Detection of Escherichia coli Resistance in Chicken Meat Against Various Types of Antibiotics at Tradisional Markets, Banyuwangi, Indonesia

Chicken meat has emerged as one of the most preferred protein sources globally, widely consumed due to its exceptional nutritional profile and palatability (Mir et al., 2017). Chicken meat is rich in essential vitamins and minerals, including B-complex vitamins, phosphorus, selenium, zinc, and iron, while maintaining relatively low-fat content and moderate caloric density (Liu et al., 2023). Despite these nutritional benefits, chicken meat exhibits heightened susceptibility to microbial contamination, which poses significant food safety risks. The high protein content, moisture levels, and neutral pH of chicken meat create an ideal environment for bacterial proliferation, especially when proper storage and handling protocols are not maintained (Silva et al., 2018).

The primary bacterial agents responsible for human intestinal infections from meat source include several bacteria such as Campylobacter spp., Staphylococcus spp., Escherichia coli, and Salmonella spp. (Uddin et al., 2021). The persistence and prevalence of these foodborne pathogens throughout the poultry processing chain, from slaughterhouses to retail markets, present substantial public health challenges. The risk of foodborne diseases is further amplified by cross-contamination occurring throughout various stages of food handling and preparation, particularly when temperature abuse takes place during storage and distribution (Akinsemolu et al., 2024; Hamaideh et al., 2024). Among the various foodborne pathogens, bacterial contamination represents the most serious threat to food safety in chicken meat. Bacterial pathogens are particularly dangerous because they can multiply rapidly under favourable conditions, produce toxins, and develop resistance to antimicrobial treatments (Salam et al., 2024). E. coli represents one of the most significant bacterial threats associated with chicken meat consumption. Although E. coli typically lives harmlessly in the intestines of both humans and animals some pathogenic strains can lead to serious illness in humans (Brătfelan et al., 2023). Research has documented E. coli contamination rates ranging from 44.26% to 100% in broiler chicken meat samples across various studies (Wibawati et al., 2023; Suswati et al., 2025). Virulence factors and toxins produced by pathogenic E. coli strains can lead to a wide spectrum of illnesses, starting with mild intestinal disturbances and potentially progressing to severe complications such as hemolytic uremic syndrome and hemorrhagic colitis (Whang et al., 2021). The organism’s ability to survive under diverse environmental conditions and to undergo horizontal gene transfer make it a particularly concerning foodborne pathogen (Vinayamohan et al., 2022).

The emergence of antibiotic-resistant E. coli in chicken meat has become an increasingly critical public health concern. Contaminated chicken meat serves as a reservoir for multidrug-resistant (MDR) E. coli strains that carry resistance genes against clinically important antibiotics (Suswati et al., 2025). Research has reported resistance levels greater than 60% against frequently used antibiotics such as erythromycin, trimethoprim, and ampicillin in E. coli isolates from chicken meat (Wibawati et al., 2023; Jaiswal et al., 2024; Wibisono et al., 2025). The detection of Extended Spectrum Beta-Lactamase (ESBL)-producing E. coli is especially concerning because these strains resist third-generation cephalosporins and other vital antimicrobials used in human healthcare (Wardhana et al., 2021). Studies have reported that the prevalence of ESBL-producing E. coli in chicken meat samples ranges from 14.84% to 100%. Frequently detected resistance genes include β-lactamase TEM (blaTEM), which confers resistance to penicillin and some cephalosporins; β-lactamase Cefotaximase-Munich (blaCTX-M), which is highly effective at hydrolyzing cefotaxime; and mobilized colistin resistance-1 (mcr-1), which modifies lipid A in the bacterial outer membrane and reduces affinity for colistin (Faridah et al., 2023; Lemlem et al., 2023; Handayani et al., 2024; Jaiswal et al., 2024).

Global monitoring studies have repeatedly shown a high occurrence of multidrug-resistant E. coli in chicken meat from various regions. Research from Peru documented that non-organic chicken isolates were more resistant to most antibiotics tested compared to human isolates, with E. coli strains containing clinically relevant resistance genes including mcr-1 for colistin resistance and blaCTX-M ESBLs (Murray et al., 2021). Studies from Saudi Arabia found 57% of E. coli isolates from chicken meat to be multidrug-resistant, with maximum resistance observed against penicillin G (95%) and amoxicillin (85%) (Ali and Chidrawar, 2021). In Bangladesh, research revealed 84.71% of multidrug resistance was observed in E. coli isolates from broiler meat, with high resistance rates recorded for tetracycline reach 86.78% and sulfamethoxazole-trimethoprim until 88.84% (Khanom et al., 2025). Indonesia, studies have documented E. coli prevalence rates of 44.26% in East Java broiler meat, with 59.26% of isolates showing multidrug resistance patterns. E. coli isolates exhibited resistance to ampicillin reach (59.3%), trimethoprim (61.1%), and erythromycin (66.7%) (Wibawati et al., 2023).

Traditional marketplaces present particularly high-risk environments for microbial contamination and growth due to suboptimal hygiene and sanitation practices. These markets often lack adequate refrigeration systems, proper waste management, and clean water supplies, creating conditions conducive to bacterial proliferation and cross-contamination (DeWaal et al., 2022; Gusti and Fitriyani, 2022). Studies have documented that traditional markets frequently fail to meet sanitation requirements, with vendors often not implementing proper personal hygiene behaviours and lacking access to clean water and appropriate waste disposal facilities (Jannah and Asyfiradayati, 2024). The absence of temperature control, combined with prolonged exposure to ambient conditions, increases the risk of bacterial multiplication and toxin production in chicken meat (Young et al., 2017). The physical infrastructure of traditional markets, including wooden surfaces that are difficult to sanitize and inadequate drainage systems, further contributes to the persistence and spread of pathogenic microorganisms (Ngan et al., 2020). The antimicrobial resistance profile of E. coli isolates from broiler chicken meat sold in Blambangan and Banyuwangi traditional markets has not been previously examined. This study aimed to determine whether chicken meat from these community-associated markets contains E. coli strains resistant to multiple antibiotics.

This study was conducted at the Faculty of Health, Medicine, and Life Science, Universitas Airlangga, Banyuwangi, East Java, Indonesia from August to October 2024.

An exploratory laboratory study was performed to isolate, identify, and determine the antimicrobial resistance profile of E. coli from broiler chicken meat. A quota sampling method was used, with inclusion criteria consisting of broiler chicken breast meat (Musculus pectoralis) sold in poultry meat sections, not adjacent to stalls selling other food products. Ten samples were collected from each market, resulting in a total of 20 samples.

A total of 20 samples of fresh broiler chicken meat were collected from two wet markets (Blambangan and Banyuwangi Traditional Market) in Banyuwangi Subdistrict (lat 8°13′; long 114°22′), Banyuwangi District, East Java, Indonesia during the dry season. Each sample weighing approximately 100 g was placed into a sterile polypropylene bag, labelled according to market and sample number, and transported to the laboratory under chilled conditions. Sterilization of laboratory equipment and materials was performed using an oven, autoclave, and UV light. Petri dishes, inoculating loops, and test tubes were sterilized in a hot air oven at 160 °C for 2 h, while plastic materials and cotton swabs were sterilized under UV light. Media used for bacterial isolation and identification were sterilized in an autoclave at 121 °C for 15 min.

Bacterial isolation was carried out by swabbing the surface of chicken meat and streaking it onto Eosin Methylene Blue Agar (EMBA) (Merck, Germany) using the zig-zag method. The plates were then incubated for 24 hours at 37°C. E. coli colonies displayed a metallic green sheen, resulting from lactose fermentation that increased medium acidity and precipitated methylene blue in the medium. Suspected positive samples of E. coli were further identified using Gram staining and IMViC biochemical tests, including Indole (I), Methyl Red (MR), Voges–Proskauer (VP), and Citrate (C) tests. Samples confirmed as E. coli showed positive results for Indole and MR tests (Merck, Germany), but negative results for VP (Merck, Germany) and Simmons Citrate Agar (SCA) (Merck, Germany). In Grams’ staining, E. coli colonies appeared pink, short rod-shaped, and non-spore forming (Khoiriyah et al., 2023). In the SIM test (Merck, Germany), E. coli exhibited positive growth indicated by bacterial proliferation beyond the stab line and the formation of a pink layer (ring) on the surface after the addition of Kovac’s or Ehrlich’s reagent. The MR test was positive, as indicated by the medium turning red, whereas the VP test was negative, showing no color change in the medium (Roy et al., 2023). The Citrate test also yielded a negative result, as the medium remained green and did not turn blue (Gita et al., 2021).

Antibiotic susceptibility was assessed using the Kirby–Bauer disc diffusion method on Mueller Hinton Agar (Oxoid, United Kingdom). Bacterial suspensions were adjusted to the 0.5 McFarland standard (approximately 1.5 × 108 CFU/ml) and evenly spread over the agar surface. After 5 min, antibiotic discs containing streptomycin 10 μg (Oxoid, United Kingdom), tetracycline 30 μg (Oxoid, United Kingdom), erythromycin 15 μg (Oxoid, United Kingdom), chloramphenicol 30 μg (Oxoid, United Kingdom), trimethoprim 5 μg (Oxoid, United Kingdom), aztreonam 10 μg (Oxoid, United Kingdom), and ciprofloxacin 5 μg (Oxoid, United Kingdom) were applied and plates were incubated at 37 °C for 24 hours, after which inhibition zones were measured using a caliper and interpreted based on the CLSI (2018) guidelines.

Primary data were obtained from bacterial isolation, identification, and antibiotic susceptibility testing. All results were recorded directly by the researchers. Data were tabulated and analysed descriptively to present the resistance profiles of E. coli isolated from broiler chicken meat.

Distinctive metallic green colonies with dark centers were produced by E. coli on Eosin Methylene Blue Agar (EMBA). Suspected E. coli colonies were purified by subculturing the metallic green colonies onto fresh EMBA plates. The isolation results are presented in Figure 1, with an estimated E. coli recovery rate of 85%.

Based on Grams’ staining, the colony was confirmed to be Gram-negative (Figure 1B). Identification of E. coli was performed using a series of IMViC biochemical tests with positive indole production test results (+), indicated by the formation of a red ring at the interface. Positive methyl red test results (+), indicated by a color change to red. Negative Voges-Proskauer test results (-), indicating no color change in the medium. Negative citrate utilization test results (-), indicated by no color change in the medium. Isolation and identification of E. coli yielded positive results in 9 of the 20 test samples (45%). These results are presented in Table 1.

Table 1: Isolation and identification results of Escherichia coli from chicken meat.

*A= Banyuwangi Market, B= Blambangan Market. EMBA: eosin methylene blue agar; IMViC: A series of tests carried out to idetrify the E.coli isolates.

Antimicrobial resistance testing revealed that E. coli isolates were 100% resistant to erythromycin (9/9), 66.7% resistant to streptomycin (6/9), 55.6% resistant to ciprofloxacin (5/9), 55.6% resistant to trimethoprim (5/9), and 33.3% resistant to tetracycline (3/9). All E. coli isolates demonstrated 100% susceptibility to chloramphenicol and aztreonam. The findings of the antimicrobial susceptibility assessment are displayed in Table 2, while Figure 2 displays the resistance patterns obtained using the Kirby-Bauer disc diffusion method on Mueller-Hinton Agar (MHA).

This study showed that 77.7% of the E. coli (7/9) isolates exhibited multidrug resistance (MDR). MDR results on E. coli isolated from meat broiler chickens can be seen in Table 3.

Table 2: Antimicrobial resistance test results of E. coli.

A = Banyuwangi Market, B = Blambangan Market, R = Resistant, I = Intermediate, S = Sensitive, ATM = aztreonam, E = erythromycin, C = chloramphenicol, CIP = ciprofloxacin, S = streptomycin, TE = tetracycline, W = trimethoprim.

Table 3: Multidrug resistant (MDR) E. coli in blambangan and banyuwangi traditional markets.

* A = Banyuwangi Market, B = Blambangan Market. MDR (Multidrug resistant): Results of E. coli resistance to ≥3 classes of antibiotics.

Escherichia coli was identified in 45% (9/20) of chicken meat samples from Blambangan and Banyuwangi traditional markets. This prevalence aligns with findings by Febrianti et al. (2022), who reported that chicken meat is highly susceptible to E. coli contamination. Soepranianondo et al. (2019) documented a contamination rate of 32.5% in East Java, while Amalia et al. (2020) found 16.66% in Makassar. International studies have reported varying contamination rates, including 83.33% in Bangladesh (Al-Salauddin et al., 2015), 70.4% in Japan (Odoi et al., 2021), 43.93% in Ethiopia (Hagos et al., 2021), 71.2% in China (Wang et al., 2021), and 98.3% in Turkey (Sahin, 2020).

Chicken meat sold in traditional markets often exhibits lower quality and hygiene standards compared to modern markets. Traders’ lack of knowledge regarding proper handling practices negatively affects meat safety. Contamination may originate from workers, equipment, waste, insects, rodents, and environmental factors such as air and water (Susilaningrum et al., 2022). Microbial contamination of food sold in markets poses a significant public health concern in developing countries largely due to inadequate hygiene practices and limited awareness among vendors (Gizaw, 2019).

Bacterial contamination in chicken meat can result from unhygienic handling, poor sanitation, open sales areas, stacking of meat without proper separation, storage without refrigeration, and prolonged exposure to open air, which favours bacterial growth (Wibisono et al., 2025). Survival of microbes in the air both within and outside droplets is influenced by environmental factors such as humidity and ambient temperature, influencing the potential for airborne transmission. Air circulation, including ventilation rates, strongly affects the removal and distribution of airborne pathogens indoors (Praja et al., 2021; Argyropoulos et al., 2022). Poor-quality water used in carcass washing may act as a source of microbial contamination, transferring bacteria including enteric pathogens onto meat surfaces and increasing the risk of foodborne illness (Ncoko et al., 2020). Poor water sanitation during carcass cleaning has been identified as an additional source of contamination (Santos et al., 2020).

Antibiotic susceptibility testing revealed that the E. coli isolates were completely sensitive to aztreonam and chloramphenicol (100%) but exhibited complete resistance to erythromycin (100%), followed by streptomycin (66.6%), ciprofloxacin (55.5%), trimethoprim (55.5%), and tetracycline (33.3%). Resistance to erythromycin has been widely reported, including 93.33% in Ghana, 78% in Sulawesi, and 100% in Lampung (Adzitey, 2018; Milawarni et al., 2022). Erythromycin, a macrolide antibiotic, has limited ability to penetrate the outer membrane of E. coli due to the lipopolysaccharide barrier (Cetuk et al., 2021; Susilo et al., 2022). The resistance mechanism involves inhibition of protein synthesis and genetic alterations in ribosomal protein-coding genes (Peykov et al., 2025).

Resistance to streptomycin was frequently detected with a rate of 53.7%, consistent with other regional studies reporting rates between 47% and 58% (Drugea et al., 2025). Streptomycin exhibits bactericidal activity by binding to the 30S ribosomal subunit, thereby inhibiting bacterial protein synthesis and disrupting peptide chain elongation (StatPearls, 2025). The inappropriate and excessive use of antibiotics, particularly their incorporation in animal feed for growth promotion and disease prevention, is a major contributing factor to the global rise of antimicrobial resistance, posing significant risks to public health (Martin et al., 2015).

Sensitivity to ciprofloxacin among clinical bacterial isolates ranged from 55% to 65%, reflecting a decline compared to earlier reports and emphasizing the persistent challenge posed by the increasing antimicrobial resistance among bacterial pathogens (Reis et al., 2016). Ciprofloxacin acts by inhibiting bacterial enzymes DNA gyrase and topoisomerase IV prevents DNA supercoiling and blocks bacterial replication. and transcription (StatPearls, 2025). Its extensive application has been linked to a higher likelihood of resistance development and reduced treatment effectiveness (Ariyani et al., 2018).

Trimethoprim resistance was found in 41% of E. coli blood culture isolates in a cohort study, with previous trimethoprim-resistant urinary tract infections identified as a significant risk factor for resistance, consistent with similar resistance rates reported in other regions (Balfour et al., 2022). Trimethoprim inhibits dihydrofolate reductase, but genetic mutations enable continued production of tetrahydrofolic acid, resulting in resistance (Bhosle et al., 2020).

Tetracycline resistance was observed in 33.3% of isolates, which was lower than the 87.2% reported by Wardhana et al. (2021). Excessive use of first-generation tetracyclines as growth promoters in livestock production creates selective pressure that drives the emergence of antimicrobial resistance mechanisms in bacterial populations (Blake et al., 2025). This antibiotic is bacteriostatic, acting by inhibiting protein synthesis through binding to the 30S ribosomal subunit (Simanjuntak et al., 2022).

Chloramphenicol demonstrated complete sensitivity in this study, showing no resistance among the E. coli isolates tested. This finding is consistent with other reports indicating chloramphenicol remains effective against E. coli despite resistance observed for many other antibiotics (Alobaidallah et al., 2024). Chloramphenicol is bacteriostatic and acts by binding to the 50S ribosomal subunit, specifically at the peptidyl transferase centre, thereby inhibiting peptide bond formation during protein synthesis (Paranos et al., 2022). Its efficacy is maintained when bacterial purine structures remain unchanged, facilitating antibiotic entry (Hernandez et al., 2024).

Aztreonam retained high activity against E. coli isolates with susceptibility rates exceeding 95% across most countries (Helio et al., 2021). Monobactams like aztreonam prevent bacterial cell division by binding to penicillin-binding protein 3 (PBP-3), which leads to inhibition of peptidoglycan synthesis and cell death. Their spectrum of activity is largely limited to aerobic Gram-negative organisms, providing an important alternative to aminoglycosides, particularly in cases of resistance or toxicity concerns (Dean et al., 2018).

Multidrug-resistant E. coli isolates were identified in 57.3% of samples from pig farms in Kupang city, Indonesia. The high prevalence is marked by resistance to multiple classes of antibiotics, raising serious concerns about the possible transmission of resistant bacteria between animals and humans (Kallau et al., 2018). Multidrug-resistant E. coli carriage on chicken farms was highly prevalent, with a studied prevalence of 62.7% among 158 sampled farms in Uganda (Nyolimati et al., 2025). Since 2017, the World Health Organization has classified E. coli is recognized as a critical pathogen due to its ability to cause a wide range of infections and its growing resistance to multiple antibiotics highlighting the urgent need for strict antibiotic regulation in humans, animals, and food production (Scheres and Kuszewski, 2019; Mancuso et al., 2021).

E. coli frequently develops antimicrobial resistance through multiple mechanisms such as production of β-lactamases that hydrolyse β-lactam antibiotics, mutations in dihydrofolate reductase (DHFR) and DNA gyrase that alter target sites, increased expression of efflux pumps reducing intracellular drug concentrations, and loss of purine biosynthesis genes that affect metabolic pathways related to susceptibility (Poirel et al., 2018). The production of β-lactamase enzymes is the primary mechanism underlying β-lactam antimicrobial resistance in E. coli, leading to hydrolysis of the antibiotic’s beta-lactam ring and inactivation. Extended-spectrum β-lactamase (ESBL)-producing E. coli strains pose significant challenges in clinical treatment due to their ability to hydrolyse a broad spectrum of β-lactams (González et al., 2025). Antimicrobial resistance arises from genetic mutations and the horizontal transfer of resistance genes between bacteria. Human activities such as inappropriate antibiotic consumption exert selective pressure, prompting the evolution and dissemination of resistant strains via mobile genetic elements (Habboush and Guzman, 2018). The mechanisms of action of antibiotics include inhibition of cell wall biosynthesis, alteration of membrane permeability, and interruption of DNA, RNA, and protein synthesis. By targeting these critical bacterial processes, antibiotics can effectively kill or inhibit bacterial growth. Different antibiotics specialize in one or more of these modes, which form the basis for their selective toxicity against bacteria (Uddin et al., 2021).

The prevalence of multidrug-resistant E. coli in poultry meat underscores the urgent need for enhanced hygiene and monitoring systems along the production chain to safeguard public health and ensure meat quality (Ahmed et al., 2025). Foodborne infection caused by E. coli can result in abdominal pain, diarrhea, vomiting, fever, and reduced appetite (Febrianti et al., 2022). Severe diarrhea may cause fluid and electrolyte loss, potentially leading to life-threatening complications (Alharbi et al., 2024).

Antibiotic-resistant E. coli infections, particularly to erythromycin, streptomycin, ciprofloxacin, tetracycline, and trimethoprim, increase the risk of treatment failure, secondary complications, and systemic spread (Mathew et al., 2019). Broad-spectrum antibiotic therapy may produce adverse effects and limit treatment options, especially in low-income countries with limited healthcare resources. Resistance also elevates treatment costs due to the need for expensive antibiotics and additional medical procedures.

Raising awareness of the dangers of antibiotic misuse is essential for sustainable behavioural change (Mathew et al., 2019). Improvement of food hygiene practices is critical in preventing foodborne diseases and limiting the spread of antibiotic-resistant bacteria within the food supply chain. Proper handling, storage, and preparation of food reduce microbial contamination and cross-contamination risks, which are critical factors in controlling antimicrobial resistance (Farrukh et al., 2025). Training traders in sanitation, hygiene, and food safety practices is essential for ensuring compliance with Good Manufacturing Practices (GMP) and Hazard Analysis Critical Control Points (HACCP) systems, thereby improving the overall safety and quality of food products (Lee et al., 2021). Prevention of antimicrobial resistance in E. coli can be effectively achieved by enforcing strict sanitation measures, implementing HACCP principles throughout the food production chain, launching public awareness campaigns on judicious antibiotic use, maintaining antimicrobial resistance surveillance systems, and fostering the development of vaccines or alternative therapeutic agents (Khan et al., 2024).

Escherichia coli has been effectively isolated and identified from chicken meat samples in Blambangan and Banyuwangi Traditional Market at a rate of 45% (9/20). The antimicrobial resistance profile of E. coli isolated from chicken meat in the Blambangan and Banyuwangi Traditional Markets showed 100% resistance to erythromycin, 66.6% resistance to streptomycin, 55.5% resistance to ciprofloxacin and trimethoprim, and 33.3% resistance to tetracycline, while remaining 100% sensitive to aztreonam and chloramphenicol. These findings highlighting the need for improved food hygiene and antimicrobial resistance monitoring in food supply chain.

The author would like to convey heartfelt thanks to Universitas Airlangga, especially the Veterinary Medicine Study Program, Faculty of Health, Medicine, and Life Sciences, for the exceptional support and facilities provided throughout the completion of this research

This study provides the first comprehensive evaluation of antimicrobial resistance profiles in broiler chicken meat from the Blambangan and Banyuwangi traditional markets, highlighting a concerning 77.7% prevalence of multidrug-resistant (MDR) Escherichia coli. While previous studies have broadly documented E. coli in poultry, this research addresses a critical local data gap by explicitly targeting community-associated wet markets, which present unique contamination risks due to suboptimal sanitation and hygiene infrastructure. Ultimately, these findings deliver crucial epidemiological evidence that underscores the urgent need for targeted food safety management interventions and supports broader One Health initiatives to monitor and mitigate foodborne antimicrobial resistance within developing urban food networks.

RNP and AY: Designed the study, materials preparation, data analysis, and manuscript preparation. AK, EKP, and RBH: Conducted the surveys and participated in laboratory examinations. DWS, AHS, SHAF, and FK: Contributed to the milk samples collection and field examination. AB, RNP: read, reviewed, and approved the final manuscript.

Ethical approval was not required for this study because no experimental treatment was administered to the animals.

The authors declare that no generative AI and AI assisted technology was used in the creation of this manuscript.

The authors declare that they have no conflict of interest.