IncP-1ε Plasmids are Important Vectors of Antibiotic Resistance Genes in Agricultural Systems: Diversification Driven by Class 1 Integron Gene Cassettes

Abstract

The role of broad-host range IncP-1ε plasmids in the dissemination of antibiotic resistance in agricultural systems has not yet been investigated. These plasmids were detected in total DNA from all of 16 manure samples and in arable soil based on a novel 5′-nuclease assay for real-time PCR. A correlation between IncP-1ε plasmid abundance and antibiotic usage was revealed. In a soil microcosm experiment the abundance of IncP-1ε plasmids was significantly increased even 127 days after application of manure containing the antibiotic compound sulfadiazine, compared to soil receiving only manure, only sulfadiazine, or water. Fifty IncP-1ε plasmids that were captured in E. coli CV601gfp from bacterial communities of manure and arable soil were characterized by PCR and hybridization. All plasmids carried class 1 integrons with highly varying sizes of the gene cassette region and the sul1 gene. Three IncP-1ε plasmids captured from soil bacteria and one from manure were completely sequenced. The backbones were nearly identical to that of the previously described IncP-1ε plasmid pKJK5. The plasmids differed mainly in the composition of a Tn402-like transposon carrying a class 1 integron with varying gene cassettes, IS1326, and in three of the plasmids the tetracycline resistance transposon Tn1721 with various truncations. Diverse Beta- and Gammaproteobacteria were revealed as hosts of one of the IncP-1ε plasmids in soil microcosms. Our data suggest that IncP-1ε plasmids are important vectors for horizontal transfer of antibiotic resistance in agricultural systems.

Broad host range plasmids can invade an unexpectedly diverse fraction of a soil bacterial community

“…Broad host range plasmids of the IncP-1 and IncPromA group can thus provide a direct link between diverse bacterial groups. Especially, IncP1e plasmids such as pKJK5 have been identified as vectors of antibiotic resistance genes transfer among Proteobacteria by additionally hosting class 1 integron gene cassettes (Heuer et al, 2012). These class 1 integrons may not only spread in their originally identified Gram-negative Enterobacteriaceae hosts but can also be found among many Gram-positive bacteria (Nandi et al, 2004).…”

Quaternary ammonium compounds in soil: implications for antibiotic resistance development

“…Class 1 integrons including qac genes are frequently found on IncP-1 plasmids which are broad host range plasmids widely distributed in the environment such as soil, manure, sludge, digestates and river sediments (Jechalke et al 2015;Smalla et al 2006;Zhang et al 2011b;Heuer et al 2012;Popowska and Krawczyk-Balska 2013;Wolters et al 2015). For example, IncP-1e plasmids were detected by qPCR in 16 manure samples and in arable soil (Heuer et al 2012). Fifty IncP-1 plasmids were further analyzed and all carried class 1 integrons.…”

“…These plasmids and transposons include multiresistance and heavy metal resistance plasmids as, for example reported for qacA and qacB genes, respectively (Russell 1997), but they were also associated with degradative pathways in a pesticide polluted environment (Dealtry et al 2014). Class 1 integrons including qac genes are frequently found on IncP-1 plasmids which are broad host range plasmids widely distributed in the environment such as soil, manure, sludge, digestates and river sediments (Jechalke et al 2015;Smalla et al 2006;Zhang et al 2011b;Heuer et al 2012;Popowska and Krawczyk-Balska 2013;Wolters et al 2015). For example, IncP-1e plasmids were detected by qPCR in 16 manure samples and in arable soil (Heuer et al 2012).…”

Temporal changes of antibiotic-resistance genes and bacterial communities in two contrasting soils treated with cattle manure

“…In addition, manuring was shown to even increase the permissiveness of the soil community for receiving and maintaining broad-host-range plasmids (Jechalke et al 2014b). Integrons possess a site specific recombination system that could capture and express mobile gene cassettes (Heuer, Schmitt and Smalla 2011a), and they were reported to often localize in broad-host-range IncP-1ε plasmids with a wide distribution in agricultural systems that further facilitates their mobility potential (Heuer et al 2012;Jechalke et al 2014c;Wolters et al 2015). The critical roles of integrons in the dissemination of ARGs in environments have been highlighted by cultivation methods and metagenomics approaches (Hardwick et al 2008;Stalder et al 2012;Gillings et al 2015).…”

“…5) suggested an increased horizontal transfer potential of these ARGs. In particular, the potential for horizontal gene transfer would be higher when the class 1 integrons are located in the transferable IncP-1ε plasmids or other mobility elements (Heuer et al 2012;Jechalke et al 2014c;Wolters et al 2015). The class 1 integrons are also important for co-selection and mobilization of other ARGs (apart from β-lactamases), especially when selective pressure by antibiotics is present (Jechalke et al 2014c).…”