; Zhang et al., 2020).
In order to maintain human health, the World Health Organization (WHO) emphasizes the use of medicinal
plants as vital medications. In developing nations, medicinal plants are used as common treatments by about
80% of the population. Therefore, research should be done on these plants to learn more about their
characteristics, safety, and effectiveness (Lin, 2021). Numerous medicinal plants have had their
phytochemical compositions examined in order to identify their bioactive components. These bioactive
compounds, which include tannins and phenols, are used therapeutically to stop pathogenic bacteria from
growing (Cui et al., 2016). The top objective will be to find innovative and powerful therapeutic alternatives.
Long-lasting acute infections are caused by multidrug-resistant bacteria, which makes antibiotic therapy for
pathogen control ineffective (Lin, 2021). Recent research on medicinal plants and their therapeutic
applications is very beneficial in addressing the problem of bacterial antibiotic resistance. According to
reports, multi-drug-resistant bacteria are mostly linked to epidemics of human diseases. The adoption of
herbal medicine as a substitute is prompted by the emergence of antibiotic resistance (Lin, 2021).
1. Cranberry Products: Cranberries have been utilized as a therapy for urinary tract disorders for decades, and
their antibacterial properties have long been known (Das, 2020). More than 80% of it is water, 10% is made
up of carbohydrates (glucose, mannose, and fructose), and the remaining 2% is made up of ascorbic, benzoic,
and glucuronic acids. Other phytoconstituents include anthocyanins, flavonoids, terpenoids, catechins, and
organic acids (citric, malic, and quinic acid, among others). Hippuric acid, an antibacterial agent that can also
acidify the urine, has been hypothesized to be excreted in considerable levels in urine by quinic acid (Das,
2020). Additionally, a new window into the mechanism of action of cranberries as an anti-adhesive,
preventative, and therapeutic agent has been opened by the definition of the UTI etiology (Das, 2020).
2. Probiotics: According to Meštrović Popovič et al., probiotics are living bacteria that, when given in
sufficient doses, alter the microbiota mix and benefit the host by possibly having immunomodulatory and
metabolic effects as well as normalizing the makeup of the gut microbiota. Due to the probiotics' ability to
compete with pathogenic bacteria in the stomach, significant UTI-causing organisms like E. coli are less
likely to proliferate, which may reduce the chance of developing UTIs or possibly prevent them altogether
(Meštrović Popovič et al., 2022).
3. D-Mannose: D-mannose is a monosaccharide that occurs naturally in a variety of plants, fruits, and berries,
including cranberries. It is also understood that the body produces it from glucose in order to produce
glycoproteins (Ala-Jaakkola et al., 2022). D-mannose is frequently sold as a dietary supplement to support
the health of the urinary system. According to research, free D-mannose in urine may saturate E. coli FimH
structures and therefore prevent E. coli from adhering to cells in the urinary tract (Ala-Jaakkola et al., 2022).
One of the probable methods for avoiding the development of UTIs is this so-called competitive inhibition
(Scaglione et al., 2021).
4. Lemon grass (Cymbopogon citratus): another study, an ethanolic extract of Cymbopogon citratus was
tested using the disc diffusion method against isolates, and the minimal inhibitory concentration was also
established. High-profile liquid chromatography (HPLC) was used to evaluate the phytochemical
composition of C. citratus ethanol extract ((Lin et al., 2021).). Zone of inhibition (ZOI) was used to measure
the antibacterial susceptibility of various bacteria, and it was shown that E. coli, S. aureus, P. aeruginosa, and
K. pneumoniae had average ZOI values of 14.0, 13.0, 13.0, and 8mm against ethanolic extract, respectively.
In the ethanolic extract of C. citratus, HPLC revealed the presence of flavonoids and phenolic compounds. C.
citratus also significantly reduced the number of uropathogens in the mouse model.
5. Essential Oils: The secondary metabolism of aromatic plants produces essential oils, also referred to as
volatile oils. According to a review of several essential oils, they may have biological features like anti-
inflammatory, sedative, digestive, antibacterial, antiviral, or antioxidant activity (Loose et al., 2020).
Essential oils have been utilized for bactericidal, virucidal, fungicidal, parasitical, insecticidal, therapeutic,
and aesthetic purposes since the Middle Ages. These applications are still prevalent today, particularly in the
pharmaceutical, sanitary, cosmetic, agricultural, and food industries (Loose et al., 2020). Essential oils have
been shown to have direct killing and sensitizing effects on microorganisms in recent years (Loose et al.
2020). Even multi-drug resistant strains of uropathogenic E. coli were susceptible to the antibacterial actions
of essential oils from the genus Origanum L. (Zapién-Chavarra et al., 2019, Zakaria Nabti et al., 2020). In the
treatment of the acute phase of UTI, recent research, by Wagenlehner et al. in patients with lower
uncomplicated UTI demonstrated that non-antibiotic herbal therapy was non-inferior to antibiotic therapy.
Additionally, it has been demonstrated that the essential oils of Pelargonium graveolens and Coriandrum
sativum enhance the efficacy of ciprofloxacin and gentamycin against particular uropathogens, respectively
(Loose et al., 2020).
6. Honey: A potent antibacterial agent with many applications is honey. The presence of sugar, phenolic
chemicals, hydrogen peroxide, 1,2-dicarbonyl compounds, and bee defensin-1 all contribute to honey's
antibacterial effectiveness (Almasaudi 2021). Depending on the source of the nectar, the type of bee, and the
storage, each of these components is present in varying amounts. These elements cooperate to make honey
effective against a range of pathogens, including multidrug-resistant bacteria, and to control their
susceptibility to antimicrobials. The type of honey produced, which is dependent upon its botanical origin, the
health of the bee, its origin, and its processing procedure, determines its effectiveness and potency against
pathogens (Almasaudi 2021).
According to Nao Medical, manuka honey is a particular variety of honey made in New Zealand from the
manuka tree's nectar. The Maori people have used it for generations as a medicine, and recent research has
revealed that it has potent antibacterial qualities. Methylglyoxal (MGO), a substance found in manuka honey,
has been proven to be effective against a variety of bacteria, including those that cause UTIs. It functions by
rupturing the bacterial cell membrane, which causes the bacterium to perish (Nao Medical 2023).
Manuka honey has anti-inflammatory qualities in addition to antibacterial ones, which can help to lessen the
pain and discomfort brought on by UTIs (Nao Medical, 2023). People can consume manuka honey or apply it
directly to the affected area. Manuka honey can be applied topically to the urethra or taken internally three
times a day for UTI. It is significant to remember that several types of manuka honey exist. Only honey with
a high MGO content (at least 400) has been demonstrated to be effective against germs, while the level of
MGO in honey can vary (Nao Medical 2023).
In a recent study, artificial honey with varying amounts of MGO and 3-phenyllactic acid (3-PLA) was used,
and it was shown that 3-PLA in concentrations above 500 mg/kg enhances the bacteriostatic effect of the
model honeys containing 250 mg/kg MGO or more (Thierig et al., 2023). It has been shown that the effect
correlates with the contents of 3-PLA and polyphenols in commercial manuka honey samples. Additionally,
yet unknown substances further enhance the antibacterial effect of MGO in manuka honey (Thierig et al.,
2023).
7. Ginger: Gull et al. investigated the antimicrobial activity of ginger extract (100 mg/ml to 0.01 mg/ml)
against Salmonella typhi, Shigella spp., Bacillus subtilis, Pseudomonas aeruginosa, Klebsiella pneumoniae,
and Staphylococcus aureus. For each kind of bacteria, a different minimum inhibitory concentration (MIC)
was required. Additionally, ginger has been said to be useful in treating a number of illnesses, including
infectious infections. To ascertain ginger's MIC and minimum bactericidal concentration (MBC),
antimicrobial susceptibility testing against a variety of microorganisms was conducted (Felicia et al.2022).
According to an in vitro investigation, ginger can be either bacteriostatic or bactericidal against intestinal
Escherichia coli depending on the concentration used (Felicia et al., 2022).
In an experimental study, Felicia et al. used the agar dilution method to determine the minimum inhibitory
concentration (MIC) and minimum bactericidal concentration (MBC). Six different ginger concentrations,
including 2,000 mg/ml, 1,000 mg/ml, 500 mg/ml, 250 mg/ml, 125 mg/ml, and 62.5 mg/ml, were used to
inoculate the McFarland 0.5 solution of UPEC on agar. The lowest concentrations, the MIC and MBC, were
recorded as being free of growth. bacteria on agar did not appear to be growing at concentrations of 1,000 or
2,000 mg/ml. As a result, the MIC and MBC values for UPEC were 1,000 mg/ml. They discovered that
ginger ethanol extract has antibacterial action that is effective against UPEC. The MIC and MBC for UPEC
in this investigation were 1,000 mg/ml (Felicia et al. 2022). There is limited research on the synergistic
antibacterial effect of honey and ginger on UTI-causing bacteria. Many natural remedies for UTIs lack
conclusive scientific evidence, and more rigorous clinical trials are needed to establish their safety and
efficacy. While some natural remedies show promise, more research is required to fully understand their
mechanisms of action and to develop evidence-based recommendations for their use in managing UTIs.
CHAPTER 3
3.0 MATERIALS AND METHODS
3.1 Materials
Petri dishes, Loops, Cooler box, Test tubes, Filter papers, Raw Ginger, Natural Malawian honey, Motor &
pestle, Antibiotics (Ciproflaxicin, sxt-trimethoprim, vancomycin & amoxicillin), Nutrient agar, Mueller
Hinton agar, Normal saline, Stirring rod
3.2 Study Design

This was a cross-sectional experimental laboratory study. It involved conducting a series of controlled
experiments to investigate and evaluate the antimicrobial properties of the mixture of ginger and honey
against UTI-causing bacteria (Yang et al., 2021). The study focused on systematic testing and analysis of
bacterial isolates in a controlled laboratory setting to determine the efficacy of these natural substances as
antimicrobial agents.
3.3 Study Setting

The study was conducted across two distinct settings. The initial phase involved sample collection, which
took place at Queen Elizabeth Central Hospital (QECH) in Blantyre. Bacterial isolates were collected after
getting approval from the Hospital Director of QECH. Following this, the laboratory testing and
experimentation phase occurred at Malawi University of Science and Technology (MUST) in Thyolo, where
additional samples were collected. The university's well-equipped laboratory facilities were utilized for
preparing ginger extract, honey samples, and conducting antimicrobial testing against the isolated UTI-
causing bacteria. As part of ethics considerations, privacy and confidentiality of the study samples were
protected. The full confidentiality of the people associated with the samples were maintained at all times.
This adherence to moral standards demonstrated our commitment to maintaining the participants' rights and
dignity as well as safeguarding any private information involved. We strived to perform a responsible and
ethical research project that respects the rights and well-being of everyone engaged by preserving the privacy
and confidentiality of our samples. Further to respecting ethical issues arising in the study, we managed to
seek approval from the Research Ethics Committees of the Malawi University of Science and Technology
(MUSTREC) and the sample collection was also approved by Research Ethics Office of Queen Elizabeth
Central Hospital
Figure 1: Map showing the study area

3.4 Sampling Techniques
This research project employed specific sampling techniques tailored to its objectives. Bacterial isolates were
obtained through purposive sampling (Thomas 2022) from QECH in Blantyre. The selection of isolates
included UTI-causing bacteria; K. pneumoniae, E. coli MRSA and E. faecalis were collected from queen
elizabeth central hospital. 4 samples were taken per every bacterial isolate that is a total of 16 samples were
used.
However, samples of E. coli, MRSA and Enterococcus species were collected from MUST which were used
for preliminary pilot experiments before collection of samples from QUECH.
1 2
Figure 2: Bacterial isolate samples
The figure above shows the frozen bacterial isolates from Malawi University of Science & Technology (1) and
QUECH (2).
3.5 Preparation of Growth Media
3.5.1 Nutrient Agar (NA)

Nutrient Agar (NA) is a general-purpose culture medium that is used to grow a wide variety of bacteria. It is
commonly used for the routine culture of non-fastidious microorganisms that grow and thrive without
specific nutritional or environmental conditions. It is a rich medium that contains all the essential nutrients
that bacteria need to grow, including peptone, beef extract, and agar (Tantray et al., 2023).
In the experimental procedure, 28 grams of NA powder was weighed out, and it was subsequently added to 1
liter of distilled water. The mixture was thoroughly mixed until complete dissolution of the powder was
achieved. The resulting medium was then heated to boiling state in order to dissolve the agar completely.
Following this, the medium had undergone autoclaving at 121°C for a duration of 15 minutes to ensure
sterilization. Once the medium was cooled down to a temperature range of 45-50°C, it was poured into 16
sterile Petri dishes. The medium was left to solidify completely before it could be utilized for culturing.
3.5.2 Mueller-Hinton Agar (MH)

Mueller Hinton Agar (MHA) is a standardized culture medium that is used for antimicrobial susceptibility
testing (AST). It is important to use a standardized medium for AST to ensure that the results are accurate and
reproducible (EUCAST 2023).
In the experimental procedure, 60 grams of MHA powder was weighed out, and it was subsequently added to
1 litre of distilled water. The mixture was thoroughly mixed until complete dissolution of the powder is
achieved. The resulting medium was then heated to boiling state in order to dissolve the agar completely.
Following this, the medium had undergone autoclaving at 121°C for a duration of 15 minutes to ensure
sterilization. Once the medium was cooled down to a temperature range of 45-50°C, it was poured into 16
sterile Petri dishes. The medium was left to solidify completely before it could be utilized for culturing.
Image showing the cooling stage of
culture media during the preparation of
nutrient agar and Mueller-Hinton agar
Figure 3: Media preparation
3.6 Sample Processing and Culture

The pure isolates of each bacterium were sub cultured on nutrient agar Petri-dishes using sterile loops for
resuscitation through the streak plate method for uniform multiplication before the commencement of the
antibacterial sensitivity testing (Allen et al., 2023). Four NA plates were used for each of the 4 bacterial
isolates. The plates were incubated for 24 hours.
Figure 4: Sub-cultured bacterial isolates
Image showing growth of bacteria colonies (K. pneumoniae, E. coli, Enterococcus faecalis and MRSA) on
nutrient agar for resuscitation after 24 hours of incubation at 37 degree Celsius in an incubator.
3.7 Processing of Natural Remedy Mixture
3.7.1 Honey
When I was analysisng the data I removed the part of having different concentrations to make the data easy
for analysis. Remove concentrations in all extracts below
The study was to use honey purchased from local farmers within the study area. The honey was bought at
Goliati, Thyolo market and Mulanje. In order to create different concentrations of honey, including 75%,
50%, and 25%, the honey was diluted with normal distributed. The volume of each extract concentration was
to be made in 10 ml. Using the C1V1=C2V2 formula, the dilutions were made by mixing matching volumes of
normal saline with the honey. Each concentration's aqueous extracts were to be kept in their own glass vials
with tight-fitting closures and labels, which were then to sterilized in an autoclave at 121°C for 15 minutes at
12 Psi (Saxena et al.,2020). After that the honey samples were ready for use in antimicrobial susceptibility
testing.
3.7.2 Ginger extracts

The ginger rhizomes were bought at the same locations the honey was bought. The ginger rhizomes were
cleaned with sterile distilled water and given an hour to dry naturally. The exterior coatings were manually
peeled off and rinsed once more. The ginger was crushed using a motor and pestle. A sterile cloth was used to
filter the ginger liquid after it had been crushed. The juice was regarded as the extract's purest form. To create
varying concentrations, the extract was diluted with normal saline, in varying concentrations of 75%, 50%,
and 25% respectively. The volume of each extract concentration was made in 10 ml. Using the C 1V1=C2V2
formula, the crude extract was diluted with equivalent amounts of normal saline to create the dilutions. Each
concentration's aqueous extracts were kept in their own glass vials with tight-fitting closures and labels,
sterilized in the autoclave at 121 °C for 15 minutes at 12 psi, and then were put into each glass vial.
3.7.3 Combination of honey and ginger extracts

The honey-ginger combo was made using only pure honey and ginger extracts. A 50:50 ratio was used to
prepare the combination. In a test tube, 10 ml of honey extract and 10 ml of ginger extract were combined to
create a honey-ginger stock. Then, using a stirring rod, the honey and ginger extracts were blended together.
Four different honey-ginger combinations were created from the honey-ginger stock: 100%, 75%, 50%, and
25%. 10 ml of the stock was added to a glass vial to create the 100% dilution.
Figure 5: Preparation of honey and ginger

Figure 6: Preparation of diffusion discs for AST
3.8 Antimicrobial Susceptibility Testing

Antimicrobial susceptibility testing (AST) is a vital process used to determine the effectiveness of antibiotics
against specific bacteria. This testing helps doctors identify which antibiotics will effectively treat an
infection and which ones might be resistant. By exposing bacteria to different antibiotics and observing their
reactions, healthcare professionals can make informed decisions about the most appropriate and targeted
treatment. Antimicrobial susceptibility provides an in vitro measure of bacterial response to an antimicrobial
agent that predicts therapeutic efficacy. Standard AST methods require isolated organisms and effectively
take a day to perform. Thus, a minimum of two days is required to obtain susceptibility information from a
clinical sample.
AST methods based on broth or agar dilution are used to determine the minimal inhibitory concentration
(MIC), defined as the lowest concentration of antimicrobial inhibiting growth in vitro. Organizations such as
the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial
Susceptibility Testing (EUCAST) correlate MICs with pharmacokinetic/pharmacodynamics (PK/PD) studies
and clinical outcome to derive categorical interpretive criteria (sensitive, resistant, etc.) upon which
antimicrobial therapeutic selections are made (Brennan-Krohn et al., 2017).
Pure bacterial isolates were obtained from the NA agar then a suspension of the bacterial isolate was prepared
to match a specific turbidity standard, often equivalent to a 0.5 of Macfarland solution.
McFarland standard, which represents a bacterial concentration of approximately 1-2 x 10^8 colony-forming
units per ml (CFU/mL). This standardization is crucial to ensure consistent results. A sterile cotton swab was
dipped into the bacterial suspension and used to evenly spread the bacteria across the surface of the Mueller-
Hinton agar plate. Next, disks impregnated with specific concentrations of honey and ginger combination
were placed on the surface of the agar plate. These disks were made by cutting out circles, with a diameter of
6mm, from filter papers. The disks were dipped in varying concentrations of honey, ginger and the mixture of
both. The disks were left to air dry for 24 hours before use. The plates were then incubated at 37°C for 16-18
hours (Ewnetu et al., 2014).
After incubation, the plates were examined. Zones of inhibition, which are areas where bacterial growth is
inhibited around the antibiotic disks, were measured in ml using a ruler. The size of the inhibition zone was
inversely proportional to the susceptibility of the bacteria to the antibiotic: larger zones indicated greater
susceptibility.
Interpretation of results was based on established standards and guidelines, such as those provided by
organizations like the Clinical and Laboratory Standards Institute (CLSI) or the European Committee on
Antimicrobial Susceptibility Testing (EUCAST).
CHAPTER 4
4.0 RESEARCH FINDINGS
Here we only present data that has been analysed. You can attach these reseults at the end if you want. Here
include tables and graphs that have gone through analysis
4.1 Control antibiotics

Table 1: shows Ciprofloxacin against Klebsiella Pneumoniae with EUCAST breakpoints interpretation
ISOLATE ZONE OF INHIBITION INTERPRETATION
1 29mm Sensitive
2 30mm Sensitive
3 31mm Sensitive
4 30mm Sensitive
Table 2: shows sxt-trimethoprim against Escherichia coli with EUCAST breakpoints interpretation
1 21mm Sensitive
2 20mm Sensitive
3 15mm Sensitive
4 28mm Sensitive
Table 3: shows vancomycin against Methicillin-resistant Staphylococcus Aureus with EUCAST breakpoints
interpretation
1 8mm Resistant
2 11mm Resistant
3 6mm Resistant
4 13mm Resistant
Table 4: shows amoxicillin against Enterococcus faecalis with EUCAST breakpoints interpretation
1 17mm Resistant
2 21mm Sensitive
3 20mm Sensitive
4 19mm Sensitive
Tables 1-4 show control drugs on all the four test isolates and the EUCAST breakpoint interpretation. Malawi
treatment guidelines was used to choose the antibiotics to which the bacterial isolates are most susceptible to
hence using Ciprofloxacin against Klebsiella Pneumoniae, sxt-trimethoprim against Escherichia coli,
vancomycin against MRSA and amoxicillin against Enterococcus faecalis. These antibiotics were to be used
as a positive control.
These tables are not necessary. Put analysed tables
4.2 Antimicrobial effects of honey, ginger and honey-ginger mixture

The honey, ginger and their combination exhibited a noteworthy antimicrobial effect against the test isolates.
This was confirmed by the culture plates below which revealed zones of inhibition for Klebsiella
Pneumoniae, Escherichia coli, Enterecoccus faecalis and MRSA at varying concentrations of each substance
as well as their mixtures. The measured results are documented in the accompanying tables.
Figure 7:
Honey
Ginger
Mixture of
honey and
ginger
Figure 7: Inhibition zones by honey and ginger on enterococcus spp.

Control plate
Ginger
Honey-ginger
mixture
Honey
Figure 8: Inhibition zones by hone and ginger on all bacterial species

Table 5: showing zones of inhibition by 100% honey, ginger and their mixture in their natural form against different bacterial
isolates
ORGANISM HONEY GINGER HONEY-GINGER

MIXTURE
K. pneumonia isolate 1 21mm 14mm 32mm
MRSA isolate 1 8mm 6mm 18mm
E. coli isolate 1 33mm 20mm 37mm
E. faecalis isolate 1 40mm 15mm 28mm
K. pneumoniae isolate 2 34mm 30mm 30mm
MRSA isolate 4 9mm 8mm
20mm

Table 6: showing zones of inhibition by 75% honey, ginger and their mixture against different bacterial isolates

MIXTURE
K. pneumonia isolate 9mm 12mm 20mm

1
E.coli isolate 1 12mm 13mm 8mm
E. FAECALIS isolate 40mm 11mm 31mm

1
K. pneumoniae isolate 29mm 20mm 17mm

2

2

3

3

4

4
Table 7: showing zones of inhibition by 50% honey, ginger and their mixture against different bacterial isolates

MIXTURE
E. FAECALIS isolate 1 6mm 6mm 6mm
K. pneumoniae isolate 2 6mm 6mm 6mm
Tables above show the zones of inhibition created around the filter discs impregnated with honey, ginger and
their combination in ranging concentration as shown in each respective table
The tables should not be included here
Average inhibition in mm for ginger extracts, honey, mixture of honey-ginger extracts and standard antibiotic
discs (Ciproflaxicin, Co-trimaxole, Vancomycin and Amoxicillin).
Figure 9: Figure 1: Average inhibition in mm for ginger powder water extracts, honey, mixture of honey-ginger
extracts and standard antibiotic discs (Ciproflaxicin, Co-trimaxole, Vancomycin and Amoxicillin).
Table 8: Results of one-way ANOVA and descriptive statistics of the inhibitions of honey, ginger extracts and honey –
ginger extracts mixtures on the bacteria isolates
___________________________________________________________________________
Remedy Test Organisms N Mean Standard Deviation Minimum Maximum
___________________________________________________________________________
Ginger K.pneumonae 4 24 8.42 14 31
MRSA 4 9.75 3.304 6 13
E-coli 4 18.25 3.862 13 22
E-faecalis 4 12.50 3.000 9 15
Total 16 64.5 18.586 42 81
___________________________________________________________________________
Honey K.pneumonae 4 33.50 8.963 21 42
MRSA 4 9.00 2.944 6 13
E-coli 4 30.50 12.61 12 39
E-faecalis 4 36.75 4.992 30 41
Total 16 109.75 29.509 69 135
___________________________________________________________________________
Mixture K.pneumonae 4 34.00 7.439 29 45
MRSA 4 20.00 1.414 18 21
E-coli 4 38.00 2.708 36 42
E-faecalis 4 27.75 9.570 19 41
Total 16 119.75 21.131 102 149
___________________________________________________________________________
Ciproflaxicin K.pneumonae 4 30.00 0.816 29 31
Vancomycin MRSA 4 9.50 3.09 6 13
Cofrimaxole E.coli 4 21.00 5.354 15 28
Amoxicilin E.faecalis 4 19.25 1.708 17 21
___________________________________________________________________________
*The mean difference is significant at the 0.05 level.
Table 9: Results of Turkeys Honestly Significant Difference (HSD) test of honey-ginger extract mixtures on the test
bacteria isolates
___________________________________________________________________________
Post Hoc Test (I) Remedy (J) Remedy Mean Difference (I-J) Std Error Sig
___________________________________________________________________________
K.pneumonae Mixture Ginger 10.000 5.012 0.243
Honey 0.500 5.012 1.000
Ciproflaxicin 4.000 5.012 0.854
MRSA Mixture Ginger 10.250 * 1.976 0.001

Honey 11.000* 1.976 < 0.001
Vancomycin 10.500* 1.976 < 0.001
E-coli Mixture Ginger 19.750* 5.122 0.011

Honey 7.500 5.122 0.487
Cofrimaxole 17.000* 5.122 0.027
E.faecalis Mixture Ginger 15.250* 4.007 0.012

Honey - 9.000 4.007 0.166
Amoxicillin 8.500 4.007 0.201
___________________________________________________________________________
*The mean difference is significant at the 0.05 level.
Make corrections on results…present results in form of the graph and two tables I
sent you. Remove the ones that contain raw data.
CHAPTER 5
Here I told you to go through the sample review paper that I sent you. Please let
that guide you. Under discussion note the following
Discussion is not literature review
Here you need to interpret your results. Say something about the p-values
Explain your results based on your objectives. Here you are answering your
objectives and research hypothesis…..take note of he newly proposed objectives
based on how we analysed the data. You have done a good job here but now we
need to discuss based on the objectives given. The change is due to some missing
items in the data that you collected
Compare and contrast all your findings with what others have found and reported.
Explain the diff. and simmi.
Explain why differences arose. Make informed explanations.
*******please read the paper I sent you. It will help you come up with a good
discussion.
5.0 DISCUSSIONS
5.1 Control antibiotics

The inhibition zones of antibiotics on bacterial cultures are commonly determined through a technique called
the Kirby-Bauer disk diffusion method. In this method, paper disks containing a known concentration of
antibiotics are placed on an agar plate inoculated with the bacteria of interest. The antibiotic diffuses from the
disk into the agar, creating a concentration gradient. If the bacteria are susceptible to the antibiotic, they will
not grow near the disk, creating a clear zone known as the inhibition zone. Looking at different inhibition
zones of Ciprofloxacin, Co-trimazole, Vancomycin, and Amoxicillin on four different bacterial strains of
interest, it was established that Klebsiella pneumoniae is susceptible to fluoroquinolones like ciprofloxacin.
The inhibition zone size had mean value of 30mm after doing four samples, compared to the EUCAST break
point of ≥21mm which reads sensitive, but the inhibition zone can vary depending on factors like the
concentration of the antibiotic and the strain's susceptibility profile. E. coli showed susceptibility to co-
trimazole, and the inhibition zone was significantly about 21mm compared to the EUCAST sensitive break
point of ≥15mm. Methicillin-resistant Staphylococcus aureus (MRSA) showed not susceptibility to
Vancomycin with mean value of 9.5mm on four samples, as compared to EUCAST break point of ≤13mm
which reads not sensitive. It typically showed a small inhibition zone. Enterococcus faecalis was susceptible
5.2 Honey, ginger and their mixture against UTI-causing bacteria
The research aimed to comprehensively assess the effectiveness of honey, ginger, and their combination
against bacterial isolates known to cause urinary tract infections (UTIs) in this case Escherichia coli,
Klebsiella Pneumoniae, Enterococcus faecalis, and MRSA. At a 100% concentration, honey displayed
notable antibacterial efficacy, generating substantial zones of inhibition against Klebsiella Pneumoniae
(42mm) and Enterococcus faecalis (40mm) and E. coli with total mean zone of inhibition being 27mm.
However, MRSA exhibited resistance to honey, evidenced by a limited zones of inhibition such as 9mm
which can be considered resistance in comparison to the used control antibiotic. For the honey, the mean
zones of 34mm (Klebsiella Pneumoniae), 31mm Escherichia coli and 37mm (Enterococcus faecalis) signify
substantial antibacterial activity in comparison to standard zones of inhibition by EUCAST. However, the
lower mean of 9mm against MRSA reflects its resistance to honey. Ginger, in contrast, demonstrated mixed
results, with resistance against MRSA and Enterococcus faecalis (13mm) but effectiveness against Klebsiella
Pneumoniae (31mm) and E. coli the most, though the results had a huge variation. Ginger exhibited a
moderate mean zone of 24mm against Klebsiella Pneumoniae, 18mm against Escherichia coli and 13mm
against Enterococcus faecalis but demonstrated resistance with a mean zone of 10mm against MRSA.
The combination of honey and ginger surpassed individual treatments, yielding mean zones of 34mm
(Klebsiella Pneumoniae), 20mm (MRSA), 38mm (E. coli), and 28mm (Enterococcus faecalis). The statistical
significance of these means is supported by minimal variability, as indicated by the low standard deviations,
reinforcing the reliability of the observed effects. The combination of honey and ginger produced synergistic
effects, particularly excelling against MRSA (20mm) which was the most resistant to honey and ginger as
single formulations, emphasizing the potential of this combination as an enhanced antibacterial strategy to
even AMR. Overall, the combination of honey and ginger had a mean inhibitory zone of 30mm beating the
27mm and 16mm of honey and ginger respectively.
The observed resistance of MRSA to honey and ginger individually, but susceptibility to their combination,
underscores the potential for synergistic effects. This aligns with the initial hypothesis and supports the
overall conclusion that the combination of honey and ginger at a 100% concentration exhibits enhanced
antibacterial efficacy against UTI-causing bacterial isolates compared to individual treatments. However, the
concentrations at 50% and 75% did not produce the maximum required inhibition zones, indicating potential
concentration-dependent effects hence results drawn from these concentrations are not analyzed or discussed
further. This limitation prompts further exploration into optimal concentrations and potential dose-response
relationships.
The factors responsible for the high or low susceptibility of the test organisms to ginger are not exactly
known but may be attributed the secondary metabolite (inhibins) and phytochemicals (gingerol and shagelol,
flavonods) (Stewart et al., 1991). The results obtained in this study agree with multiple previous studies such
as that of Ewnetu et al., (2014), Ficker et al., (2003), Grange and Davey (1990) and Zahra et al., (2009). The
100% natural honey showed antibacterial activity on the bacterial isolates possibly due to redox potential of
ascorbic acids present in the honey (Rahmanian et al., 1970). The inability of the ginger, honey and their
combination to produce significant zones at less than 100% concentration also agree completely with those of
other authors where some honey samples tested were found not to have anti-bacterial activity at 50%, 60%
and 70% dilutions. The ability of honey to kill microorganisms has been attributed to its higher content of
tetracycline derivatives, peroxidases, fatty acids, phenols, ascorbic acids and amylases (Jeddar et al., 1985;
Nzeako and Hamdi, 2020).
The antimicrobial substances in the ginger and honey samples were not estimated in this study, however, the
fact remains that the samples tested all showed a high degree of activity on the tested microbes. In a nut shell,
the research findings present compelling evidence of the synergistic antibacterial effects of honey and ginger
against UTI-causing bacteria. While individual treatments showed efficacy, the combination outperformed
them, especially against the resistant MRSA strain. The findings contribute to the evolving understanding of
natural compounds as alternative therapeutic options against antibiotic-resistant bacterial strains.
CHAPTER 6
6.0 CONCLUSIONS AND RECOMMENDATIONS
6.1 Conclusions
Antimicrobial susceptibility testing was carried out using disc diffusion method on Muller-hinton agar in
order to assess the sensitivity of bacterial isolates: Klebsiella Pneumoniae, Enterococcus faecalis, Escherichia
coli and MRSA to honey and ginger as single formulated compounds and also as a mixture. Accordingly,
honey is a valuable food not only for its nutritious value, but also for its physical, chemical, and biological
capabilities. These bioactive functions are mostly due to the biochemical compounds found in honey, which
may be used alone as a functional food or as potential additives in a variety of foods so as ginger. Generally,
Honey demonstrated a higher efficiency as a bacteriostatic agent while ginger demonstrated a significant
antibacterial activity but less than the activity of honey, however honey and ginger as a mixture had a higher
antibacterial activity as compared to honey and ginger as separate entities and in other circumstances
produced greater inhibition zones as compared to the zones produced by clinically used antibiotics.
Not to be included in the conclusion. Summarise ur finding here. You have

made a good conclusion in the abstract. Read conclusions from other studies
and learn from there.
However, there were also limitations to the study such as the use of a small sample size whereby the results
generate from the study cannot be generalized but also the study was done in vitro hence not defining the real
pharmacological efficacy and interaction of the antibacterial agents in vivo. Lastly, the study did not explore
potential adverse effects or toxicity associated with prolonged use of honey and ginger. Future research could
address these gaps and provide a more comprehensive understanding of the therapeutic potential and safety
profile of honey and ginger combinations for combating UTIs.
6.2 Recommendations
The findings of our research reveal the remarkable potential of honey and ginger in combating bacterial
infections caused by Escherichia coli, Klebsiella pneumoniae, MRSA, and Enterococcus faecalis.
Specifically, our investigations demonstrate that the combined use of honey and ginger exhibits a more
pronounced inhibitory effect on these bacterial strains compared to when honey and ginger are administered
separately. This synergy underscores the therapeutic value of utilizing these natural substances in tandem for
the treatment of bacterial infections.
Despite the promising outcomes observed in our study, it is imperative to emphasize that individuals
experiencing persistent symptoms following the administration of honey and ginger remedies should
promptly seek medical attention from a qualified healthcare provider. While these natural remedies offer
notable antibacterial properties, they may not always suffice to address severe or prolonged infections
necessitating professional intervention. Furthermore, our research underscores the importance of considering
regional variations in the medicinal application of honey and ginger. Given the potential variability in honey
quality, particularly in regions with limited regulatory oversight, further investigations are warranted to assess
the bioavailability of antibacterial agents present in honey post-consumption. This research endeavor will
facilitate a deeper understanding of the therapeutic efficacy of honey and ginger preparations and inform the
development of evidence-based treatment protocols for bacterial infections. In a nut shell, our research
underscores the therapeutic potential of honey and ginger in combating bacterial infections, however
continued exploration and scrutiny are essential to optimize their efficacy and ensure their safe and effective
utilization in clinical practice.
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APPENDICES
Appendix 1: Calculations for different concentrations of honey and ginger

Different concentrations of honey, ginger and their combination were achieved through the dilution of the
original remedies with distilled water.
100% Honey, Ginger and their combination
100% Honey was the original natural honey (undiluted) which had a concentration of 2.6g/ml
Where 1.5ml of honey was used with a mass of 4.1968g-0.3050g (mass of measuring pipette)
Thus (4.1968-0.3050) ÷ 1.5ml = 2.6g/ml
100% ginger followed a similar procedure where the original concentration of the extracts was calculated to
be (2.1456-0.3050)g ÷ 1.5ml =1.2g/ml
100% Combination of honey and ginger was made by combining honey and ginger extracts in their natural
form where (3.0933-0.3050)g ÷1.5ml = 1.9g/ml
To achieve a lower estimated concentration of 75% of honey, ginger and their combination,3ml extra
distilled water was added to the 100% concentrations as shown below:
75% Honey 3.8918g ÷3ml =1.3g/ml
75% Ginger 1.8406g ÷3ml = 0.6g/ml
75% Combination of honey and ginger 2.7883g ÷ 3ml =0.9g/ml
Note that the masses of honey, ginger and their combination do not include the mass of the weighing pipette
which at this point was estimated at 0.305g

To make a lower dilution; an estimate of 50% initial concentration 10ml of distilled water was added to the
initial concentrations of the honey, ginger and their combination to give the following lower concentrations:
50% Honey: 3.8918g÷10ml =0.4g/ml
50% Ginger: 1.8406g ÷10ml = 0.2g/ml
50% Combination of honey and ginger: 2.7883g ÷10ml = 0.3g/ml
Appendix 2: Certificates of approval from MUSTREC and QUECH Research Office
Figure 10: Approval of research by QUECH

Figure 11: MUSTREC approval of research
Appendix 3: Growth media details

1. Nutrient Agar
a. Ingredients and their uses:
i. Agar: A solidifying agent to give the medium a gel-like consistency.

ii. Peptone: Provides nitrogen and other nutrients for bacterial growth.
iii. Beef extract or yeast extract: Adds vitamins and minerals necessary for microbial growth.
iv. NaCl (Salt): Maintains osmotic balance.
v. Water: Constitutes the solvent for dissolving the dry ingredients.
b. Preparation procedure: 38g of blood agar base was suspended in 1000ml of distilled water, agitated
and brought to boil to dissolve completely. The media was then sterilized by autoclaving at 121˚C (15psi)
for 15 minutes after which it was allowed to cool down then poured into 20 petri dishes.
Figure 12: Nutrient agar
2. Mueller-Hinton Agar
a. Ingredients and their use:
i. Beef extract and casein hydrolysate: Provide nitrogen, vitamins, carbon, amino acids,
sulphur, and other essential nutrients for bacterial growth.
ii. Starch: Acts as a colloid and is added to absorb any toxic metabolites produced by bacteria.
It also yields dextrose when hydrolysed which serves as a source of energy.
iii. Agar: Serves as the solidifying agent.
b. Preparation procedure: 19.76g of Mueller Hinton powder was added to 520 ml of distilled water. The
solution was brought to a boil in a heating mantle so as to dissolve the medium completely. The medium
was then sterilized by autoclaving at 121˚C for 15 minutes after which it was allowed to cool down before
pouring into 20 petri dishes (25ml in each).
Figure 13: Mueller-Hinton agar
Appendix 4: Some of the equipment used

1. Precision balance
2. Autoclave
3. Centrifuge
4. Incubator
5. Refrigerator
6. Water bath
7. Light microscope

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MALAWI UNIVERSITY OF SCIENCE AND TECHNOLOGY

ACADEMY OF MEDICAL SCIENCES

DEPARTMENT OF BIOLOGICAL SCIENCES

UNDERGRADUATE RESEARCH DISSERTATION

ASSESSING THE EFFECTIVENESS OF ANTIBACTERIAL ACTIVITY OF HONEY AND

ANTHONY NKHUNGUNI (MMB/026/19)

SOLIUM NGULUWE (MMB/044/16)

CHAWEZI CHIRWA (MMB/014/19)

SUBMITTED TO UNDERGRADUATE’S OFFICE

IN PARTIAL FULFILMENT FOR THE REQUIREMENTS FOR

THE AWARD OF BACHELOR OF SCIENCE DEGREE IN MEDICAL MICROBIOLOGY

Anthony Nkhunguni 4th February 2024

Chawezi Chirwa 4th February 2024

Solium Nguluwe 4th February 2024

Dr. Willard Mbewe 1st March 2024

Key words: Escherichia coli, Klebsiella Pneumoniae, Methicillin-resistant Staphylococcus

1.1. Bacteria Species and Diversity................................................................................................2

1.1.2 Urinary tract infections (UTI) in humans..........................................................................3

1.1.3 Synthetic antibiotics against UTIs in clinical use.............................................................3

1.2 Exploring natural antibacterial remedies..................................................................................4

1.3 problem statement....................................................................................................................5

1.4.1 Main Objective..................................................................................................................6

1.4.2 Specific Objectives............................................................................................................7

1.5 Hypothesis of the Study...........................................................................................................7

1.5.1 Main Hypothesis...............................................................................................................7

1.5.2 Specific Hypotheses..........................................................................................................7

1.6 Significance of the study..........................................................................................................7

2.0 LITERATURE REVIEW.............................................................................................................9

2.1 Bacterial species associated with UTI......................................................................................9

2.2Antimicrobial properties of honey and ginger........................................................................12

3.2 Study Design..........................................................................................................................18

3.3 Study Setting..........................................................................................................................18

3.4 Sampling Techniques.............................................................................................................19

3.5 Preparation of Growth Media.................................................................................................20

3.5.1 Nutrient Agar (NA).........................................................................................................20

3.5.2 Mueller-Hinton Agar (MH).............................................................................................21

3.6 Sample Processing and Culture..............................................................................................22

3.7 Processing of Natural Remedy Mixture.................................................................................23

3.7.2 Ginger extracts................................................................................................................23

3.7.3 Combination of honey and ginger extracts.....................................................................24

3.8 Antimicrobial Susceptibility Testing.....................................................................................25

4.0 RESEARCH FINDINGS...........................................................................................................27

4.1 Control antibiotics..................................................................................................................27

4.2 Antimicrobial effects of honey, ginger and honey-ginger mixture........................................29

5.1 Control antibiotics..................................................................................................................38

6.0 CONCLUSIONS AND RECOMMENDATIONS.....................................................................41

Appendix 1: Calculations for different concentrations of honey and ginger...............................50

Appendix 3: Growth media details...............................................................................................53

Appendix 4: Some of the equipment used...................................................................................55

Figure 1: Map showing the study area.............................................................................................19

Table 1: shows Ciprofloxacin against Klebsiella Pneumoniae with EUCAST breakpoints

AST: Antimicrobial Susceptibility Testing

UTI: Urinary Tract Infection(s)

MRSA: Methicillin-Resistant Staphylococcus Aureus

AMR: Antimicrobial resistance

EUCAST: European Committee of Antimicrobial Susceptibility Tests

1.1. Bacteria Species and Diversity

1.1.2 Urinary tract infections (UTI) in humans

1.1.3 Synthetic antibiotics against UTIs in clinical use

1.3 problem statement

1.4.1 Main Objective

1.4.2 Specific Objectives

1.5 Hypothesis of the Study