1、传统的药敏实验传统的药敏实验 S J Kim, The Union药药敏敏试验试验的目的的目的/药敏试验的校正药敏试验的校正获获得最相得最相关关的的临临床耐床耐药标药标准准 可能的敏感株的敏感可能的敏感株的敏感级别级别在治在治疗疗反反应变应变化方面化方面测测定敏感性水平定敏感性水平从从未接受未接受过过治治疗疗的患者的患者(PS)与与至少接受至少接受5/6个个月治月治疗疗的患的患者者(PR)的的标标本菌株的敏感性本菌株的敏感性进进行比行比较较 PS = 100 菌株菌株; PR = 50 菌株菌株Cantti G, et al. Bull WHO 1969; 41:21-43.可能敏感菌株可能敏
2、感菌株/ /(可能无耐药性菌株)在不同药(可能无耐药性菌株)在不同药物浓度下的抑菌分布物浓度下的抑菌分布Minimal inhibitory concentrations (MIC)最小抑菌最小抑菌浓浓度度Frequency of strainsClinicallysusceptiblestrains (PS)Breakpoint1 SD3 SDFrequency of strainsMinimal inhibitory concentrations (MIC)Clinicallysusceptiblestrains (PS)Clinicallyresistantstrains (PR)不同不
3、同药药物物浓浓度下可能敏感菌株和可能耐度下可能敏感菌株和可能耐药药菌株最低抑菌菌株最低抑菌浓浓度的分布度的分布Frequency of strainsMinimal inhibitory concentrations (MIC)PSPR2 mcg/mlBreakpoint13.1 as “R”75.811.1 as “S”可能敏感可能敏感/可能耐可能耐药药菌株菌株对对利福平的最小抑菌利福平的最小抑菌浓浓度的累度的累积积分布分布(此(此试验试验采用采用 L-J 培培养养基用基用绝对浓绝对浓度法)度法)85.8Frequency of strainsMinimal inhibitory conce
4、ntrations (MIC)Clinicallysusceptiblestrains (PS)Clinicallyresistantstrains (PR)采用采用 L-J 培培养养基基Do not quote this data!(n=81) (n=111) 校正的乙校正的乙胺胺丁醇的丁醇的临临界界浓浓度度(此试验采用改良罗氏培养基的绝对浓度法)42%50%In L-J MediumIn 7H10 Medium67%67%抗抗结结核核药药物的最低抑菌物的最低抑菌浓浓度比平均峰度比平均峰值值EMBETHOFXCFXINHRMPKMCPMPASSM推荐的临临界浓浓度药药物LJ*7H107H11
5、Bactec 460MGIT 960异异烟肼肼0.20.20.20.10.1利福平40.01.01.02.01.0乙胺胺丁醇2.05.07.52.55.0链链霉素4.02.02.02.01.0吡嗪酰胺吡嗪酰胺NRNRNR100.0100.0卡那霉素30.05.06.04.0阿米卡星40.01.01.0卷曲霉素40.010.010.01.252.5环环丙沙星2.02.02.02.01.0氧氟氧氟沙星2.02.02.02.02.0莫西沙星0.5 0.25 加替沙星1.0乙硫异酰胺异酰胺5.010.02.55.0丙硫异异烟胺胺40.01.252.5PAS1.02.08.02.0环丝酰胺环丝酰胺40.
6、0NRNRNRNR氨苯氨苯硫脲脲NRNRNRNRNR利奈唑酮唑酮1.01.0*Proportion WHO推荐推荐药药敏敏试验试验的的临临界界浓浓度度, 2007(4) 浓缩过程中利福平的稳定性浓缩过程中利福平的稳定性抗结核药物的储存抗结核药物的储存在密封的硅在密封的硅胶胶容器容器内储内储存存(最好在最好在真真空下空下储储存存)比例法比例法 (by Middlebrook & Cohn) Drug free Isoniazid Ethambutol Inoculum media 0.2 mcg/ml 2.0 mcg/ml 10-3 + + 3 10-5 26 9 0 % resistant 3
7、4.6 0.1Interpretation Resistant Susceptible药药敏敏试验结试验结果果-11 . If use nichrome wire (0.7 mm wire/ 3 mm of internal diameter), inoculate oneloopful of bacterial suspension of 10-2 / or if use pipette, inoculate 0.1 ml of 10-3 / of bacterial suspension2. If use nichrome wire, inoculate one loopful of 10
8、-4 / bacterial suspension or if use pipette, inoculate 0.1 ml of 10-5 /.DrugNoneINHRIFSMEMBPNBConcentrations(/)00.240.04.02.0500.0Tube NoC1H1R1S1E1PBInoculum (10-4 )13+2+11870Tube NoC2H2R2S2E2Inoculum (10-6 )2282000InterpretationR (7.1) (2/28x100)S (0.03)(1/2800 x100)S (0.6)(18/2800 x100)S (0.3)(7/2
9、800 x100)MTB药药敏敏试验结试验结果果-21 . If use nichrome wire (0.7 mm wire/ 3 mm of internal diameter), inoculate oneloopful of bacterial suspension of 10-2 / or if use pipette, inoculate 0.1 ml of 10-3 / of bacterial suspension2. If use nichrome wire, inoculate one loopful of 10-4 / bacterial suspension or if
10、 use pipette, inoculate 0.1 ml of 10-5 /.DrugNoneINHRIFSMEMBPNBConcentrations(/)00.240.04.02.0500.0Tube NoC1H1R1S1E1PBInoculum (10-4 )14+2+21+3+0Tube NoC2H2R2S2E2Inoculum (10-6 )22+50010Interpretation?S?MTB药药敏敏试验结试验结果果-31 . If use nichrome wire (0.7 mm wire/ 3 mm of internal diameter), inoculate one
11、loopful of bacterial suspension of 10-2 / or if use pipette, inoculate 0.1 ml of 10-3 / of bacterial suspension2. If use nichrome wire, inoculate one loopful of 10-4 / bacterial suspension or if use pipette, inoculate 0.1 ml of 10-5 /.DrugNoneINHRIFSMEMBPNBConcentrations(/)00.240.04.02.0500.0Tube No
12、C1H1R1S1E1PBInoculum (10-4 )13515000Tube NoC2H2R2S2E2Inoculum (10-6 )200000InterpretationRR?MTB 17个个跨跨国参国参比比实验实验室室网络网络第第14轮轮熟熟练练度度测试结测试结果果(数数据据来来源于源于Armand Van Deun 2007年)年)敏感性敏感性特特异异性性效能效能可重可重复复性性Isoniazid异异烟烟肼肼S敏感敏感=13R耐耐药药=17Average (%)99999998100%1314111495%4313Rifampicin利福平利福平S敏感敏感=10R耐耐药药=17Av
13、erage (%)99999998100%1414121495%3323StreptomycinS敏感敏感=11R耐耐药药=16Average (%)97919594100%12117995%5688EthambutolS敏感敏感=17R耐耐药药=4Average (%)88979697100%111381195%6476蓝色数字表示实验室数量蓝色数字表示实验室数量 51个国个国家(或地家(或地区区)参参比比实验实验室利福平和室利福平和异异烟烟肼药肼药敏敏试验结试验结果的一致性果的一致性(世界(世界卫卫生生组织组织第四期全球耐第四期全球耐药报药报告,告,2008)% Agreement一致性一
14、致性Isoniazid异异烟烟肼肼Rifampicin利福平利福平10038 (77.6)37 (72.5)95-996 (12.2)9 (17.6)90-944 (8.2)5 (9.8)901 (2.0)0Total number of labs 合合计计49 (100.0)51 (100.0)TestDrug药药物敏感性(%)特异异性(%)患病情况况下的耐药结药结果预测值预测值(%)1%15%110%130%1KM2100909.234.552.681.1CPM21009620.056.873.591.5OFX21009725.064.178.793.5ETH279918.232.049.
15、479.0二二线线抗抗结结核核药药物耐物耐药预测值药预测值1 Prevalence rates2 Results of SLDST proficiency tests done once in 2004 (coordinated by Dr. Nuria Martine Casabona) (participated SRL=9)3 Based on sensitivity/ specificity of 9 rounds proficiency tests of SRLs, 1994-2002Resistance predictive values (%)Resistance prevalence 敏感性敏感性= 99.6%特特异异性性= 97.9%
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