中华微生物学和免疫学杂志
中華微生物學和免疫學雜誌
중화미생물학화면역학잡지
CHINESE JOURNAL OF MICROBIOLOGY AND IMMUNOLOGY
2012年
1期
1-5
,共5页
eDNA%表皮葡萄球菌%细菌生物被膜%微量板静置培养法
eDNA%錶皮葡萄毬菌%細菌生物被膜%微量闆靜置培養法
eDNA%표피포도구균%세균생물피막%미량판정치배양법
eDNA%Staphylococccus epidermidis%Bacterial biofilm%Microtitre plate static culture
目的 检测表皮葡萄球菌(表葡菌)临床株生物被膜的形成能力,研究胞外DNA(eDNA)水平差异对表葡菌临床株生物被膜形成能力的影响.方法 收集227株临床分离表葡菌,黏附试验检测其生物被膜的形成能力,PCR方法扩增icaA基因片段,黏附试验阳性和icaA基因扩增阳性的表葡菌为成膜组,黏附试验阴性和icaA基因扩增阴性的表葡菌为非成膜组,应用浮游培养法和微量板静置培养法检测eDNA水平,激光共聚焦显微镜(CLSM)观察生物被膜中eDNA的分布.结果 227株表葡菌中黏附试验阳性26株,icaA基因阳性32株.选取黏附试验阳性和icaA基因扩增阳性的成膜组表葡菌20株,黏附试验阴性和icaA基因扩增阴性的非成膜组表葡菌19株,浮游培养2、4、6h后,成膜组菌株的eDNA水平分别为(32.2±10.1) μg/ml、(33.6±11.9) μg/ml、(34.3±10.0)μg/ml,非成膜组菌株的eDNA水平分别为(28.7±8.9)μg/ml、(31.5±11.7) μg/ml、(31.8±l2.7)μg/ml,浮游培养成膜组菌株各时相eDNA水平分别高于非成膜组菌株,但差异尚无显著性(P＞0.05).微量板静置培养法成膜组20株eDNA水平为(740.0±264.4) ng/A600,高于非成膜组19株eDNA水平(80.1±31.1) ng,/A600,两组之间比较差异具有显著性(P＜0.05).通过AO-PI荧光染色于CLSM下观察静置培养的成膜株表葡菌Y36,其生物被膜中可见eDNA分布；非成膜株Y26没有生物被膜结构形成,未见eDNA分布.结论 表葡菌临床株具有形成生物被膜的能力,eDNA是表葡菌形成生物被膜的重要基质成分,在判断表葡菌生物被膜形成能力方面微量板静置培养法检测eDNA水平优于浮游培养法.
目的 檢測錶皮葡萄毬菌(錶葡菌)臨床株生物被膜的形成能力,研究胞外DNA(eDNA)水平差異對錶葡菌臨床株生物被膜形成能力的影響.方法 收集227株臨床分離錶葡菌,黏附試驗檢測其生物被膜的形成能力,PCR方法擴增icaA基因片段,黏附試驗暘性和icaA基因擴增暘性的錶葡菌為成膜組,黏附試驗陰性和icaA基因擴增陰性的錶葡菌為非成膜組,應用浮遊培養法和微量闆靜置培養法檢測eDNA水平,激光共聚焦顯微鏡(CLSM)觀察生物被膜中eDNA的分佈.結果 227株錶葡菌中黏附試驗暘性26株,icaA基因暘性32株.選取黏附試驗暘性和icaA基因擴增暘性的成膜組錶葡菌20株,黏附試驗陰性和icaA基因擴增陰性的非成膜組錶葡菌19株,浮遊培養2、4、6h後,成膜組菌株的eDNA水平分彆為(32.2±10.1) μg/ml、(33.6±11.9) μg/ml、(34.3±10.0)μg/ml,非成膜組菌株的eDNA水平分彆為(28.7±8.9)μg/ml、(31.5±11.7) μg/ml、(31.8±l2.7)μg/ml,浮遊培養成膜組菌株各時相eDNA水平分彆高于非成膜組菌株,但差異尚無顯著性(P＞0.05).微量闆靜置培養法成膜組20株eDNA水平為(740.0±264.4) ng/A600,高于非成膜組19株eDNA水平(80.1±31.1) ng,/A600,兩組之間比較差異具有顯著性(P＜0.05).通過AO-PI熒光染色于CLSM下觀察靜置培養的成膜株錶葡菌Y36,其生物被膜中可見eDNA分佈；非成膜株Y26沒有生物被膜結構形成,未見eDNA分佈.結論 錶葡菌臨床株具有形成生物被膜的能力,eDNA是錶葡菌形成生物被膜的重要基質成分,在判斷錶葡菌生物被膜形成能力方麵微量闆靜置培養法檢測eDNA水平優于浮遊培養法.
목적 검측표피포도구균(표포균)림상주생물피막적형성능력,연구포외DNA(eDNA)수평차이대표포균림상주생물피막형성능력적영향.방법 수집227주림상분리표포균,점부시험검측기생물피막적형성능력,PCR방법확증icaA기인편단,점부시험양성화icaA기인확증양성적표포균위성막조,점부시험음성화icaA기인확증음성적표포균위비성막조,응용부유배양법화미량판정치배양법검측eDNA수평,격광공취초현미경(CLSM)관찰생물피막중eDNA적분포.결과 227주표포균중점부시험양성26주,icaA기인양성32주.선취점부시험양성화icaA기인확증양성적성막조표포균20주,점부시험음성화icaA기인확증음성적비성막조표포균19주,부유배양2、4、6h후,성막조균주적eDNA수평분별위(32.2±10.1) μg/ml、(33.6±11.9) μg/ml、(34.3±10.0)μg/ml,비성막조균주적eDNA수평분별위(28.7±8.9)μg/ml、(31.5±11.7) μg/ml、(31.8±l2.7)μg/ml,부유배양성막조균주각시상eDNA수평분별고우비성막조균주,단차이상무현저성(P＞0.05).미량판정치배양법성막조20주eDNA수평위(740.0±264.4) ng/A600,고우비성막조19주eDNA수평(80.1±31.1) ng,/A600,량조지간비교차이구유현저성(P＜0.05).통과AO-PI형광염색우CLSM하관찰정치배양적성막주표포균Y36,기생물피막중가견eDNA분포；비성막주Y26몰유생물피막결구형성,미견eDNA분포.결론 표포균림상주구유형성생물피막적능력,eDNA시표포균형성생물피막적중요기질성분,재판단표포균생물피막형성능력방면미량판정치배양법검측eDNA수평우우부유배양법.
Objective To determine the ability of biofilm formation of Staphylococcus epidermidis isolates and study the influence of different extracellular DNA(eDNA) levels in S.epidermidis isolates on the ability of biofilm formation.Methods Detect the biofilm-formation ability of 227 S.epidermidis isolates with adhesion assays,amplify the icaA gene fragment with PCR.The S.epidermidis isolates were divided into biofilm formation(BF) group and non-biofilm formation (NBF) group according to adhesion assays and icaA amplification.Detect eDNA levels of S.epidermidis in planktonic culture and microtitre plate static culture.The eDNA in S.epidermidis biofilms stained by AO-PI was observed by CLSM.Results 26 isolates were positive in adhesion assays and 32 isolates existed icaA gene among 227 S.epidermidis isolates.Select 20 isolates with positive adhesion assays and positive icaA amplification for BF group.Select 19 isolates with negative adhesion assays and negative icaA amplification for NBF group.The eDNA levels were (32.2±10.1)μg/ml,(33.6±11.9) μg/ml,(34.3±10.0) μg/ml in BF group when cultured in planktonic condition for 2,4,6 h,while the eDNA levels in NBF group were (28.7±8.9) μg/ml,(31.5±11.7) μg/ml,(31.8±12.7) μg/ml respectively.There were no significant differences between the two groups for these three phases(P＞0.05),though the eDNA levels of BF group were higher than that of NBF group.The eDNA levels were (740.0±264.4) ng/A600 in BF group when cultured in static microtitre plate,higher than that of NBF group,(80.1 ±31.1) ng/A600,and the difference between these two groups was significant.The eDNA in BF isolate Y36 biofilms could be visualized by staining with AO and PI when observed by CLSM,while neither biofilm structure nor eDNA appeard when NBF isolate Y26 was cultured for 24 h.Conclusion S.epidermidis isolates have the ability of biofilm formation.eDNA is one of the important matrix components in the S.epidermidis biofilm-forming process.The eDNA of static culture in microtitre plate was more efficient than planktonic culture in the case of estimating the ability of biofilm formation of S.epidermidis.
