Quhao Wei

A novel functional class 2 integron in clinical Proteus mirabilis isolates

OBJECTIVES To describe a novel functional class 2 integron that was found in clinical Proteus mirabilis isolates. METHODS Class 1 and 2 integrons were screened by PCR in 153 clinical Proteus isolates. The variable regions of class 1 and 2 integrons were determined by restriction analysis and sequencing. The mutations of internal stop codons in class 2 integrons and their common promoters were also determined by sequencing. Enterobacterial repetitive intergenic consensus (ERIC)-PCR was used to analyse the phylogenetic relations of class 2 integron-positive P. mirabilis isolates. RESULTS Class 1 integrons were detected in 96 (63%) of 153 Proteus isolates: eight different gene cassette arrays were detected, including dfrA32-ereA1-aadA2, which was detected for the first time in P. mirabilis. Class 2 integrons were detected in 101 (66%) of 153 Proteus isolates: four different gene cassette arrays were detected, including dfrA1-catB2-sat2-aadA1, which was detected for the first time in a class 2 integron. A novel functional class 2 integron was detected in 38 P. mirabilis isolates with a common promoter (-35 TTTAAT|16 bp|-10 TAAAGT). The variable region of this functional class 2 integron contained dfrA14 and three novel open reading frames with unknown functions. Very similar ERIC-PCR fingerprinting patterns were detected in these 38 P. mirabilis isolates and were different from other class 2 integron-positive isolates. CONCLUSIONS A novel functional class 2 integron was found for the first time in P. mirabilis. These functional class 2 integron-harbouring P. mirabilis isolates were likely to be clonally spread in our hospital.

Transcription of integron-harboured gene cassette impacts integration efficiency in class 1 integron

Class 1 integrons play important roles in the dissemination of antibiotic resistance genes among bacteria. Generally, class 1 integron consists of an integrase gene (intI1), a recombination site (attI1) and a promoter (Pc) that drives the transcription of the downstreamed gene cassettes. Occasionally, there is a second promoter P2 downstream of the Pc promoter. Several Pc variants and Pc–P2 combinations have been defined and they display different transcription strengths, but the influence of the transcription of integron‐harboured gene cassette on the integration efficiency has never been comprehensively studied. In this study, the integration frequencies of gene cassettes into the attI1 sites that downstream of four different Pc variants as well as their combinations with P2 promoter were measured. The results showed that there was an inverse correlation between the strength of Pc promoter and the integration efficiency and, with the same Pc promoter, the integration efficiency was significantly decreased when a P2 promoter preceded the attI1 site. Our findings indicate there is a relationship between the transcription of integron‐harboured gene cassette and the integration of exogenous gene cassettes. The interrelationship between these two relatively independent processes may throw a light on our understanding the regulation system of class 1 integron.

A novel and rapid method for determining integration frequency catalyzed by integron integrase intI1

We developed a faster and more convenient method to determine integration frequency mediated by integron integrase intI1. This method based on real-time fluorescent quantitative PCR. By using this method, we revealed that the integration frequency of aadA2 gene cassette was 1.87x10(-4) when integrase intI1 was present, and the background frequency was less than 5.23x10(-8) without integrase intI1.