Depression of biofilm formation and antibiotic resistance by sarA disruption in Staphylococcus epidermidis.
OBJECTIVE
To study the effects of disruption of sarA gene on biofilm formation and antibiotic resistance of Staphylococcus epidermidis (S. epidermidis).


METHODS
In order to disrupt sarA gene, the double-crossover homologous recombination was applied in S. epidermidis RP62A, and tetracycline resistance gene (tet) was used as the selective marker which was amplified by PCR from the pBR322 and inserted into the locus between sarA upstream and downstream, resulting in pBT2delta sarA. By electroporation, the plasmid pBT2delta sarA was transformed into S. epidermidis. Gene transcription was detected by real-time reverse transcription-PCR (RT-PCR). Determination of biofilm was performed in 96-well flat-bottomed culture plates, and antibiotic resistance was analyzed with test tube culture by spectrophotometry at 570 nm respectively.


RESULTS
A sarA disrupted strain named S. epidermidis RP62Adelta sarA was constructed, which was completely defective in biofilm formation, while the sarA complement strain RP62Adelta sarA (pHPS9sarA) restored the biofilm formation phenotype. Additionally, the knockout of sarA resulted in decreased erythromycin and kanamycin resistance of S. epidermidis RP62A. Compared to the original strain, S. epidermidis RP62Adelta sarA had an increase of the sensitivity to erythromycin at 200-400 microg/mL and kanamycin at 200-800 microg/mL respectively.


CONCLUSIONS
The knockout of sarA can result in the defect in biofilm formation and the decreased erythromycin and kanamycin resistance in S. epidermidis RP62A.
Tao
Ju-Hong
JH
Department of Microbiology, School of Life Sciences, Fudan University, 220 Han Dan Road, Shanghai 200433, China.
Fan
Chang-Sheng
CS
Gao
Shan-E
SE
Wang
Hai-Jiao
HJ
Liang
Guo-Xin
GX
Zhang
Qing
Q
D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology*; D018441:Biofilms / Q000254:growth & development*; D002876:Chromosomes, Bacterial; D024881:Drug Resistance, Bacterial / Q000235:genetics; D017353:Gene Deletion; D020868:Gene Silencing; D005898:Glass; D007613:Kanamycin Resistance / Q000235:genetics*; D009876:Operon; D010641:Phenotype; D013212:Staphylococcus epidermidis / Q000235:genetics* / Q000254:growth & development; D015534:Trans-Activators / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; C088021:SarA protein, bacterial; D015534:Trans-Activators