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19 pages, 2130 KiB  
Article
Isolation of ESBL-Producing Enterobacteriaceae in Food of Animal and Plant Origin: Genomic Analysis and Implications for Food Safety
by Rosa Fraccalvieri, Stefano Castellana, Angelica Bianco, Laura Maria Difato, Loredana Capozzi, Laura Del Sambro, Adelia Donatiello, Domenico Pugliese, Maria Tempesta, Antonio Parisi and Marta Caruso
Microorganisms 2025, 13(8), 1770; https://doi.org/10.3390/microorganisms13081770 - 29 Jul 2025
Viewed by 301
Abstract
Background: The spread of ESBL-producing Enterobacteriaceae (ESBL-PE) strains in food poses a potential risk to human health. The aim of the study was to determine the occurrence of ESBL-PE and to investigate their distribution on foods. Methods: A total of 1000 food [...] Read more.
Background: The spread of ESBL-producing Enterobacteriaceae (ESBL-PE) strains in food poses a potential risk to human health. The aim of the study was to determine the occurrence of ESBL-PE and to investigate their distribution on foods. Methods: A total of 1000 food samples, including both raw and ready-to-eat products, was analyzed for the presence of ESBL-producing Enterobacteriaceae using chromogenic selective agar. Antibiotic resistance in the isolated strains was assessed using conventional methods, while whole-genome sequencing was employed to predict antimicrobial resistance and virulence genes. Results: The overall occurrence of ESBL-PE strains was 2.8%, with the highest contamination in raw meat samples (10%). A total of 31 multidrug-resistant (MDR) strains was isolated, mainly Escherichia coli, followed by Klebsiella pneumoniae, Salmonella enterica, and Enterobacter hormaechei. All strains exhibited high levels of resistance to at least four different β-lactam antibiotics, as well as to other antimicrobial classes including sulfonamides, tetracyclines, aminoglycosides, and quinolones. Whole-genome sequencing identified 63 antimicrobial resistance genes, with blaCTX-M being the most prevalent ESBL gene. Twenty-eight (90%) isolates carried Inc plasmids, known vectors of multiple antimicrobial resistance genes, including those associated with ESBLs. Furthermore, several virulence genes were identified. Conclusions: The contamination of food with ESBL-PE represents a potential public health risk, underscoring the importance of the implementation of genomic surveillance to monitor and control the spread of antimicrobial resistance. Full article
(This article belongs to the Special Issue Food Microorganisms and Genomics, 2nd Edition)
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14 pages, 384 KiB  
Article
Outbreak Caused by VIM-1- and VIM-4-Positive Proteus mirabilis in a Hospital in Zagreb
by Branka Bedenić, Gernot Zarfel, Josefa Luxner, Andrea Grisold, Marina Nađ, Maja Anušić, Vladimira Tičić, Verena Dobretzberger, Ivan Barišić and Jasmina Vraneš
Pathogens 2025, 14(8), 737; https://doi.org/10.3390/pathogens14080737 - 26 Jul 2025
Viewed by 287
Abstract
Background/objectives: Proteus mirabilis is a frequent causative agent of urinary and wound infections in both community and hospital settings. It develops resistance to expanded-spectrum cephalosporins (ESCs) due to the production of extended-spectrum β-lactamases (ESBLs) or plasmid-mediated AmpC β-lactamases (p-AmpCs). Recently, carbapenem-resistant isolates of [...] Read more.
Background/objectives: Proteus mirabilis is a frequent causative agent of urinary and wound infections in both community and hospital settings. It develops resistance to expanded-spectrum cephalosporins (ESCs) due to the production of extended-spectrum β-lactamases (ESBLs) or plasmid-mediated AmpC β-lactamases (p-AmpCs). Recently, carbapenem-resistant isolates of P. mirabilis emerged due to the production of carbapenemases, mostly belonging to Ambler classes B and D. Here, we report an outbreak of infections due to carbapenem-resistant P. mirabilis that were observed in a psychiatric hospital in Zagreb, Croatia. The characteristics of ESBL and carbapenemase-producing P. mirabilis isolates, associated with an outbreak, were analyzed. Materials and methods: The antibiotic susceptibility testing was performed by the disk-diffusion and broth dilution methods. The double-disk synergy test (DDST) and inhibitor-based test with clavulanic and phenylboronic acid were applied to screen for ESBLs and p-AmpCs, respectively. Carbapenemases were screened by the modified Hodge test (MHT), while carbapenem hydrolysis was investigated by the carbapenem inactivation method (CIM) and EDTA-carbapenem-inactivation method (eCIM). The nature of the ESBLs, carbapenemases, and fluoroquinolone-resistance determinants was investigated by PCR. Plasmids were characterized by PCR-based replicon typing (PBRT). Selected isolates were subjected to molecular characterization of the resistome by an Inter-Array Genotyping Kit CarbaResisit and whole-genome sequencing (WGS). Results: In total, 20 isolates were collected and analyzed. All isolates exhibited resistance to amoxicillin alone and when combined with clavulanic acid, cefuroxime, cefotaxime, ceftriaxone, cefepime, imipenem, ceftazidime–avibactam, ceftolozane–tazobactam, gentamicin, amikacin, and ciprofloxacin. There was uniform susceptibility to ertapenem, meropenem, and cefiderocol. The DDST and combined disk test with clavulanic acid were positive, indicating the production of an ESBL. The MHT was negative in all except one isolate, while the CIM showed moderate sensitivity, but only with imipenem as the indicator disk. Furthermore, eCIM tested positive in all of the CIM-positive isolates, consistent with a metallo-β-lactamase (MBL). PCR and sequencing of the selected amplicons identified VIM-1 and VIM-4. The Inter-Array Genotyping Kit CarbaResist and WGS identified β-lactam resistance genes blaVIM, blaCTX-M-15, and blaTEM genes; aminoglycoside resistance genes aac(3)-IId, aph(6)-Id, aph(3″)-Ib, aadA1, armA, and aac(6′)-IIc; as well as resistance genes for sulphonamides sul1 and sul2, trimethoprim dfr1, chloramphenicol cat, and tetracycline tet(J). Conclusions: This study revealed an epidemic spread of carbapenemase-producing P. mirabilis in two wards in a psychiatric hospital. Due to the extensively resistant phenotype (XDR), therapeutic options were limited. This is the first report of carbapenemase-producing P. mirabilis in Croatia. Full article
(This article belongs to the Special Issue Emerging and Neglected Pathogens in the Balkans)
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20 pages, 2552 KiB  
Article
Environmental Dispersion of Multiresistant Enterobacteriaceae in Aquatic Ecosystems in an Area of Spain with a High Density of Pig Farming
by Javier Díez de los Ríos, Noemí Párraga-Niño, María Navarro, Judit Serra-Pladevall, Anna Vilamala, Elisenda Arqué, María Baldà, Tamar Nerea Blanco, Luisa Pedro-Botet, Óscar Mascaró and Esteban Reynaga
Antibiotics 2025, 14(8), 753; https://doi.org/10.3390/antibiotics14080753 - 25 Jul 2025
Viewed by 298
Abstract
Background: This study aimed to (a) assess the prevalence of multidrug-resistant (MDR) Enterobacteriaceae in the waters of two rivers and wastewater treatment plants (WWTPs) in a region of Catalonia, Spain; (b) genetically characterize the MDR strains; and (c) compare extended-spectrum β-lactamase (ESBL)-producing [...] Read more.
Background: This study aimed to (a) assess the prevalence of multidrug-resistant (MDR) Enterobacteriaceae in the waters of two rivers and wastewater treatment plants (WWTPs) in a region of Catalonia, Spain; (b) genetically characterize the MDR strains; and (c) compare extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates from environmental and human sources. Methods: A total of 62 samples were collected from the influent and effluent of 31 WWTPs and 29 river water samples from 11 sites. Simultaneously, 382 hospitalized patients were screened for MDR Enterobacteriaceae using rectal swabs. All isolates underwent antibiotic susceptibility testing and whole-genome sequencing. Results: MDR Enterobacteriaceae were detected in 48.4% of WWTP samples, with 18.5% ESBL-producing E. coli and 1.5% (one sample) OXA-48-producing K. pneumoniae in influents, and 12.8% ESBL-producing E. coli in effluents. In river waters, 5.6% of samples contained ESBL-producing E. coli and 1.4% (1 sample) contained VIM-producing Enterobacter cloacae complex strains. Among patients, 10.2% (39/382) carried MDR Gram-negative bacilli, of which 66.7% were ESBL-producing E. coli. In aquatic ecosystems E. coli ST131 (13.3%) and ST162 (13.3%) were the most common strains, while in humans the common were E. coli ST131 (33.3%), ST69 (11.1%) and ST410 (7.4%) in humans. The most frequent environmental antibiotic resistance genes (ARG) were blaCTX-M-15 (24%) and blaTEM-1B (20%), while the most common ARGs were blaTEM-1B (20.4%), blaCTX-M15 (18.4%) and blaCTX-M-27 (14.3%). IncF plasmids were predominant in environmental and human strains. Conclusions: ESBL-producing E. coli and carbapenemase-producing Enterobacteriaceae are present in aquatic environments in the region. Phylogenetic similarities between environmental and clinical strains suggest a possible similar origin. Further studies are necessary to clarify transmission routes and environmental impact. Full article
(This article belongs to the Special Issue A One Health Approach to Antimicrobial Resistance, 2nd Edition)
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16 pages, 3400 KiB  
Article
Antimicrobial Resistance Genotypes and Mobile Genetic Elements of Poultry-Derived Escherichia coli: A Retrospective Genomic Study from the United States
by Sohyun Cho, Hazem Ramadan, Lari M. Hiott, Jonathan G. Frye and Charlene R. Jackson
Pathogens 2025, 14(8), 726; https://doi.org/10.3390/pathogens14080726 - 23 Jul 2025
Viewed by 618
Abstract
The presence of antibiotic resistance in commensal bacteria may be an influential factor in the persistence of resistance in pathogens. This is especially critical for Escherichia coli that consumers may be exposed to through the consumption of uncooked meat. In this study, E. [...] Read more.
The presence of antibiotic resistance in commensal bacteria may be an influential factor in the persistence of resistance in pathogens. This is especially critical for Escherichia coli that consumers may be exposed to through the consumption of uncooked meat. In this study, E. coli isolates previously recovered from poultry in the US between 2001 and 2012 were whole-genome sequenced to identify their antibiotic resistance genes and mobile genetic elements. The genomes of 98 E. coli isolates from poultry carcass rinsates and 2 isolates from poultry diagnostic samples with multidrug resistance or potential extended-spectrum β-lactam (ESBL)-producing phenotypes as well as the genetic variabilities among the E. coli were assessed. All E. coli isolates were positive for at least one antibiotic resistance gene and plasmid replicon, with 37 resistance genes and 27 plasmid replicons detected among the isolates. While no ESBL genes were detected, blaCMY-2 was the most common β-lactamase gene, and blaTEM and blaCARB-2 were also identified. Most isolates (95%) harbored at least one intact phage, and as many as seven intact phages were identified in one isolate. These results show the occurrence of antibiotic resistance genes and mobile genetic elements in these 100 poultry-associated E. coli isolates, which may be responsible for the resistance phenotypes exhibited by the isolates. This retrospective study also enables comparisons of resistance genes and mobile genetic elements from more recent E. coli isolates associated with poultry to aid in understanding the trends of both antibiotic resistance phenotypes and genotypes in the poultry setting over time. Full article
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11 pages, 775 KiB  
Article
Whole Genome Sequencing of Klebsiella variicola Strains Isolated from Patients with Cancer
by Alicja Sękowska, Andrés Carrazco-Montalvo and Yulian Konechnyi
Antibiotics 2025, 14(8), 735; https://doi.org/10.3390/antibiotics14080735 - 22 Jul 2025
Viewed by 456
Abstract
Background: Klebsiella variicola is a Gram-negative, capsulated, nonmotile, facultative anaerobic rod. It is one of the species belonging to the K. pneumoniae complex. The objective of this study was to gain insights into the antimicrobial resistance and virulence of K. variicola [...] Read more.
Background: Klebsiella variicola is a Gram-negative, capsulated, nonmotile, facultative anaerobic rod. It is one of the species belonging to the K. pneumoniae complex. The objective of this study was to gain insights into the antimicrobial resistance and virulence of K. variicola strains isolated from clinical samples from oncologic patients. Methods: Strain identification was performed using a mass spectrometry method. Whole genome sequencing was conducted for all analyzed strains. Antimicrobial susceptibility was determined using an automated method. The presence of antimicrobial resistance mechanisms and genes encoding extended-spectrum beta-lactamases (ESBL) was assessed using the double-disc synergy test and genotypic methods. Results: All isolates were identified as K. variicola using mass spectrometry and whole genome sequencing (WGS). All isolates were ESBL-positive, and two of them harbored the blaCTX-M-15 gene. In our study, the blaLEN-17 gene was detected in all strains. Genome sequence analysis of the K. variicola isolates revealed the presence of virulence factor genes, including entAB, fepC, ompA, ykgK, and yagWXYZ. Two different plasmids, IncFIB(K) and IncFII, were identified in all of the analyzed K. variicola strains. The detected virulence factors suggest the ability of the bacteria to survive in the environment and infect host cells. All isolates demonstrated in vitro susceptibility to carbapenems. Conclusions: Further studies are needed to confirm whether multidrug-resistant K. variicola strains represent an important pathogen in infections among oncologic patients. Full article
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15 pages, 2742 KiB  
Article
Resistome and Phylogenomics of Escherichia coli Strains Obtained from Diverse Sources in Jimma, Ethiopia
by Mulatu Gashaw, Esayas Kebede Gudina, Guenter Froeschl, Ralph Matar, Solomon Ali, Liegl Gabriele, Amelie Hohensee, Thomas Seeholzer, Arne Kroidl and Andreas Wieser
Antibiotics 2025, 14(7), 706; https://doi.org/10.3390/antibiotics14070706 - 14 Jul 2025
Viewed by 356
Abstract
Introduction: In recent years, antimicrobial resistance (AMR) rates have increased significantly in bacterial pathogens, particularly extended beta-lactam resistance. This study aimed to investigate resistome and phylogenomics of Escherichia coli (E. coli) strains isolated from various sources in Jimma, Ethiopia. Methods [...] Read more.
Introduction: In recent years, antimicrobial resistance (AMR) rates have increased significantly in bacterial pathogens, particularly extended beta-lactam resistance. This study aimed to investigate resistome and phylogenomics of Escherichia coli (E. coli) strains isolated from various sources in Jimma, Ethiopia. Methods: Phenotypic antibiotic resistance patterns of E. coli isolates were determined using automated Kirby–Bauer disc diffusion and minimum inhibitory concentration (MIC). Isolates exhibiting phenotypic resistance to beta-lactam antibiotics were further analyzed with a DNA microarray to confirm the presence of resistance-encoding genes. Additionally, multilocus sequence typing (MLST) of seven housekeeping genes was conducted using PCR and Oxford Nanopore-Technology (ONT) to assess the phylogenetic relationships among the E. coli isolates. Results: A total of 611 E. coli isolates from human, animal, and environmental sources were analyzed. Of these, 41.6% (254) showed phenotypic resistance to at least one of the tested beta-lactams, 96.1% (244) thereof were confirmed genotypically. More than half of the isolates (53.3%) had two or more resistance genes present. The most frequent ESBL-encoding gene was CTX-M-15 (74.2%; 181), followed by TEM (59.4%; 145) and CTX-M-9 (4.1%; 10). The predominant carbapenemase gene was NDM-1, detected in 80% (12 out of 15) of carbapenem-resistant isolates. A phylogenetic analysis revealed clonality among the strains obtained from various sources, with international high-risk clones such as ST131, ST648, ST38, ST73, and ST405 identified across various niches. Conclusions: The high prevalence of CTX-M-15 and NDM-1 in multidrug-resistant E. coli isolates indicates the growing threat of AMR in Ethiopia. The discovery of these high-risk clones in various niches shows possible routes of transmission and highlights the necessity of a One Health approach to intervention and surveillance. Strengthening antimicrobial stewardship, infection prevention, and control measures are crucial to mitigate the spread of these resistant strains. Full article
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16 pages, 1767 KiB  
Article
Population Structure, Genomic Features, and Antibiotic Resistance of Avian Pathogenic Escherichia coli in Shandong Province and Adjacent Regions, China (2008–2023)
by Shikai Song, Yao Wang, Zhihai Liu, Rongling Zhang, Kaiyuan Li, Bin Yin, Zunxiang Yan, Shifa Yang, Shuqian Lin and Yunpeng Yi
Microorganisms 2025, 13(7), 1655; https://doi.org/10.3390/microorganisms13071655 - 13 Jul 2025
Viewed by 649
Abstract
Avian pathogenic Escherichia coli (APEC) poses a global threat to poultry health and public safety due to its high lethality, limited treatment options, and potential for zoonotic transmission via the food chain. However, long-term genomic surveillance remains limited, especially in countries like China [...] Read more.
Avian pathogenic Escherichia coli (APEC) poses a global threat to poultry health and public safety due to its high lethality, limited treatment options, and potential for zoonotic transmission via the food chain. However, long-term genomic surveillance remains limited, especially in countries like China where poultry farming is highly intensive. This study aimed to characterize the population structure, virulence traits, and antimicrobial resistance of 81 APEC isolates from diseased chickens collected over 16 years from Shandong and neighboring provinces in eastern China. The isolates were grouped into seven Clermont phylogroups, with A and B1 being dominant. MLST revealed 27 STs, and serotyping identified 29 O and 16 H antigens, showing high genetic diversity. The minor phylogroups (B2, C, D, E, G) encoded more virulence genes and had higher virulence-plasmid ColV carriage, with enrichment for iron-uptake, protectins, and extraintestinal toxins. In contrast, the dominant phylogroups A and B1 primarily carried adhesin and enterotoxin genes. Antimicrobial resistance was widespread: 76.5% of isolates were multidrug-resistant. The minor phylogroups exhibited higher tetracycline resistance (mediated by tet(A)), whereas the major phylogroups showed increased resistance to third- and fourth-generation cephalosporins (due to blaCTX-M-type ESBL genes). These findings offer crucial data for APEC prevention and control, safeguarding the poultry industry and public health. Full article
(This article belongs to the Section Antimicrobial Agents and Resistance)
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16 pages, 613 KiB  
Article
Isolation and Molecular Characterization of Antimicrobial-Resistant Bacteria from Vegetable Foods
by Annamaria Castello, Chiara Massaro, Erine Seghers, Clelia Ferraro, Antonella Costa, Rosa Alduina and Cinzia Cardamone
Pathogens 2025, 14(7), 682; https://doi.org/10.3390/pathogens14070682 - 10 Jul 2025
Viewed by 370
Abstract
Antimicrobial resistance (AMR) poses a growing threat to global health, and its spread through the food chain is gaining increasing attention. While AMR in food of animal origin has been extensively studied, less is known about its prevalence in plant-based foods, particularly fresh [...] Read more.
Antimicrobial resistance (AMR) poses a growing threat to global health, and its spread through the food chain is gaining increasing attention. While AMR in food of animal origin has been extensively studied, less is known about its prevalence in plant-based foods, particularly fresh and ready-to-eat (RTE) vegetables. This study investigated the occurrence of antimicrobial-resistant bacteria in fresh and RTE vegetables. Isolates were subjected to antimicrobial susceptibility testing and molecular analyses for the characterization of antimicrobial resistance genes (ARGs). A significant proportion of samples were found to harbor antimicrobial-resistant bacteria, including multidrug-resistant strains. Several ARGs, including those encoding extended-spectrum β-lactamases (ESBLs) and resistance to critically important antimicrobials, were detected. The findings point to environmental contamination—potentially originating from wastewater reuse and agricultural practices—as a likely contributor to AMR dissemination in vegetables. The presence of antimicrobial-resistant bacteria and ARGs in fresh produce raises concerns about food safety and public health. The current regulatory framework lacks specific criteria for monitoring AMR in vegetables, highlighting the urgent need for surveillance programs and risk mitigation strategies. This study contributes to a better understanding of AMR in the plant-based food sector and supports the implementation of a One Health approach to address this issue. Full article
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22 pages, 1347 KiB  
Article
The Microbiological Characteristics and Genomic Surveillance of Carbapenem-Resistant Klebsiella pneumoniae Isolated from Clinical Samples
by Mehwish Rizvi, Noman Khan, Ambreen Fatima, Rabia Bushra, Ale Zehra, Farah Saeed and Khitab Gul
Microorganisms 2025, 13(7), 1577; https://doi.org/10.3390/microorganisms13071577 - 4 Jul 2025
Viewed by 593
Abstract
Klebsiella pneumoniae is a major public health concern due to its role in Gram-negative bacteremia, which leads to high mortality and increased healthcare costs. This study characterizes phenotypic and genomic features of K. pneumoniae isolates from clinical samples in Karachi, Pakistan. Among 507 [...] Read more.
Klebsiella pneumoniae is a major public health concern due to its role in Gram-negative bacteremia, which leads to high mortality and increased healthcare costs. This study characterizes phenotypic and genomic features of K. pneumoniae isolates from clinical samples in Karachi, Pakistan. Among 507 isolates, 213 (42%) were carbapenem-resistant based on disk diffusion and MIC testing. Urine (29.7%) and blood (28.3%) were the most common sources, with infections predominantly affecting males (64.7%) and individuals aged 50–70 years. Colistin was the only antibiotic showing consistent activity against these isolates. The whole-genome sequencing of 24 carbapenem-resistant K. pneumoniae (CR-KP) isolates revealed blaNDM-5 (45.8%) as the dominant carbapenemase gene, followed by blaNDM-1 (12.5%) and blaOXA-232 (54.2%). Other detected blaOXA variants included blaOXA-1, blaOXA-4, blaOXA-10, and blaOXA-18. The predominant beta-lactamase gene was blaCTX-M-15 (91.6%), followed by blaCTX-M-163, blaCTX-M-186, and blaCTX-M-194. Sequence types ST147, ST231, ST29, and ST11 were associated with resistance. Plasmid profiling revealed IncR (61.5%), IncL (15.4%), and IncC (7.7%) as common plasmid types. Importantly, resistance was driven not only by acquired genes but also by chromosomal mutations. Porin mutations in OmpK36 and OmpK37 (e.g., P170M, I128M, N230G, A217S) reduced drug influx, while acrR and ramR mutations (e.g., P161R, G164A, P157*) led to efflux pump overexpression, enhancing resistance to fluoroquinolones and tigecycline. These findings highlight a complex resistance landscape driven by diverse carbapenemases and ESBLs, underlining the urgent need for robust antimicrobial stewardship and surveillance strategies. Full article
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20 pages, 1599 KiB  
Article
Amoxicillin Resistance: An In Vivo Study on the Effects of an Approved Formulation on Antibiotic Resistance in Broiler Chickens
by Ádám Kerek, Ábel Szabó and Ákos Jerzsele
Animals 2025, 15(13), 1944; https://doi.org/10.3390/ani15131944 - 1 Jul 2025
Viewed by 432
Abstract
Background: Antimicrobial resistance (AMR) is a growing global concern in poultry production, where antibiotic use can disrupt gut microbiota and enrich antimicrobial resistance genes (ARGs). Objectives: This study aimed to assess the in vivo effects of a veterinary-approved amoxicillin formulation on gut microbiome [...] Read more.
Background: Antimicrobial resistance (AMR) is a growing global concern in poultry production, where antibiotic use can disrupt gut microbiota and enrich antimicrobial resistance genes (ARGs). Objectives: This study aimed to assess the in vivo effects of a veterinary-approved amoxicillin formulation on gut microbiome composition and ARG profiles in broiler chickens. Methods: A total of 120 Ross-308 broiler chickens were randomly allocated into 12 experimental groups (n = 10 per group), with three replicates per treatment. Birds received either full-dose (1×), a subtherapeutic quarter-dose (¼×) of amoxicillin, a placebo (starch), or no treatment. Cloacal swabs were collected on days 0, 14, and 28 for shotgun metagenomic sequencing. One-way ANOVA was used to evaluate treatment effects on body weight, with significant differences observed from day 14 onward (p < 0.0001). Results: The ¼× dose caused a more pronounced microbiome shift than the 1× dose, with a marked reduction in Pseudomonadota and increase in Bacillota and Bacteroidota. ARG abundance declined in the ¼× group (from 1386 to 1012). While TEM-type ESBL genes were ubiquitous, CTX-M-1 emerged only after ¼× treatment. Worryingly, 20 types of vancomycin resistance genes were detected across all samples. Plasmid-borne ARGs and mobile genetic elements decreased in the ¼× group. Conclusions: Even subtherapeutic antibiotic exposure significantly reshapes the gut microbiota composition and ARG landscape, highlighting the need for refined risk assessments and microbiome-conscious antimicrobial policies in poultry farming. Full article
(This article belongs to the Section Poultry)
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14 pages, 1389 KiB  
Article
Lack of Association Between qacE and qacE∆1 Gene Variants and Sodium Hypochlorite Resistance in Clinical Isolates of ESBL- and Carbapenemase-Producing Klebsiella spp. and Enterobacter spp., from Gaborone, Botswana
by Pearl Ntshonga, Tlhalefo Dudu Ntereke, Tshiamo Zankere, Daniel Paul Morse, Garesego Koto, Irene Gobe and Giacomo Maria Paganotti
Antibiotics 2025, 14(7), 662; https://doi.org/10.3390/antibiotics14070662 - 30 Jun 2025
Viewed by 371
Abstract
Background: The qacE gene and its variant, qacE∆1, have been associated with resistance to antimicrobials and biocides. This poses a threat to infection prevention, control and treatment. Several studies investigated this relationship with conflicting results. The aim of this research was [...] Read more.
Background: The qacE gene and its variant, qacE∆1, have been associated with resistance to antimicrobials and biocides. This poses a threat to infection prevention, control and treatment. Several studies investigated this relationship with conflicting results. The aim of this research was to determine the prevalence of qacE and qacE∆1 in clinical extended spectrum β-lactamase- (ESBL) and carbapenemase-producing Klebsiella spp. and Enterobacter spp. and elucidate the association of these genes with resistance to sodium hypochlorite. Methods: This study included 216 isolates of ESBL- and carbapenemase-producing multidrug-resistant (MDR) Klebsiella spp. and Enterobacter spp. These isolates were identified by VITEK-2 analyser. The MIC and MBC of sodium hypochlorite were determined using the microbroth serial-dilution method. PCR was used to detect gene variants. A regression analysis investigated any association between qacE genotypes, MIC and MBC, as well as antimicrobial drug resistance profiles. Results: Overall, there was a high prevalence of qacE and qacE∆1 variants (84.7%; 95% CI, 79.2–89.2). There was a high prevalence of qacE∆1 (80.6%; 95% CI, 74.6–85.6) as compared to qacE (15.3%, 95% CI, 10.8–20.8). The MIC50 and MIC90 of the isolates ranged between 7031 mg/L and 9375 mg/L and 14,060 mg/L and 18,750 mg/L, respectively, while the MBC ranged from 48,750 mg/L to 18,750 mg/L. There was no association between qacE genotypes and high MIC and MBC as well as antimicrobial drug resistance. Conclusions: The MIC and MBC of sodium hypochlorite are higher than what is currently used for disinfection in Botswana. There is a high prevalence of qacE and qacE∆1; however, these genes do not seem to be associated with resistance to sodium hypochlorite. Full article
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21 pages, 1726 KiB  
Article
Exploring Virulence Characteristics of Clinical Escherichia coli Isolates from Greece
by Lazaros A. Gagaletsios, Elisavet Kikidou, Christos Galbenis, Ibrahim Bitar and Costas C. Papagiannitsis
Microorganisms 2025, 13(7), 1488; https://doi.org/10.3390/microorganisms13071488 - 26 Jun 2025
Viewed by 377
Abstract
The aim of this study was to examine the genetic characteristics that could be associated with the virulence characteristics of Escherichia coli collected from clinical samples. A collection of 100 non-repetitive E. coli isolates was analyzed. All isolates were typed by MLST. String [...] Read more.
The aim of this study was to examine the genetic characteristics that could be associated with the virulence characteristics of Escherichia coli collected from clinical samples. A collection of 100 non-repetitive E. coli isolates was analyzed. All isolates were typed by MLST. String production, biofilm formation and serum resistance were examined for all isolates. Twenty E. coli isolates were completely sequenced Illumina platform. The results showed that the majority of E. coli isolates (87%) produced significant levels of biofilm, while none of the isolates were positive for string test and resistance to serum. Additionally, the presence of CRISPR/Cas systems (type I-E or I-F) was found in 18% of the isolates. Analysis of WGS data found that all sequenced isolates harbored a variety of virulence genes that could be implicated in adherence, invasion, iron uptake. Also, WGS data confirmed the presence of a wide variety of resistance genes, including ESBL- and carbapenemase-encoding genes. In conclusion, an important percentage (87%) of the E. coli isolates had a significant ability to form biofilm. Biofilms, due to their heterogeneous nature and ability to make microorganisms tolerant to multiple antimicrobials, complicate treatment strategies. Thus, in combination with the presence of multidrug resistance, expression of virulence factors could challenge antimicrobial therapy of infections caused by such bacteria. Full article
(This article belongs to the Section Molecular Microbiology and Immunology)
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19 pages, 2649 KiB  
Article
Integrating Nanopore MinION Sequencing into National Animal Health AMR Surveillance Programs: An Indonesian Pilot Study of Chicken Slaughterhouse Effluent and Rivers
by Rallya Telussa, Puji Rahayu, Thufeil Yunindika, Curtis J. Kapsak, Kanti Puji Rahayu, Oli Susanti, Imron Suandy, Nuraini Triwijayanti, Aji B. Niasono, Syamsul Ma’arif, Hendra Wibawa, Lestari Lestari, Gunawan B. Utomo, Farida C. Zenal, Luuk Schoonman and Lee E. Voth-Gaeddert
Antibiotics 2025, 14(7), 624; https://doi.org/10.3390/antibiotics14070624 - 20 Jun 2025
Viewed by 644
Abstract
Background: Antimicrobial resistance (AMR) poses significant risks to human and animal health, while the environment can contribute to its spread. National AMR surveillance programs are pivotal for assessing AMR prevalence, trends, and intervention outcomes; however, integrating advanced surveillance tools can be difficult. This [...] Read more.
Background: Antimicrobial resistance (AMR) poses significant risks to human and animal health, while the environment can contribute to its spread. National AMR surveillance programs are pivotal for assessing AMR prevalence, trends, and intervention outcomes; however, integrating advanced surveillance tools can be difficult. This pilot study, conducted by FAO ECTAD Indonesia and DGLAHS, the Indonesian Ministry of Agriculture, evaluated the costs and benefits of integrating the Nanopore MinION, Illumina MiSeq, and Sensititre system into a culture-based slaughterhouse–river surveillance system. Methods: Water samples were collected from six chicken slaughterhouses and adjacent rivers (pre- and post-treatment effluent, upstream, and downstream). Culture-based ESBL and general E. coli concentrations were estimated via the WHO Tricycle Protocol, while isolates (n = 42) were sequenced (MinION, MiSeq) and antimicrobial susceptibility testing conducted (Sensititre). Results: The Tricycle Protocol results provided estimates of effluent and river concentrations of ESBL and general E. coli identifying ESBL-to-general E. coli ratios of 13.8% and 6.2%, respectively. Compared to hybrid sequencing assemblies, MinION had a higher concordance than MiSeq for ARG identification (98%), virulence genes (96%), and locations for both (predominately plasmids). Furthermore, MinION concordance with Sensititre AST was 91%. Conclusions: Cost–benefit comparisons suggest sequencing can complement culture-based methods but is dependent on the value placed on the additional information gained. Full article
(This article belongs to the Special Issue Microbial Resistance Surveillance and Management in Food Systems)
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25 pages, 3717 KiB  
Article
Genotypic Characterisation and Risk Assessment of Virulent ESBL-Producing E. coli in Chicken Meat in Tunisia: Insights from Multi-Omics Machine Learning Perspective
by Khaled Abdallah, Ghassan Tayh, Elaa Maamar, Amine Mosbah, Omar Abbes, Ismail Fliss and Lilia Messadi
Microbiol. Res. 2025, 16(6), 131; https://doi.org/10.3390/microbiolres16060131 - 18 Jun 2025
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Abstract
Antibiotics are frequently used in the poultry industry, which has led to the emergence of bacterial strains that are resistant to antimicrobial treatments. The main objectives of this research were to conduct a multimodal risk assessment, to determine the extent of contamination of [...] Read more.
Antibiotics are frequently used in the poultry industry, which has led to the emergence of bacterial strains that are resistant to antimicrobial treatments. The main objectives of this research were to conduct a multimodal risk assessment, to determine the extent of contamination of chicken meat with Escherichia coli, assess the prevalence of strains resistant to extended-spectrum cephalosporins (ESC), and characterise the genes associated with resistance and virulence. A standardised procedure involving enrichment in buffered peptone water and isolation of E. coli on MacConkey agar was carried out on 100 chicken carcasses. Subsequently, the sensitivity of the strains was tested against 21 antibiotic discs. Additionally, ESBL production was detected using a double synergy test. Specific PCRs were employed to identify resistance to critical antibiotics in human medicine (such as cephalosporins, carbapenems, fluoroquinolones, and colistin), as well as the presence of virulence genes. The contamination rate of chicken meat with E. coli was 82%. The prevalence of ESC-resistant isolates was 91.2%. Furthermore, 76.5% of the isolates exhibited ESBL production, with the different beta-lactamase genes (blaCTXM, blaTEM, and blaSHV). The mcr-1 gene, associated with colistin resistance, was detected in four strains (5.9%). Some isolates also carried resistance genes such as sul1, sul2, sul3, tetA, tetB, qnrB, and qnrS. In addition, several virulence genes were detected. In our study, we were able to link the expression of AMR to the iron metabolic regulatory elements using a multimodal machine learning approach; this mechanism could be targeted to mitigate the bacteria virulence and resistance. The high prevalence of ESBL-producing and multi-resistant E. coli strains in poultry presents significant human health risks, with the focus on antibiotic-resistant uropathogenic strains since poultry meat could be an important source of uropathogenic strains, underscoring the danger of hard-to-treat urinary tract infections, stressing the need for controlled antibiotic use and thorough monitoring. Full article
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11 pages, 257 KiB  
Article
Antibiotic Resistance Profiles of Diarrhoeagenic Enterobacterales in Bioko Island, Equatorial Guinea
by Úrsula-Eva Eñeso Efuá, Silvia Herrera-León, Fátima Patabobe, Pascual Erasmo Owono and Agustín Benito
Acta Microbiol. Hell. 2025, 70(2), 24; https://doi.org/10.3390/amh70020024 - 10 Jun 2025
Viewed by 955
Abstract
Acute diarrhoeal disease caused by antibiotic-resistant diarrhoeagenic bacteria is a significant global public health issue, particularly in low- and middle-income countries. This study provides the first molecular characterisation of antimicrobial resistance profiles, including the detection of CTX-M-15 and CTX-M-55 extended-spectrum beta-lactamases (ESBLs), among [...] Read more.
Acute diarrhoeal disease caused by antibiotic-resistant diarrhoeagenic bacteria is a significant global public health issue, particularly in low- and middle-income countries. This study provides the first molecular characterisation of antimicrobial resistance profiles, including the detection of CTX-M-15 and CTX-M-55 extended-spectrum beta-lactamases (ESBLs), among diarrhoeagenic Enterobacterales in Bioko Island, Equatorial Guinea, offering novel epidemiological insights into an understudied region. This study investigated the antibiotic resistance profiles of pathogenic bacteria isolated from diarrhoeal samples on Bioko Island. A total of 153 clinical isolates were collected between 1 February and 30 May 2014, and antimicrobial susceptibility testing was performed at Loeri Comba Polyclinic (Malabo) using the Kirby–Bauer method. The molecular characterisation of β-lactamase-associated genes was performed on different isolates of diarrhoeagenic pathotypes—144 Escherichia coli, 7 Salmonella enterica, and 2 Shigella flexneri—at the National Centre for Microbiology (Majadahonda, Spain). High resistance rates were detected against ampicillin (98%), tetracycline (93.5%), sulfonamides (94.8%), sulfamethoxazole–trimethoprim (88.2%), and cefotaxime (78.8%), while moderate rates of resistance were noted for ciprofloxacin (26.7%), and all isolates remained susceptible to imipenem. Of the isolates, 107 (69.9%) produced either single or multiple β-lactamases. Among these, 73 (68.2%) harbored classical β-lactamases, specifically TEM and OXA-1 types, representing 47.7% of the total sample. Additionally, 34 (31.8%) of the isolates were identified as producers of extended-spectrum β-lactamases (ESBLs), specifically CTX-M enzymes. Sequencing identified CTX-M-15 and CTX-M-55 variants. The predominant ESBL-producing bacteria were enteroaggregative Escherichia coli (56.2%), followed by enteropathogenic and enterotoxigenic E. coli. These findings confirm the circulation of multidrug-resistant diarrhoeagenic Enterobacterales in Equatorial Guinea, raising concerns about limited treatment options due to widespread resistance to multiple antibiotic classes, including third-generation cephalosporins and quinolones. The most important conclusion drawn from this study is that a high percentage of diarrhoeagenic bacteria have an antibiotic resistance and multi-resistance profile, especially to beta-lactams and other groups of antibiotics such as tetracyclines and sulphonamides. There is also a moderate prevalence of isolates carrying ESBLs on Bioko Island, Equatorial Guinea, which could indicate the inappropriate use of antimicrobials. Full article
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