Identification and molecular characterization of class 1 integrons in plasmid ?carrying bacterial isolates recovered from a biopurification system used for pesticide removal

VIETA, MP; MARTINI MC ; QUIROGA MP; CENTRÓN D; LAGARES A; DEL PAPA M. F.

Córdoba

Congreso; XI SAMIGE; 2015

SAMIGE

Biopurification systems (BPS) are complex soil-related and artificially-generated environments usually designed for the removal of toxic compounds from polluted wastewater sources. BPS receives high loads of pesticides and other toxic compounds, thus imposing a strong, constant and long-term selective pressure on the evolution and growth of the associated microbiome. Recently, we have isolated and characterized a collection of 35 plasmid-carrying bacterial isolates recovered from a BPS contaminated with more than 50 different pesticides. These cultivable bacteria were selected using a plasmid-screening based method, in order to study the role of plasmids in such environment. It is well known that mobile genetic elements (MGEs) play an essential role in the evolution and adaptation of bacteria as they encode accessory genes that increase bacterial fitness in a given environment. In order to go deeper in the analysis of MGEs, we decided to expand our study looking for the presence of type 1 integrons, which have been found in plasmid backbones. Integrons are assembly platforms that incorporate exogenous open reading frames harbouring attC sites by site-specific recombination and convert them to functional gene cassettes by ensuring their correct expression. In particular, class 1 integrons are widely distributed in the clinical setting as well as in natural environments. Their ability to acquire, exchange, and express antimicrobial resistance gene cassettes, makes them an important MGE to be studied. The aim of this work was to investigate the presence of class 1 integrons as well as their genetic content, in a plasmid-containing bacteria collection obtained from a model on-farm biofilter used for wastewater decontamination in intensive agricultural production. In order to evaluate the presence of class 1 integrons, we performed a PCR-based screening for detection of the integrase of type 1 (intI1) . Out of the screened isolates, 23% were positives for intI1, thus suggesting the presence of class 1 integrons in the collection. In a second step, we investigated the presence of genes that frequently constitute the 3´conserved fragment in this type of integrons. The screening revealed that all isolates that contain the intI1 gene posses also the sul1 and qacE genes, wich confer resistance to sulfonamides and quaternary ammonium compounds, respectively. Besides, 4 of these isolates were positive for qacE1, a qacE gene variant. In order to go deeper in the integron characterization, a second PCR-based screening was performed using primers that bind to the 5´and 3´conserved fragments, flanking the variable region. Out of the 8 integron-containing isolates, 3 of them gave a PCR fragment whose sequencing and analysis revealed the presence of a adenylyltransferase gene, which in all cases confers resistance to streptomycin. The analysis of MGE in soil bacteria is a critical issue for improving our current understanding of the mechanisms underlying microbial-genome dynamics.