HORIZONTAL TRANSFERENCE OF ANTIMICROBIAL RESISTANCE GENES BETWEEN A POSITIVE AND A NEGATIVE CLASS 1 INTEGRON STRAIN OF ESCHERICHIA COLI

PÉREZ GAUDIO D.S., COLELLO R., FERNÁNDEZ D., ETCHEVERRÍA A., PADOLA N.L., SORACI A.L.

Atlanta, Georgia

Congreso; 3rd International Congress on Microbiology & Pharmaceutical Microbiology; 2017

Pulsus Group

Statement of the Problem: A consequence of an intensive and inappropriateuse of antimicrobials in veterinary medicine is the spreading of antibiotic resistance, which implies a problem for public health due to the potential risk of resistance transference through the food chain by bacteria such as E. coli (2; 5). The participation of integrons has been studied in recent years (2; 4). They are gene-capture and expression systems characterized by the presence of an int1 gene encoding an integrase (Int1), a recombination site (attI), and a promoter. The most frequently reported mobile integrons are class 1 and class 2, which have been shown to contribute to the spread of antimicrobial resistance genes (3). Methodology & Theoretical Orientation: A conjugation assay was performed with two E. coli strains to determine the horizontal transference of int1 from an antibiotic resistant bacteria (both sorbitol and int1 positive), isolated from a pig farm, to a pathogenic bacteria (both sorbitol and int1 negative, MV-VO-A O157 strain). Each strain was cultured separately for 18 h at 37°C and agitated at100 rpm in 30 mL of LB broth. To perform the co-culture, 10 μL of each strain were placed at 37°C and agitated at100 rpm in 20 mL of LB broth. At 0.5; 2 and 4 h aliquots of the co-culture were seeded on McConkey sorbitol agar. Dilutions were performed for the different times, in order to distinguish colonies without difficulty. The sorbitol positive (red) and sorbitol negative (white) colonies were identified, spiked into tubes and allowed to grow in 800 μL of LB broth. Then, DNA was extracted to perform PCR and to evaluate the presence or absence of intl1. The amplified products were visualized by 2% agarose electrophoresis gel in TBE buffer containing SyBR Safe. Samples (10 μL) were run for 30 min, at 90 V and visualized with UV illuminator. Molecular weight markers were included in each gel. Findings: Figure 1 shows the gel corresponding to 14 samples of initially int1 negative E. coli colonies. Note that at the 4 h of assay, six out of nine repetitions acquired the int1 (892 bp) and they probably had become resistant to antimicrobials. Conclusion & Significance: On a previous work of our group it was demonstrated that commensal E. coli isolated from farm sows, harbored the genes for Int1 and/or 2. It was also confirmed the presence of integrons in strains isolated from piglets younger than 12 h of age suggesting the importance of the transmission of resistant strains at birth (1). On this work we have proven the transference of integrons from an E. coli resistant strain, isolated from the mentioned swine farm, to a negative and pathogenic one, in a period of only 4 h. This fact is extremely important considering our previous findings on pig farms. As it was previously shown animals and farm environment can act as reservoirs for potential spread of resistant bacteria by means of integrons, allowing negative int1 bacteria (probably pathogenic) to become resistant to antibiotics in a very short period of time, constituting a problem for public health, due to the dilemma that it means for the future treatment of such bacterial infections.