INTERNATIONAL JOURNAL OF RESEARCH AND SCIENTIFIC INNOVATION (IJRSI)  
ISSN No. 2321-2705 | DOI: 10.51244/IJRSI |Volume XII Issue XI November 2025  
Prevalence and Antibacterial Susceptibility Pattern of Escherichia  
Coli in Fermented Cow Milk Collected from Selected Markets in  
Maiduguri, Borno State.  
1Jarafu Umar Ngunupu, 2Nasiru Adoyiza Usman, 3Emeka Walter Ndubuisi, 4Wadai Smith, 5Isaac  
Geofrey Gwary, 6Apeh Enoch Odekina, 7Uchechukwu Chisom Chimezie, 8Desmond Senguro  
1,2 Department of Pharmaceutical Microbiology, University of Maiduguri, Maiduguri  
3,4,5,7 Department of Pharmacology and Toxicology, University of Maiduguri, Maiduguri  
6Department of Pharmacology and Toxicology University of Igbinedion, okada.  
8Department Pharmaceutical Chemistry, University of Maiduguri, Maiduguri.  
DOI: https://dx.doi.org/10.51244/IJRSI.2025.12110003  
Received: 10 November 2025; Accepted: 20 November 2025; Published: 27 November 2025  
ABSTRACT  
Escherichia coli is an enteric food borne pathogen associated with life-threatening disease conditions. The  
enterobacteria are frequently found in the cattle gastrointestinal tract with a high potential of contaminated  
animal products such as milk, meat, and cheese. A cross-sectional study was conducted to investigate the  
presence of Escherichia coli. in fermented milk products sold within Maiduguri, Borno State. Thirty (30) samples  
were collected from different sources Bacteriological isolation and biochemical characterization yielded  
Escherichia coli and the prevalence of E. coli was 3.33 % (2/60). Antibacterial susceptibility test was carried out  
and the bacteria were sensitive to ciprofloxacin (17 mm) and gentamycin (21 mm), while the remaining  
antibiotics were non sensitive, i.e. cephalexin, cefoxitin and suphamethoxazole-trimethoprim. Based on  
EUCAST the isolates were resistant to all the antibiotics except gentamycin. The result revealed the presence of  
Escherichia coli in the fermented cow milk samples. The finding indicates possible faecal contamination of the  
milk product with serious public health consequences. This necessitates the need to screen other milk products  
in the area. Health authorities in the state need to enlighten dairy product farmers on the zoonotic potential of  
Escherichia coli.  
Key words: Escherichia coli, Fermented milk, Antimicrobial susceptibility, Foodborne pathogens, Public health  
.
INTRODUCTION  
Foods made from or containing the milk of mammals are referred to as dairy goods or milk products. Mammals  
like cattle, water buffaloes, goats, sheep, camels, and humans are the main source of them. Dairy goods include  
foods like cheese, butter, and yogurt. A dairy, or dairy factory, is a facility that manufactures dairy  
products.(Joseph F. Sullivan Center, 2020). Except for regions of central Africa and the majority of East and  
Southeast Asia, dairy products are consumed all over the world.  
Since milk contains significant macro- and micronutrients, it is a popular beverage that is vital to the diets of  
millions of people worldwide. Due to its composition, milk is known to be beneficial during children and  
adolescence; yet, its comparatively large concentration of saturated fat raises concerns about possible negative  
consequences, including on the cardiovascular system.(Visioli & Strata, 2014).  
Milk contains all of the necessary nutrients and several anabolic hormones because it is naturally used to feed  
and support the growth of newborn mammals. In order to meet nutritional needs for calcium and lower the risk  
of bone fractures, the recommended consumption level has been justified. However, there are worries about the  
potential negative health effects and the lack of evidence supporting the health benefits of consuming large  
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amounts of milk products. Consequently, a thorough evaluation of the contribution of dairy consumption to  
human nutrition and disease prevention is necessary (Haug et al., 2007). In addition to lactose, the primary milk  
sugar, milk also includes vital amino acids, lipids, vitamins, minerals, and proteins (caseins, whey proteins, and  
minor proteins). Thus, its use may have played a role in the centuries-long success of human evolution. But in  
addition to being extremely nourishing for people, milk is also a great place for microbes to develop (Fasce et  
al., 2020). A vast and diverse variety of bacteria may grow in milk because of its high nutritional content. As a  
result, in addition to its natural microbiota, a wide variety of additional microorganisms that come from the udder  
skin, teat canal, milking equipment, tanks, and storage containers all of which represent the farm and pasture  
environments may colonize the milk as soon as it is milked.  
Scientific classification of Escherichia coli:  
Domain:  
Phylum:  
Class:  
Bacteria  
Proteobacteria  
Gammaproteobacteria  
Enterobacteriales  
Enterobacteriaeceae  
Escherichia  
Order:  
Family:  
Genus:  
Species:  
E. coli  
Gram-negative, rod-shaped, flagellated, facultative anaerobic, and nonsporulating, Escherichia coli is a member  
of the enterobacteriaceae family of bacteria. Enterotoxigenic Escherichia coli (ETEC), attaching and effacing  
Escherichia coli (AEEC), enterophathogenic Escherichia coli (EPEC), enterohemorrhagic Escherichia coli  
(EHEC), and Shiga-toxin producing Escherichia coli (STEC or VTEC) are some of the classifications for this  
bacterium based on its virulence factor, (Holko et al., 2006; karch et al., 2005; Frohlicher et al., 2008; Solomakos  
et al., 2009; Wang et al., 2010;).  
The bacteria has been the subject of much research for more than 60 years and is simple and affordable to grow  
and cultivate in a lab setting. The majority of Escherichia coli's chemically defined mediums use carbon as an  
energy source, making them chemoheterophs (Tortora et al., 2010).  
Escherichia coli is a member of the Enterobacteriaceae, a broad family of gram-negative rods (Cheesebrough,  
2005). German pediatrician Theodor Escherichia initially identified this species in the faces of healthy people in  
1885. Because it is found in the colon, he named it bacterium coli commune. It was reclassified as Bacillus coli  
after bacterial taxonomy was revised (Migula, 1985).  
They naturally occur in warm-blooded creatures' lower gastrointestinal tracts (Singleton et al., 1999). Many of  
the more well-known diseases, including Salmonella, Yesinia pestis, Klebsiella, and Shigella, are members of  
the Enterobacteriaceae, a broad family of Gram-negative bacteria, along with numerous innocuous symbionts.  
Proteus, Enterobacter, Serratia, and Citrobacter are some of the other bacteria in this family that cause illness.  
In the phylum Proteobacteria, this family is the sole representative of the class Gammaproteobacteria within  
the order Enterobacteriales (Brenner et al., 2005). The innocuous strains, which are a typical component of the  
gut's flora, can help their hosts by generating vitamin K2 (Bentley et al., 1982) and preventing colonization of  
the intestine with pathogenic bacteria (Hudault et al., 2001). About 0.1% of the gut flora is made up of facultative  
anaerobes like Escherichia coli, (Russell et al., 2001) and the main way that harmful forms of the bacterium  
spread illness is by fecal oral transfer. Because cells have a short half-life outside the body, they can be used as  
indicator organisms to check environmental samples for fecal contamination (Feng et al., 2002).  
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Strains of Escherichia coli  
One of the most researched bacterial species, Escherichia coli (E. coli), is well-known for its wide range of  
genetic and phenotypic variations (Kaper et al., 2004; Croxen et al., 2013). It can be found in a variety of settings,  
including soil, water, and the intestines of animals, including people. While commensal strains aid in digestion  
and the production of vitamin K, pathogenic strains are linked to illnesses such intestinal infections, newborn  
meningitis, and urinary tract infections (Todar, 2020; Leimbach et al., 2013). Globally, foodborne outbreaks  
brought on by toxic E. coli have become more significant for public health (CDC, 2024). Diarrhea is a common  
sign of intestinal infections, and symptom profiles frequently reveal the type of pathogenic strain (WHO, 2023).  
Intestinal E. coli infections are typically categorized into five major pathotypes:  
Enterotoxigenic (E. coli, ETEC)  
Enteropathogenic (E. coli, EPEC)  
Enteroinvasive (E. coli, EIEC)  
Enterohemorrhagic (E. coli, EHEC)  
Enteroaggregative (E. coli, EAEC)  
(Kaper et al., 2004; Nataro & Kaper, 1998).  
Enterotoxigenic (ETEC) Strains  
In underdeveloped nations, ETEC is a leading cause of infantile and traveler's diarrhea (Qadri et al., 2005).  
Contaminated food or water is the means of transmission, and after repeated exposure, immunity is developed.  
Secretory diarrhea is caused by ETEC's expression of colonization factor antigens (CFAs), which mediate  
adhesion to intestinal epithelial cells and release heat-labile (LT) and heat-stable (ST) enterotoxins (Gaastra &  
Svennerholm, 1996).  
Enteropathogenic (EPEC) Strains  
In low-income nations, infantile diarrhea is still frequently caused by EPEC, which is transferred through  
contaminated water or fecal-oral pathways (Nataro & Kaper, 1998). Despite lacking LT and ST toxins, EPEC  
produces distinctive attaching-and-effacing (AE) lesions by forming localized microcolonies on intestinal  
epithelium (Dean et al., 2005). Watery diarrhea is brought on by these lesions, which interfere with intestinal  
absorption.  
Enteroinvasive (EIEC) Strains  
Similar to Shigella species, EIEC causes sickness akin to dysentery by entering intestinal epithelial cells,  
proliferating intracellularly, and damaging host cells (Lan et al., 2004). Children in developing nations are  
primarily affected by infections, while tainted food has been implicated in outbreaks in developed areas  
(Sansonetti, 2001).  
Enterohemorrhagic (EHEC) Strains  
One of the deadliest strains of E. coli is EHEC, which includes E. coli O157:H7. It is spread by contaminated  
vegetables, unpasteurized milk, and undercooked meat, and its infectious dose is minimal (CDC, 2024). Shiga  
toxins (Stx1, Stx2), which are produced by EHEC, prevent protein synthesis, kill epithelial cells, and cause  
hemolytic uremic syndrome (HUS) and bloody diarrhea (Tarr et al., 2005). Because E. coli O157:H7 cannot  
ferment sorbitol, it produces colorless colonies on sorbitol-MacConkey agar, allowing for quick laboratory  
screening (March & Ratnam, 1986).  
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Enteroaggregative (EAEC) Strains  
EAEC strains generate biofilms without causing noticeable lesions by adhering to intestinal mucosa in a stacked-  
brick pattern (Nataro et al., 1995). In children and immunocompromised patients, they are associated with  
chronic watery or mucoid diarrhea (Harrington et al., 2006).  
Uropathogenic (E. coli, UPEC) and verotoxin-producing (E. coli, VTEC) strains are two more noteworthy E.  
coli pathotypes that are important contributors to systemic and urinary tract infections (Johnson & Russo, 2005).  
Characteristics of Escherichia coli  
Escherichia coli is a facultatively anaerobic, Gram-negative rod that is a member of the Enterobacteriaceae  
family (Madigan et al., 2022). Usually found as solitary rods, cells are 1.1–1.5 μm wide and 2–6 μm long  
(Tortora et al., 2021). They exhibit both fermentative and respiratory metabolisms and are oxidase-negative but  
catalase-positive. Peritrichous (lateral) flagella allow many strains to move, although there are other non-motile  
varieties (Madigan et al., 2022).  
Furthermore, E. coli expresses proteinaceous appendages called fimbriae and pili, which facilitate adherence to  
host tissues and surfaces and aid in colonization and pathogenicity (Proft & Baker, 2009). E. coli is a pathogen  
of significant medicinal importance as well as a model organism in biotechnology due to these structural  
characteristics and metabolic adaptability (Leimbach et al., 2013).  
The cell wall of Escherichia coli contains strain-specific O lipopolysaccharide antigens (175 O antigens are now  
known), as well as flagella or H antigens, if they are present (56 H kinds are known). Additionally, there are  
eighty distinct capsular polysaccharide (K) antigens. Although only the O and H types are typically given, such  
as E. coli O157:H7, Escherichia coli are serotyped depending on the combination of O, H, and K antigens. E.  
Coli serotyping is a valuable epidemiological tool when combined with molecular and phage typing (Bat, 2014).  
Lactate, succinate, ethanol, acetate, and carbon dioxide are all products of mixed-acid fermentation, which  
Escherichia coli employs in anaerobic environments and can survive on a broad range of substrates. When  
Escherichia coli coexists alongside hydrogen-consuming species like methanogens or sulphate-reducing  
bacteria, the hydrogen levels must be low since several routes in mixed-acid fermentation produce hydrogen gas.  
(Madigan et al., 2006).  
Escherichia coli grows best around 37 °C (98.6 °F), while certain lab strains can grow at temperatures as high  
as 49 °C (120 °F). Any medium that contains glucose, ammonium phosphate, monobasic, sodium chloride,  
magnesium sulfate, potassium phosphate, dibasic, and water is suitable for Escherichia coli growth, as is  
lysogeny broth. Using a wide range of redox pairs, including as the oxidation of pyruvic acid, formic acid,  
hydrogen, and amino acids, and the reduction of substrates like oxygen, nitrate, fumarate, dimethylsulfoxide,  
and trimethylamine N-oxide, growth can be fueled by either aerobic or anaerobic respiration (Fortadar et al.,  
2005).  
Isolation of Escherichia coli  
Numerous clinical specimens, such as urine, pus, feces, cerebral fluid (especially in newborns), endocervical  
swabs, high vaginal swabs, and blood cultures, can be used to isolate Escherichia coli (Cheesbrough, 2010;  
Forbes et al., 2022). The presence of E. coli in sterile bodily locations like blood, cerebrospinal fluid, or urine  
usually signifies infection because it is a frequent commensal bacterium in the digestive system (WHO, 2023).  
To avoid contamination and preserve bacterial viability during isolation, aseptic sample collection and suitable  
transport media are essential (Collee et al., 1996). To differentiate E. coli from other Enterobacteriaceae,  
selective and differential media are used for fecal and enteric samples (Cappuccino & Sherman, 2018).  
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Identification of Escherichia coli  
Based on its biochemical and cultural traits, Escherichia coli is initially identified. Blood agar, MacConkey agar,  
and Eosin Methylene Blue (EMB) agar are the first media on which clinical specimens suspected of containing  
enteric Gram-negative rods are inoculated (Forbes et al., 2022).  
Lactose fermentation and acid generation cause E. coli to develop pink to crimson colonies on MacConkey agar,  
but non-lactose fermenters show up colorless (Madigan et al., 2022). Strong acid production causes the dyes  
eosin and methylene blue to precipitate, giving E. coli colonies on EMB agar a characteristic metallic green  
sheen (Tortora et al., 2021). In clinical microbiology labs, these differential reactions serve as important markers  
for presumptive identification.  
Biochemical identification further confirms the organism. Distinguishing features of E. coli include:  
1. Production of indole from tryptophan, detected by Kovac’s reagent (Cappuccino & Sherman, 2018).  
2. Positive lysine decarboxylase activity, differentiating E. coli from other coliforms (Forbes et al., 2022).  
3. Ability to utilize acetate as a sole carbon source, reflecting its metabolic versatility (Madigan et al., 2022).  
4. Motility due to peritrichous flagella, although some clinical strains may be non-motile (Tortora et al.,  
2021).  
Pathogenicity of Escherichia coli  
Viral strains of Escherichia coli can cause a variety of illnesses in both domestic animals and people. UTIs,  
newborn meningitis, and gastroenteritis are common human infection manifestations (Todar, 2007; Kaper et al.,  
2004). Furthermore, especially in immunocompromised people, E. coli is linked to more serious systemic  
infections like gram-negative pneumonia, septicemia, peritonitis, and mastitis (Forbes et al., 2022; Donnenberg,  
2015).  
Because they produce strong toxins or have virulence characteristics that improve adhesion, invasion, or immune  
evasion, certain strains of Escherichia coli are extremely harmful. Hemorrhagic colitis and hemolytic uremic  
syndrome (HUS), a dangerous illness marked by hemolytic anemia, thrombocytopenia, and renal failure, can be  
brought on by Shiga toxin-producing E. coli (STEC), which includes serotypes O157:H7, O104:H4, O26, O103,  
O111, O121, O145, and O104:H21 (Kaper et al., 2004; Tarr et al., 2005; WHO, 2023).  
The E. coli O157:H7 strain is a prominent example; it caused the June 2011 outbreak in Europe, which resulted  
in hundreds of illnesses and multiple deaths from complications mediated by Shiga toxin (Frank et al., 2011).  
The outbreak brought attention to the dangers that contaminated E. coli poses to public health and emphasized  
the significance of food safety protocols and surveillance (WHO, 2023).  
The acquisition of virulence genes through horizontal gene transfer is a major factor in the pathogenic potential  
of E. coli strains. This allows the bacteria to colonize a variety of host niches and cause illnesses ranging from  
mild diarrhea to infections that can be fatal (Nataro & Kaper, 1998; Todar, 2007).  
Medical importance of Escherichia coli  
In medicine, probiotics such as the non-pathogenic Escherichia coli strain Mutaflor and the E. coli O83:K24:H31  
strain Colinfant are used mostly to treat inflammatory bowel disease and other gastroenterological conditions  
(Grozdanov et al., 2004). In contemporary industrial microbiology and biological engineering, Escherichia coli  
is crucial (Lee, 1996). Biotechnology was founded on the work of Stanley Norman Cohen and Herbert Boyes in  
Escherichia coli, who used plasmids and restriction enzymes to produce recombinant DNA (Russo, 2003). The  
production of human insulin by manipulating Escherichia coli was one of the earliest practical uses of  
recombinant DNA technology (Cornelis, 2000). In microbiology research, Escherichia coli is also employed as  
a model organism. Using Escherichia coli as a model bacterium, Joshua Lederberg and Edward Tatum initially  
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identified the phenomenon known as bacterial conjugation in 1946 (Lederberg et al., 1946). It continues to be  
the major paradigm for conjugation research.  
Disease by Escherichia coli  
Escherichia coli is a rod-shaped, flagellated, gram-negative bacterium. Even though it is a vital part of the  
bacterial gut flora, illnesses can be brought on by intestinal bacteria spreading to other organs (cystitis,  
pneumonia) or by direct ingestion of pathogenic Escherichia coli subtypes (contaminated food, for example).  
For example, enterohemorrhagic Escherichia coli (EHEC) can cause hemolytic-uremic syndrome (HUS) and  
severe colitis, especially in young children and newborns.  
Genetic basis of Escherichia coli resistance to antibiotics  
The issue of antibiotic resistance is getting worse. The use of antibiotics as growth promoters in animal feeds is  
likely responsible for some of this, but human abuse of antibiotics is also a contributing factor (Johnson et al.,  
2006). The rate of adaptative mutations in Escherichia coli is "on the order of 10 −5 per genome per generation,  
which is 1,000 times as high as previous estimates," according to a study published in the Journal Science in  
August 2007. This finding may be important for the investigation and treatment of bacterial antibiotic resistance  
(Perfeito et al., 2007). Through a mechanism known as horizontal gene transfer, antibiotic-resistant Escherichia  
coli can also transfer the genes that cause antibiotic resistance to other bacterial species, including  
Staphylococcus aureus. Escherichia coli frequently possess several drug-resistant plasmids, which they can  
easily spread to other species when they are under stress. Plasmids from and to other bacteria can be accepted  
and transferred by Escherichia coli due to species mixing in the intestines. Accordingly, Escherichia coli and  
other Enterobacteria are significant sources of antibiotic resistance that can be transferred (Salyers et al., 2004).  
Since types of bacteria that manufacture extended-spectrum beta-lactamases have become more prevalent in  
recent decades, resistance to beta-lactam antibiotics has become a specific concern. Many, if not all, of the  
cephalosporins and penicillin’s are rendered ineffective as treatments by these beta-lactamase enzymes (Paterson  
et al., 2005).  
Genetic Origin of Drug Resistance  
(i) Chromosomal Resistance  
Spontaneous mutations at chromosomal regions that control bacterial susceptibility to antimicrobial drugs give  
rise to chromosome-mediated resistance (Brooks et al., 2022; Forbes et al., 2022). In the presence of the  
antimicrobial medication, which inhibits susceptible bacterial populations and promotes the growth of resistant  
mutants, these mutations are chosen for (Prescott et al., 2020).  
The rpoB gene, which codes for the β-subunit of RNA polymerase, is mutated in spontaneous chromosomal  
mutations, which contribute significantly to resistance to several medications, including rifampicin, even if they  
are not the most common source of clinical drug resistance (Davies & Davies, 2010). Furthermore, bacteria may  
become resistant to β-lactam antibiotics like cephalosporins and penicillin’s due to chromosomal abnormalities  
that cause the loss or modification of penicillin-binding proteins (PBPs) (Leclercq & Courvalin, 2002).  
(ii) Extrachromosomal Resistance  
(a) Plasmid-Mediated Resistance  
Extra-chromosomal genetic elements called plasmids, which frequently contain genes encoding antimicrobial  
resistance, are involved in plasmid-mediated resistance (Davies & Davies, 2010). In clinical practice, plasmid-  
mediated resistance is more prevalent than chromosomal resistance (Prescott et al., 2020). Multi-drug resistance  
(MDR) is a condition whereby resistance plasmids, or R factors, carry numerous genes that give resistance to  
multiple medications at the same time (Todar, 2020).  
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For instance, genes conferring resistance to tetracyclines, sulfonamides, streptomycin, and chloramphenicol may  
be found on a single plasmid (Levin et al., 2017). Enzymes like β-lactamases, which hydrolyze penicillins and  
cephalosporins' β-lactam ring and eliminate their antibacterial properties, can be encoded via plasmid-borne  
genes (Forbes et al., 2022).  
(b) Transposon-Mediated Resistance  
The use of transposons, or mobile genetic elements, is another extrachromosomal resistance strategy. These  
DNA segments have the ability to transfer resistance genes between bacteria by moving between plasmid and  
chromosomal DNA (Frost et al., 2005). Thus, resistance features may spread when the genetic material causing  
antimicrobial resistance in a donor cell is transferred to a susceptible recipient cell (Brooks et al., 2022).  
The intercellular transfer of resistance genes occurs through several mechanisms:  
Conjugation: The direct, frequently plasmid-mediated transfer of genetic material between bacterial cells  
through physical contact. Antimicrobial resistance spreads primarily through this route (Levin et al.,  
2017).  
Transduction: The process by which bacteriophages (phages) move bacterial DNA from one cell to  
another. The transfer of resistance genes, like those encoding β-lactamase, may be mediated via this  
pathway (Todar, 2020).  
Transformation: A spontaneous process wherein a bacterial cell integrates donor genetic material,  
possibly including resistance determinants, by directly absorbing free DNA from its surroundings  
(Prescott et al., 2020).  
Non-Genetic Origin of Drug Resistance  
(i) Metabolic Inactivity  
Due to metabolic inactivity rather than genetic changes, several microbes display resistance to antimicrobial  
medications (Brooks et al., 2022). Since many antibacterial medications only work on cells that are actively  
dividing, latent or non-replicating cells develop temporary resistance (Levin et al., 2017). For instance,  
Mycobacterium TB "persister cells" can endure in host tissues for years without proliferating. Antimicrobials  
have no effect on them while they are in this dormant state, but once their metabolic activity resumes, they  
become susceptible again (Zhang et al., 2012).  
(ii) Loss of Specific Target Structures  
The loss or modification of target structures that the antimicrobial drug works upon can also result in microbial  
resistance (Prescott et al., 2020). For example, certain susceptible bacteria may change into cell wall-deficient  
forms (L-forms) during penicillin therapy, making them resistant to inhibitors of cell wall synthesis such  
cephalosporins and penicillin’s (Brooks et al., 2022). Susceptibility to these antibiotics is regained after the  
bacteria return to their typical shape.  
Antibiotic used against Escherichia coli  
Bacterial infections are frequently treated with antibiotics or antimicrobial medicines. Penicillin’s (such as  
ampicillin and amoxicillin), cephalosporins (such as cefuroxime, cefotaxime, and ceftriaxone), aminoglycosides  
(such as streptomycin and gentamycin), fluoroquinolones (such as ciprofloxacin, levofloxacin, and norfloxacin),  
nitrofurantoin, and cotrimoxazole are among the classes of antibiotics frequently used to treat infections brought  
on by Escherichia coli.  
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Penicillin  
A class of bactericidal antibiotics known as penicillin's prevents the production of cell walls. Certain penicillin’s  
have a limited range of action (i.e. Penicillin’s work well against gram-positive bacteria like staphylococci and  
streptococci. Additionally, they exhibit efficacy against gram-negative bacteria, including salmonella, Shigella,  
hemophilia, protease, and pseudomonas (Prescott, 2005).  
Cephalosporin  
Similar to penicillin’s, cephalosporins are a class of antibiotics. Additionally, during the formation of  
peptidoglycans, cephalosporin suppresses the transpeptidase process. They are extensively categorized into  
generations and have a wide range of characteristics. Cefazolin, cephalexin, and cefadroxil are examples of first-  
generation cephalosporins that work better against gram-positive bacteria than gram-negative ones. Cefuroxime,  
cefprozil, and cefoxitin are examples of second-generation cephalosporins that exhibit enhanced effectiveness  
against gram-negative bacteria with some anaerobic coverage. With increased action against Enterobacteria and  
pseudomonas, the third generation is more powerful than the second (Prescott, 2005).  
Aminoglycosides  
Gram-negative rods are specifically targeted by aminoglycosides, which are bactericidal antibiotics. Some  
aminoglycosides are used to treat other organisms; for example, gentamicin is used in conjunction with penicillin  
G to treat enterococci, while streptomycin is used to treat tuberculosis. Aminoglycosides work through two key  
mechanisms: misreading of messenger RNA (mRNA) and suppression of the initiation complex. Gentamycin,  
Streptomycin, Amikacin, and Neomycin are among the aminoglycosides. When treating confirmed or suspected  
gram-negative infections, particularly those caused by P. aeruginosa, Enterobacter, Klebsiella, Serratia, and  
other species resistant to less toxic antibiotics, such as urinary tract infections, bacteremia, infected burns,  
osteomyelitis, pneumonia, peritonitis, and otitis, aminoglycosides are commonly used in conjunction with  
penicillin’s or cephalosporins (Prescott, 2005).  
Fluoroquinolones  
Nucleic acid synthesis is inhibited by these significant antibiotics. A growing number of infections are being  
treated with them. Topoisomerase II and bacterial DNA gyrase are inhibited by quinolones. They are antibiotics  
with a broad spectrum. Enteric bacteria like Klebsiella pneumoniae and Escherichia coli are effectively  
combatted by fluoroquinolones. They can be applied to gram-negative infections such as Haemophlilus and  
pseudomonas auruginosa. Staphylococcus aureus and Streptococcus pyrogenes are two gram-positive bacteria  
that they are also effective against. Fluoroquinolones include, for example, pefloxacin, ciprofloxacin,  
levofloxacin, norfloxacin, and nalidixic acid (Prescott, 2005).  
Chloramphenicol  
As a bacteriostatic antibiotic, chloramphenicol works by preventing the creation of new proteins. By blocking  
the bacterial ribosome's peptidyl transferase function, they stop proteins from elongating. A broad-spectrum  
antibiotic, chloramphenicol is effective in treating a variety of bacterial infections, particularly those caused by  
Neisseria meningitides, Streptococcus pneumoniae, and Hemophilus influenzae (Prescott, 2005).  
Nitrofurantoin  
Since 1953, nitrofurantoin has been used to treat urinary tract infections (UTI). Currently, it is used to treat  
simple UTIs and prevent UTIs in those who are at risk of getting them again. Interest in employing nitrofurantoin  
has grown as a result of growing bacterial antibiotic resistance to other widely used drugs, including  
fluoroquinolones and trimethoprim/-sulfamethoxazole (Garau, 2008). Nitrofurantoin is one of the first-line  
treatments for simple UTIs due to its effectiveness in treating the condition and the low rate of bacterial resistance  
to it (Gupta et al., 2011).  
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One of the few medications frequently used to treat UTIs during pregnancy is nitrofuratoin (Lee et al., 2008).  
The medication should not be administered to women in the latter stages of pregnancy because it may cause  
oxidative damage to the red blood cells and increase the risk of hemolytic anemia in the unborn child, who does  
not yet have the enzyme pathways required for glutathione metabolism. Neonatal jaundice was more likely to  
occur in babies whose mothers took this medication late in pregnancy (Nordeng et al., 2013).  
Cotrimoxazole  
Co-trimoxazole, also known as trimethoprim/sulfamethoxazole (TMP/SMX), is an antibiotic used to treat a  
range of bacterial and protozoal illnesses. It works as a folate antagonist by preventing the synthesis and  
metabolism of folate. Trimethoprim's effects result in a backlog of dihydrofolate (DHF), which can counteract  
the drug's inhibitory effect on tetrahydrofolate biosynthesis. Sulfamethoxazole is used to get rid of the excess  
DHF by stopping its synthesis in the first place (Wormser et al., 1982).  
In treating bacterial infections, co-trimoxazole was said to be more successful than each of its constituents alone;  
however, this was later called into question. In many countries, its usage has been limited to relatively specific  
situations where its greater efficacy has been established because to its higher prevalence of adverse effects,  
particularly allergic responses (Brumfit et al., 1993).  
MATERIALS AND METHOD  
Culture media  
Peptone saline diluents (PSD)  
Nutrient agar  
MacConkey agar  
Eosin methylene blue (EMB)  
Moeller Hinton agar.  
Reagents used  
Indole, urease, citrate, Normal saline.  
Solvents and disinfectants used  
Dettol soap, Dettol liquid, distilled water.  
Antibiotics used  
The panel of five antimicrobial agents [Ciprofloxacin, Gentamycin, Cephalothin, Cefoxitin,  
Sulphametoxazole/Trimethoprim]—was strategically selected to represent the different mechanistic classes of  
antibiotics, including agents commonly used for the empirical treatment of gastrointestinal and diarrheal  
infections associated with E. coli in both veterinary and human public health sectors within the Maiduguri region.  
METHODOLOGY  
Collection and labelling of samples  
A total of thirty (60) Dairy product (nono) samples were collected from different vendors across Maiduguri  
metropolis. These were labelled accordingly.  
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Serial dilution  
A serial dilution is any dilution in which the concentration decreases by the same factor in each successive step.  
In this research work, peptone saline diluents (PSD) was prepared by autoclaving a mixture of 1g of peptone  
water and 8.5g of NaCl dissolved in 1L of distilled water. 45ml of the diluent was pippeted into each of thirty  
sterilized bottles each containing 5ml of the nono samples, mixed thoroughly to form homogenates 1 to 20 (of  
1:10 dilutions). Next, 5 ml of each of the homogenates was measured into another thirty sterile bottles each  
containing 45 ml of peptone saline diluent, mixed thoroughly to form the second set of homogenates (of 1:100  
dilutions).  
Media preparation  
Each medium was accurately weighted and dissolved in an appropriate quantity of distilled water, according to  
the manufacturer’s specification; heated on a hot plate till the agar powder melted, and the medium was sterilized  
in an autoclave at 121°C for 15 minutes,  
Plating  
On freshly prepared nutrient agars, the plating (inoculation) of the homogenates, both 1:10 and 1:100 was carried  
out by using a sterilized wire loop to pick drops of the homogenates and carefully streaking on the surfaces of  
the agar. The plates were labelled accordingly and incubated at 37◦C for 24 hours.  
Isolation of Escherichia coli  
On freshly prepared MacConkey agar plates, colonies from the nutrient agar plates were inoculated by using a  
sterilized wire loop. These were also labelled accordingly and incubated at 37°C for 24 hours. Then the growth  
on MacConkey agar which shows pinkish reddish colouration which signifies that gram negative organism is  
present, was then inoculated on freshly prepared EMB (eosin methylene blue) to identify Escherichia coli from  
the colonies present. Escherichia coli will give a greenish metallic sheen of the plate.  
Biochemical test  
Biochemical tests were carried out based on Gram reactions. Among the tests carried out were indole, citrate,  
and methyl red.  
Indole test  
Indole is a nitrogen-containing compound formed when the amino acid tryptophan is hydrolyzed by bacteria that  
have tryptophanase. The test was carried out by inoculating one loopful of each test isolate separately into pre-  
sterilized Bijou bottles containing 3 ml of tryptone water. These were incubated at 37°C for 48 h after which 0.5  
ml Kovac’s reagent was added. The set up was examined by shaking after 1min. A red colour at the interphase  
was indicative of indole production.  
Citrate test  
This test uses a media, in which sodium citrate is the only source of carbon and energy. The medium used was  
Simmon’s medium. The medium was prepared according to the manufacturer’s instruction into Bijou bottles  
and autoclaved at 120°C for 15 minutes. The bottles are allowed to solidify as slanting slopes. They were  
inoculated with cultures of the isolates and incubated for 24 h at 37°C. It is positive when it is blue and negative  
when it appears green.  
Methyl red  
The methyl red (MR) test detects the production of sufficient acid during the fermentation of glucose and some  
bacteria have the ability to utilize glucose and convert it to stable acid like lactic acid, acetate or formic acid as  
the end product. Inside a test tube the microorganism to be tested was inoculated inside the test tube containing  
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5ml distilled water and peptone water, after 24 hours when the organism grows inside the tube, 2-3 drop of  
methyl red was drop inside the tube and it form a reddish ring which indicate positive  
Antibiotic Susceptibility Test  
The antibiotic susceptibility of isolates to commonly used antibiotics was determined using modified Kirby-  
Bauer technique as modified by Clinical and laboratory standard institute, (CLSI, 2014). The cultures were  
standardized by transferring four to five similar colonies of overnight culture with sterile wire loop into separate  
test tubes containing 5ml normal saline. The turbidity was adjusted to match that of McFarland 0.5, sterile plates  
of Mueller-Hinton agar (MHA) were prepared according to the manufacturer’s instruction. Surface of dried agar  
media were inoculated with the standardized cultures. After letting the plates dry for five minutes at room  
temperature, forceps were used to equally put antibiotic sensitivity discs on the plates' dried surface. The plates  
were incubated for eighteen hours at 37°C in order to stop the development of bacteria while the antibiotics  
diffused. The CLSI (2014) guideline was used to measure and interpret the zones of inhibition's sizes  
(Cheesbrough, 2005).  
The five different antibiotics used for the sensitivity testing include:  
1-Gentamycin (30µg)  
2-Ciprofloxacin (5µg)  
3-Cefoxitin (30µg)  
4-Cephalothin(30µg)  
5-Sulphamethoxazole/Trimethoprim(25µg)  
Determination of MAR (Multiple Antibiotic Resistance) Index  
The MAR Index of an isolate is defined as a/b, where 'a' represents the number of antibiotics to which the isolate  
was resistant and 'b' represents the number of antibiotics to which the isolate was subjected (Jayaraman et al.,  
2012). This was calculated for isolates that tested positive for Escherichia coli.  
RESULT, DISCUSSION AND CONCLUSION  
Characteristic appearance of the isolated Escherichia coli  
Out of the 60 nono samples dilutions plated on the nutrient agar plates all plates yielded bacterial growth. On  
inoculation onto MacConkey agar, a total of 21 plates yielded pinkish colony formation, suggesting the possible  
presence of Escherichia coli or other coliforms. The colonies were inoculated on freshly EMB (eosin methylene  
blue) on which 2 plate shows greenish sheen which indicate the present of Escherichia coli and the rest showed  
mucoid jelly pinkish and turbid growth which indicate the present of klebsiella spp.  
Microbiological analysis conducted on nono samples  
Below highlight the observations made from the isolation and biochemical tests conducted on the nono samples  
1
30  
1
30  
H1 -H1 and H2 -H2 . On nutrient agar which is a general purpose media, in all the sixty plate, they were  
distinctive colony growth on both the first and second homogenate. From there selected sample were inoculated  
on macconkey agar on which 21 plate shows pinkish reddish colouration. To further confirm e coli, the growth  
was inoculated on eosin methylene blue selective media to confirm e coli. Of which only two sample showed  
greenish metallic sheen which indicated that e coli is present, the reset of the plate showed pinkish mucoid jelly  
like, which indicate the presence of klebsiella ssp.  
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Table 1: Appearance of various media used in the isolation and confirmation of Escherichia coli  
NONO  
MACCONKE EOSIN  
BIOCHEMICAL TEST  
INDOLE CITRATE  
INFERENCE  
Y
METHYLENE  
SAMPLE  
S
BLUE  
METHYL RED  
H127  
PINK  
COLONIES  
GREENISH  
SHEEN  
PINK RING NO  
FORMATIO COLOR RING  
REDISH  
PRESENCE  
OF  
COLOURATIO  
N
N
CHANG FORMATIO ESCHERICHI  
E
N
A COLI  
H227B  
PINK  
COLONIES  
GREENISH  
SHEEN  
PINK RING NO  
FORMATIO COLOR RING  
REDISH  
PRESENCE  
OF  
COLOURATIO  
N
N
CHANG FORMATIO ESCHERICHI  
E
N
A COLI  
H226  
H130  
H13  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H23  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H126  
H117  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H117B  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H119  
H219  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
COLIFORMS  
N
E
N
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H16  
H26  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H222  
H224  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H124  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H125  
H225  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR  
NO RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H120  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
H128  
H129  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
N
E
N
COLIFORMS  
PINK  
COLONIES  
PINKISH  
MUCOID  
COLOUR  
NO  
RING COLOR NO  
RING PRESENCE  
FORMATIO CHANG FORMATIO OF  
COLIFORMS  
N
E
N
Result for antibiotic susceptibility test  
Table 2 shows the zones of inhibition (mm) by the various antibiotics used against the Escherichia coli A  
ANTIBIOTIC (WITH ITS CONCENTRATION)  
CIPROFLOXACIN (5 µg)  
ZONE OF INHIBITION (mm)  
17  
0
CEPHALOTIN (30 µg)  
GENTAMYCIN (30 µg)  
21  
0
SUPHAMETHOXAZOLE-TRIMETHOPRIM (25 µg)  
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CEFOXITIN (30 µg)  
0
As shown above, the isolate Escherichia coli A was resistant to three out of the five antibiotics used in test. The  
Multiple Antibiotic Resistance Index was determined using the formula, MARI = a/b Where, a is the number of  
antibiotics to which the isolate is resistant and b is the total number of antibiotics to which the test isolates have  
been evaluated for sensitivity. Therefore, the MARI of the Escherichia coli A is 0.6.  
DISCUSSION  
Only 2 out of a total of 60 nono samples tested positive for Escherichia coli. And the rest of the sample were  
positive to klebsiella. This poses danger or danger signal to the public. It is noteworthy that the isolated  
Escherichia coli was susceptible to two out of the five antibiotics used in this work. These include Ciprofloxacin,  
Gentamycin and. High sensitivity to ciprofloxacin and gentamycin have been recorded from previous studies  
conducted in Nigeria (Wariso et al.,2006) and India (Bharathi et al., 2002). The presence of coliforms indicated  
contamination and the poor level of hygiene after processing, Coliforms are not supposed to be present in nono  
because of high temperature short time pasteurization and effective cleaning and good hygienic procedures. The  
presence of coliforms from this pose’s great danger to the health of the consumers and suggest neglect on the  
part of the processors or the nono vendors. Coliforms are considered as normal flora of the intestinal tract of  
human and animals and their presence indicates direct faecal contamination. They have been used as indicator  
organisms for bacteriological quality of milk and its products (ICMSF). The level of presence of coliform and  
indicator organisms has been described as index of food hygiene. Klebsiella spp, has been related to bacterial  
pneumonia cases more severe than those produced by Streptococcus pneumonia and urinary tract infection.  
A key finding of our study is the high prevalence of resistance to sulphamethoxazole-trimethoprim (SXT) among  
the E. coli isolates from fermented cow milk. This pattern is particularly concerning given that SXT is an  
affordable and widely used first-line drug for treating various bacterial infections, including diarrheal diseases,  
in settings like Maiduguri. This widespread resistance, therefore, carries significant public health implications,  
suggesting that empirical SXT treatment for infections potentially originating from contaminated food sources,  
such as this locally fermented milk, may face high rates of treatment failure. From a mechanistic standpoint, this  
resistance is often attributable to the acquisition of mobile genetic elements, such as plasmid-mediated resistance  
genes (sul and dfrA) that encode for drug-resistant enzymes. Alternatively, while less characterized in  
environmental isolates, treatment survival in vivo could also involve phenotypic mechanisms like the formation  
of 'persister' cells—a transient, non-growing subpopulation of bacteria that can survive high antibiotic  
concentrations and later revert to a susceptible state. Further molecular studies are necessary to confirm the  
underlying genetic basis (plasmid vs. chromosomal) of the SXT resistance observed in this region.  
In most foods, the total bacterial count is often an indication for the sanitary quality, safety and utility of foods.  
It may reflect the conditions under which the product is manufactured such as contamination of raw materials  
and ingredients, the effectiveness of processing and the sanitary conditions of equipment and utensils at the  
processing plants  
CONCLUSION  
From the available result, it can be inferred from this study that there is a form of compromise in the good  
manufacturing practice of dairy products (particularly nono) being consumed in Maiduguri Metropolis.  
The efficacy of some commonly used antibiotics has been confirmed.  
RECOMMENDATION  
There is need to supervise the diary product, starting from the farm, how it was processed and transported  
to the local vendor.  
Vendors need to properly take good hygiene methods in order to prevent cross contaminations of the  
products.  
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The public should be educated on the danger of consuming dairy products from unreliable/ unhygienic  
sources.  
The vendors need to be educated on proper storage of the dairy product.  
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