Effects of short interfering RNA against methicillin-resistant Staphylococcus aureus coagulase in vitro and in vivo.
OBJECTIVE
The emergence of antibiotic-resistant bacteria such as Staphylococcus aureus calls for inventive research and development strategies. Inhibition of bacterial pathogenesis may be a promising therapeutic approach in this regard. The gene-silencing effect of short interfering RNA (siRNA) is useful for this strategy. We investigated the efficacy of siRNA on the expression of coagulase because it is the one of the most important enzymes in the pathogenesis of methicillin-resistant S. aureus (MRSA) infection.


METHODS
We designed and synthesized 21 bp siRNA duplexes against staphylococcal coagulase. RT-PCR was performed to determine whether the siRNAs inhibit the expression of the coagulase mRNA and radio-labelled siRNA was used to confirm transfection to bacteria in vitro. The efficacy of siRNA was determined in a murine model of haematogenous pulmonary infection.


RESULTS
RT-PCR showed that siRNAs significantly inhibited the expression of the coagulase mRNA. The coagulase titres in the siRNA and control groups were 8 and 32, respectively. Measurement of incorporated radioactivity indicated that the siRNAs were delivered into the bacteria. In the murine infection model, in control and siRNA groups, 7.64 +/- 0.42 and 6.29 +/- 0.23 log cfu/mL (mean +/- SEM) MRSA were detected, respectively, showing that there was a significant decrease in the number of viable bacteria in the siRNA group (P < 0.05).


CONCLUSIONS
The results show that siRNA inhibited both mRNA expression and the activity of MRSA coagulase in vitro. The in vivo results revealed that the siRNA was effective in reducing the bacterial load in a murine model of haematogenous pulmonary infection. Targeting of coagulase with siRNA appears to be a novel strategy for treating MRSA infections.
Yanagihara
Katsunori
K
Second Department of Internal Medicine, Nagasaki University School of Medicine, Nagasaki, Japan.
Tashiro
Miwa
M
Fukuda
Yuichi
Y
Ohno
Hideaki
H
Higashiyama
Yasuhito
Y
Miyazaki
Yoshitsugu
Y
Hirakata
Yoichi
Y
Tomono
Kazunori
K
Mizuta
Yohei
Y
Tsukamoto
Kazuhiro
K
Kohno
Shigeru
S
D000818:Animals; D003030:Coagulase / Q000235:genetics*; D004195:Disease Models, Animal; D020868:Gene Silencing; D018390:Gene Targeting; D008297:Male; D016106:Methicillin Resistance / Q000235:genetics*; D051379:Mice; D011023:Pneumonia, Staphylococcal / Q000628:therapy*; D034741:RNA, Small Interfering / Q000235:genetics* / Q000494:pharmacology; D013211:Staphylococcus aureus / Q000201:enzymology* / Q000235:genetics
D003030:Coagulase; D034741:RNA, Small Interfering