Widespread dissemination of class 1 integron components in soils and related ecosystems as revealed by cultivation-independent analysis

Abstract

Class 1 integrons contribute to the emerging problem of antibiotic resistance in human medicine by acquisition, exchange, and expression of resistance genes embedded within gene cassettes. Besides the clinical setting they were recently reported from environmental habitats and often located on plasmids and transposons, facilitating their transfer and spread within bacterial communities. In this study we aimed to provide insights into the occurrence of genes typically associated with the class 1 integrons in previously not studied environments with or without human impact and their association with IncP-1 plasmids. Total community DNA was extracted from manure-treated and untreated soils, lettuce and potato rhizosphere, digestates, and an on-farm biopurification system and screened by PCR with subsequent Southern blot hybridization for the presence of the class 1 integrase gene intI1 as well as qacE and qacEΔ 1 resistance genes. The results revealed a widespread dissemination of class 1 integrons in the environments analyzed, mainly related to the presence of qacEΔ 1 genes. All 28 IncP-1ε plasmids carrying class 1 integrons, which were captured exogenously in a recent study from piggery manure and soils treated with manure, carried qacEΔ 1 genes. Based on the strong hybridization signals in the rhizosphere of lettuce compared to the potato rhizosphere, the abundances of intI1, qacE/qacEΔ 1, and sul1 genes were quantified relative to the 16S rRNA gene abundance by real-time PCR in the rhizosphere of lettuce planted in three different soils and in the corresponding bulk soil. A significant enrichment of intI1 and qacE/qacEΔ 1 genes was confirmed in the rhizosphere of lettuce compared to bulk soil. Additionally, the relative abundance of korB genes specific for IncP-1 plasmids was enriched in the rhizosphere and correlated to the intI1 gene abundance indicating that IncP-1 plasmids might have contributed to the spread of class 1 integrons in the analyzed soils.

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

“…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).…”

“…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). Class 1 integrons have been extensively studied due to their widespread distribution in Gram-negative bacteria of clinical importance (Stalder et al 2012), and the intI1 genes were reported to be very common in soils and related ecosystems (Gillings et al 2008;Jechalke et al 2014c).…”

“…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).…”

“…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). However, the rate of horizontal transmission in soil was assumed to be very low, compared with the vertical transmission of ARGs due to growth of resistant microorganisms (Heuer et al 2011b).…”

Microbial Communities in the Rhizosphere Analyzed by Cultivation-Independent DNA-Based Methods

“…We could show that the abundance of sulfonamide resistance genes (sul1, sul2) was unexpectedly lower in the rhizosphere of maize and grass grown in manure-treated soils compared to control. Another interesting observation was the enrichment of class 1 integrons and IncP-1 plasmids in the rhizosphere of lettuce grown in three soils (Jechalke et al 2014). This increased abundance might be caused by aromatic compounds in the root exudates of lettuce (Neumann et al 2014).…”