摘要
目的了解烧伤临床鲍氏不动杆菌分离株生物膜形成过程中，密度感应基因aba I表达变化及其对生物膜ECM和细菌耐药性的影响。 方法2011年1-10月，收集上海交通大学医学院附属瑞金医院烧伤住院患者创面、血液和静脉导管来源的鲍氏不动杆菌耐药菌6株、敏感菌5株。以鲍氏不动杆菌标准菌株ATCC 19606为参照。（1）临床分离株和标准菌株体外常规培养10、24、48 h后，经碘化丙啶染色，用激光扫描共聚焦显微镜观察生物膜厚度。（2）采用试管培养法，将临床分离株和标准菌株振摇培养10、24、48 h，培养液内漂浮细菌为游离菌，黏附于试管壁的生物膜内细菌为膜内菌。采用实时荧光定量PCR技术检测各菌株中aba I基因及pgaB基因的相对表达量（标准菌株基因表达丰度设为1）。对实验数据进行方差分析。 结果 （1）鲍氏不动杆菌临床分离株体外培养10、24、48 h，生物膜厚度均超过标准菌株，其中耐药菌株生物膜厚度分别为（28.8±0.6）、（ 31.7±1.1）、（38.1 ＋3.l）pLm，明显高于敏感菌株[（17.1±0.4）、（20.1±1.6）、（ 25.8±1.7） pLm，F值分别为1274. 38、206. 60，61. 73，P值均小于0.05]。（2）膜内菌：体外培养10、24、48 h，耐药菌株abaI表达量分别为6.6 ＋1.7、25.7±3.5、9.8＋3.6，均高于敏感菌株（2.7＋1．O、15．O＋3.5、4.7 ＋3.2，F值分别为21. 82、25. 24、6.22，P值均小于0.05）；耐药菌株pgaB的表达量分别为37.4±1.1、44.5±3.6、33.1 ＋11.5，明显高于敏感菌株（14.6 ＋0.8、20．O＋6.9、18.7 ＋6.8，F值分别为1488. 44、57. 26、6.01，P值均小于0.05）。（3）游离菌：体外培养10、24、48 h，耐药菌株aba I表达量均与敏感菌株接近（F值分别为0. 24、2.33、0.11，P值均大于0.05）；耐药菌株pgaB表达量分别为13.8±3.8、12.5±2.9、23.7±2.1，明显高于敏感菌株（7．O＋5.9、5．O＋1.3、15.6±6.7，F值分别为5.44、28. 42、7.76，P值均小于0. 05）。（4）膜内菌与游离菌的耐药菌株比较：膜内菌各时相点abaI表达量均多于游离菌（F值分别为43. 69、286. 61、9.98．P值均小于0.05），膜内菌pgaB表达量在10、24 h多于游离菌（F值分别为214. 26、283. 20，P值均小于0.05）。（5）膜内菌与游离菌的敏感菌株比较：膜内菌aba I表达量在培养24 h多于游离菌（F=70.28，P〈0.05），pgaB表达量在10、24 h多于游离菌（F值分别为8. 03、22. 62，P值均小于0.05）。 结论烧伤临床鲍氏不动杆菌分离株生物膜形成过程中，耐药菌株的密度感应基因aba I表达明显增多，可能引起pgaB基因表达上调，导致细菌ECM及生物膜形成增加、耐药性增强。
Objective To stud,, the Acinetubacter bal^mannii （ AB ） strains isolated influences on the extracellular matrix of biofilm and five drug-sensitive AB strains isolated from changes in expression of quorum sensing gene aba I in from burn patients during t）iofilm fl）rmation process, and its and drug resistance of AB. Methods Six drug-resistant wound excretion, bhn）d and venous catheter were collected from burn patients hospitalized in Ruijin hospital of Shanghai Jiao Tong University Sc：hool of Medicine fromJanuary to October 2011. The AB standard strain ATCC 19606 was used as control. （ 1） Clinical strains andstandard strain were normally cultured 10, 24, and 48 h respectively in vitro. The bacteria samples were
stained with propidium iodide to measure biofilm thickness with confocal laser scanning microsc：ope. （2）
Clinical strains and standard strain were cultured in tubes 10, 24, and 48 h respectively in vitro under sha-
king condition. The bacteria floacing in the medium were regarded as free bacteria, while those adhered to
the cube wall as the hacteria within hiofilm （ biofilm bacteria） . Relacive expression value of genes aba I and
pgaB was detected hy real-time fluorescent quantitaLive PCR with the expression value of the standard strain
set at l. Data were processed with analysis of variance. Results （1） At post culture hour （PCH） 10,
24, 48, biofilm thic.kness of clinical strains was chicker than that of standard strain; biofilm thickness of
drug-resistant strains [ （ 28. 8 ＋0. 6） , （31. 7 ＋ 1. 1 ） , and （38. 1 ＋3. 1 ） ym ] was respectively thicker than
that of drug-sensitive strains [ （ 17. 1 ＋0. 4） , （ 20. 1 ＋ 1. 6） , and （ 25. 8 ＋ 1. 7 ） ym, with F value respec-
tively 1274. 38 , 206. 60 , and 61. 73 , P values all below 0. 05 ] . （2） Biofilm bacteria： at PCH 10 , 24 , 48 ,
expression values of aba I in drug-resistant strains （ 6. 6 ＋ 1. 7 , 25. 7 ＋3. 5 , 9. 8 ＋3. 6） were much higher
than those of drug-sensicive strains （2. 7 ＋ 1. 0 , 15. 0 ＋ 3. 5 , 4. 7 ＋ 3. 2 , with F value respectively 21. 82 ,
25. 24, and 6. 22, P values all below 0. 05 ） ; expression values of pgaB in drug-resisLant strains （37. 4 ＋
1. 1, 44. 5 ＋3. 6, 33. 1 ＋ 11. 5 ） were ohviously higher than those of drug-sensitive strains （ 14. 6 ＋ 0. 8,
20. 0 ＋ 6. 9 , 18. 7 ＋ 6. 8 , with F value respectively 1488. 44 , 57. 26 , and 6. 01 , P values all below 0. 05 ） .
（ 3 ） Free bacteria： at PCH 10, 24 , 48 , chere was no significant stacistical differenc：e between drug-resistant
strains and drug-sensitive strains in expression value ofaba I （ with F value respectively 0. 24, 2. 33 , and
0. 11 , P values all above 0. 05 ） ; expression values of pgaB in drug-resistant strains （ 13. 8 ＋ 3. 8 , 12. 5 ＋
2. 9, 23. 7 ＋ 2. 1 ） were obviously higher than those of drug-sensitive strains （7. 0 ＋5. 9 , 5. 0 ＋ 1. 3 , 15. 6 ＋
6. 7 , with F value respectively 5. 44 , 28. 42 , and 7. 76 , P values all below 0. 05 ） . （4） Comparison between
biofilm hacteria and free bacteria in resistant strains： expression value of aba I in biof＇ilm bacteria at each
time point was respectively higher than that of free bacteria （ with F value respectively 43. 69, 286.61 , and
9.98 , P values all below 0. 05） ; expression values of pgaB in biofilm bacteria at PCH 10, 24 were higher
than those in free hacteria （ with F value respectively 214. 26 and 283. 20 , P values below 0. 05 ） . （ 5 ） Com-
parison between biofilm hacteria and free bacteria in sensitive strains： expression value ofaba I in BF bac-
teria at PCH 24 was higher than that of free bacteria （ F = 70.28 , P 〈 0. 05） ; expression values of pgaB in
biofilm hacteria at PCH 10, 24 were higher than those of free bacteria （ with F value respectively 8. 03 and
22. 62, P values below 0. 05 ） . Conclusions During hiofilm formation process, the increasing expression
of quorum sensing gene aba I in drug-resistant AB strains isolated from burn patients may up-regulate che
expression of gene pgaB, which leads to high produc.tion of extracellular matrix and biofilm formation, and
enhanc：es drug resistance of AB.
出处
《中华烧伤杂志》
CAS
CSCD
北大核心
2012年第2期101-105,共5页
Chinese Journal of Burns
基金
基金项目：上海市科学技术委员会自然科学基金（11ZR1422300）
关键词
烧伤
感染
鲍氏不动杆菌
生物膜
抗药性
aba
I基因
Burns
Infec：tion
ilcin,eiobaUer baum,an,n,ii
Biofilms
Drug resistanc：e
Ceneaba I
