Extended-spectrum b-lactamase (ESBL) and AmpC b-lactamase-producing Enterobacteriaceae are no longer restricted to the health care system, but represent increased risks related to environmental integrity and food safety. Fresh produce has been increasingly reported to constitute a reservoir of multidrug-resistant (MDR) potential human pathogenic Enterobacteriaceae. This study aimed to detect, identify, and characterize the antimicrobial resistance of ESBL/AmpC-producing Enterobacteriaceae isolates from fresh vegetables at point of sale. Vegetable samples (spinach, tomatoes, lettuce, cucumber, and green beans; n=545) were purchased from retailers in Gauteng, the most densely populated province in South Africa. These included street vendors, trolley vendors, farmers' market stalls, and supermarket chain stores. Selective enrichment, plating onto chromogenic media, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDITOF MS) confirmation of isolate identities showed that 17.4% (95/545) vegetable samples analyzed were contaminated with presumptive ESBL/AmpC-producing Enterobacteriaceae. Dominant species identified included Escherichia coli, Enterobacter cloacae, Enterobacter asburiae, and Klebsiella pneumoniae. Phenotypic antibiotic resistance analysis showed that 96.1% of 77 selected isolates were MDR, while resistance to aminoglycoside (94.8%), chloramphenicol (85.7%), and tetracycline (53.2%) antibiotic classes was most prevalent. Positive phenotypic analysis for ESBL production was shown in 61 (79.2%) of the 77 isolates, and AmpC production in 41.6% of the isolates. PCR and sequencing confirmed the presence of b-lactamase genes in 75.3% isolates from all vegetable types analyzed, mainly in E. coli, Enterobacter spp., and Serratia spp. isolates. CTXM group 9 (32.8%) was the dominant ESBL type, while EBC (24.1%) was the most prevalent plasmidic type AmpC b-lactamase. Our findings document for the first time the presence of MDR ESBL/AmpC-producing Enterobacteriaceae in raw vegetables sold at selected retailers in Gauteng Province, South Africa.

The increasing occurrence of multidrug-resistant (MDR) extended-spectrum β-lactamase-(ESBL) and/or AmpC β-lactamase-producing Enterobacteriaceae in health care systems, the environment and fresh produce is a serious concern globally. Production practices, processing and subsequent consumption of contaminated raw fruit and vegetables represent a possible human transmission route. The purpose of this study was to determine the presence of ESBL/AmpC-producing Enterobacteriaceae in complete spinach supply chains and to characterize the isolated strains phenotypically (antimicrobial resistance profiles) and genotypically (ESBL/AmpC genetic determinants, detection of class 1, 2, and 3 integrons). Water, soil, fresh produce, and contact surface samples (n = 288) from two commercial spinach production systems were screened for ESBL/AmpC-producing Enterobacteriaceae. In total, 14.58% (42/288) of the samples were found to be contaminated after selective enrichment, plating onto chromogenic media and matrix-assisted laser desorption ionization time-of-flight mass spectrometry identity confirmation of presumptive ESBL/AmpC isolates. This included 15.28% (11/72) water and 12.12% (16/132) harvested-and processed spinach, while 25% (15/60) retail spinach samples were found to be contaminated with an increase in isolate abundance and diversity in both scenarios. Dominant species identified included Serratia fonticola (45.86%), Escherichia coli (20.83%), and Klebsiella pneumoniae (18.75%). In total, 48 (81.36%) isolates were phenotypically confirmed as ESBL/AmpC-producing Enterobacteriaceae of which 98% showed a MDR phenotype. Genotypic characterization (PCR of ESBL/AmpC resistance genes and integrons) further revealed the domination of the CTX-M Group 1 ESBL type, followed by TEM and SHV; whilst the CIT-type was the only plasmid-mediated AmpC genetic determinant detected. Integrons were detected in 79.17% (n = 38) of the confirmed

Contaminated fresh produce has increasingly been implicated in foodborne disease outbreaks. As microbiological safety surveillance in South Africa is limited, a total of 545 vegetable samples (spinach, tomato, lettuce, cucumber, and green beans) were purchased from retailers, street traders, trolley vendors and farmers’ markets. Escherichia coli, coliforms and Enterobacteriaceae were enumerated and the prevalence of Escherichia coli, Salmonella spp. and Listeria monocytogenes determined. E. coli isolates were characterized phenotypically (antibiotic resistance) and genotypically (diarrheagenic virulence genes). Coliforms, E. coli and Enterobacteriaceae counts were mostly not significantly different between formal and informal markets, with exceptions noted on occasion. When compared to international standards, 90% to 98% tomatoes, 70% to 94% spinach, 82% cucumbers, 93% lettuce, and 80% green bean samples, had satisfactory (≤ 100 CFU/g) E. coli counts. Of the 545 vegetable samples analyzed, 14.86% (n = 81) harbored E. coli, predominantly from leafy green vegetables. Virulence genes (lt, st, bfpA, eagg, eaeA, stx1, stx2, and ipaH) were not detected in the E. coli isolates (n = 67) characterized, however 40.30% were multidrug‐resistant. Resistance to aminoglycosides (neomycin, 73.13%; gentamycin, < 10%), penicillins (ampicillin, 38.81%; amoxicillin, 41.79%; augmentin, < 10%), sulfonamides (cotrimoxazole, 22.39%), tetracycline (19.4%), chloramphenicol (11.94%), cephalosporins (cefepime, 34.33%), and carbapenemases (imipenem, < 10%) were observed. This study highlights the need for continued surveillance of multidrug resistant foodborne pathogens in fresh produce retailed formally and informally for potential consumer health risks.Practical ApplicationThe results indicate that the microbiological quality of different vegetables were similar per product type, regardless of being purchased from formal retailers or informal street traders, trolley vendors or farmers’ markets. Although no pathogenic bacteria (diarrheagenic E. coli, Salmonella spp. or L. monocytogenes) were isolated, high levels of multidrug‐resistance was observed in the generic E. coli isolates. These findings highlight the importance of microbiological quality surveillance of fresh produce in formal and informal markets, as these products can be a reservoir of multidrug resistant bacteria harboring antibiotic resistance and virulence genes, potentially impacting human health.

BackgroundMultidrug-resistant extended-spectrum β-lactamase (ESBL)-producing Enterobacterales is regarded as a critical health issue, yet, surveillance in the water-plant-food interface remains low, especially in Africa.ObjectivesThe objective of the study was to elucidate the distribution and prevalence of antimicrobial resistance in clinically significant members of the Enterobacterales order isolated from the water-plant-food interface in Africa.MethodsA literature search was conducted using six online databases according to the PRISMA guidelines. All available published studies involving phenotypic and genotypic characterization of ESBL-producing Enterobacterales from water, fresh produce or soil in Africa were considered eligible. Identification and characterization methods used as well as a network analysis according to the isolation source and publication year were summarized. Analysis of Escherichia coli, Salmonella spp. and Klebsiella pneumoniae included the calculation of the multiple antibiotic resistance (MAR) index according to isolation sources and statistical analysis was performed using RStudio.ResultsOverall, 51 studies were included for further investigation. Twelve African countries were represented, with environmental AMR surveillance studies predominantly conducted in South Africa. In 76.47% of the studies, occurrence of antimicrobial resistant bacteria was investigated in irrigation water samples, while 50.98% of the studies included fresh produce samples. Analysis of bacterial phenotypic antimicrobial resistance profiles were reported in 94.12% of the studies, with the disk diffusion method predominantly used. When investigating the MAR indexes of the characterized Escherichia coli, Klebsiella pneumoniae and Salmonella spp., from different sources (water, fresh produce or soil), no significant differences were seen across the countries. The only genetic determinant identified using PCR detection in all the studies was the blaCTX − M resistance gene. Only four studies used whole genome sequence analysis for molecular isolate characterization.DiscussionGlobally, AMR surveillance programmes recognize ESBL- and carbapenemase-producing Enterobacterales as vectors of great importance in AMR gene dissemination. However, in low- and middle-income countries, such as those in Africa, challenges to implementing effective and sustainable AMR surveillance programmes remain. This review emphasizes the need for improved surveillance, standardized methods and documentation of resistance gene dissemination across the farm-to-fork continuum in Africa.