Antimicrobial susceptibility and integron diversity in organic and conventionally grown fruits and vegetables

Jones-Dias, Daniela; Moura, Inês Barata; Ferreira, Eugénia; Caniça, Manuela

http://hdl.handle.net/10400.18/4177

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Introduction: During growth and harvesting fresh produce can become contaminated not only with environmental, but also pathogenic bacteria from animal and human sources. This study aimed to characterize the antibiotic resistant bacteria gathered from organically and conventionally grown fruits and vegetables, and to characterize the diversity of class 1, 2 and 3 integrons detected. Methods: Between 2013 and 2014, one conventionally and one organically produced batch of six fruits and vegetables were purchased at retail stores. Products were processed as follows: 50g of product was selected at random without washing or peeling, diluted and homogenized. The selection of resistant Gram negative bacteria was performed in VRBG plates containing ten different antibiotics separately. The isolates were identified through the amplification of the 16S rDNA. Antimicrobial susceptibility was assessed by disc diffusion method using EUCAST guidelines. All isolates were investigated for the presence of class 1, 2 and 3 integrons, through PCR amplification, and their variable region was explored by a strategy of Next Generation Sequencing. Results: A total of 333 isolates showing nonsusceptibility to, at least, one antibiotic were collected among 144 samples. Among others, Enterobactericeae (n=184), Moraxellaceae (n=88), Pseudomonadaceae (n=36), were detected. Nonsusceptibility was mainly identified among β-lactam antibiotics for Enterobactericeae recovered from conventionally produced samples (45.1%); overall, cefoxitin was the most ineffective antibiotic (19.6 % for organic and 26.6% for conventionally grown produce). The molecular screening for the integrase-encoding gene showed the presence of eight isolates with class 1 (2 Enterobacter, 2 Escherichia, 1 Morganella, 2 Acinetobacter and 1 Pseudomonas spp.), three isolates with class 2 (1 Escherichia and 2 Raoutella spp.), and one isolate with class 3 integrons (Klebsiella spp.). The integrons revealed a variety of gene cassettes conferring nonsusceptibility to different classes of antibiotics. Conclusion: Resistance was more frequent in the products grown in close contact with the soil. Besides environmental bacteria, many pathogenic agents were also detected. Lettuce was significantly associated with the existence of class 1 and 2 integrons regardless of the origin. Both organic and conventional produced fruits and vegetables may constitute potential sources of resistant bacteria and of integrons.