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目的：了解我国结核分支杆菌耐链霉素（Ｓｔｒｅｐｔｏｍｙｃｉｎ，ＳＭ）分离株ｒｐｓＬ基因突变情况，建立快速检测结核分支杆菌耐药基因型的分子药敏试验方法。方法：通过聚合酶链反应（ＰｏｌｙｍｅｒａｓｅＣｈａｉｎＲｅａｃｔｉｏｎ，ＰＣＲ）－单链构象多态性（Ｓｉｎｇｌｅ－ｓｔｒａｎｄｅｄＣｏｎｆｏｒｍａｔｉｏｎＰｏｌｙｍｏｒｐｈｉｓｍ，ＳＳＣＰ）、ＰＣＲ－限制性片段长度多态性（ＲｅｓｔｒｉｃｔｉｏｎＦｒａｇｍｅｎｔＬｅｎｇｔｈＰｏｌｙｍｏｒｐｈｉｓｍ，ＲＦＬＰ）和ＰＣＲ－直接测序法（ＤｉｒｅｃｔＳｅｑｕｅｎｃｉｎｇ，ＤＳ）分析３８株结核分支杆菌耐ＳＭ分离株的ｒｐｓＬ基因。结果３８株耐ＳＭ分离株中，２５株ＳＳＣＰ异常、不被ＭｂｏⅡ消化、ＤＳ分析４３位密码子ＡＡＧ→ＡＧＧ突变；１株ＳＳＣＰ异常、可被ＭｂｏⅡ消化、ＤＳ分析３３位密码子ＧＴＡ→ＡＴＡ突变；１２株ＳＳＣＰ正常、可被ＭｂｏⅡ消化、ＤＳ分析未见异常；未发现８８位密码子突变。结论：大多数结核分支杆菌耐ＳＭ是由于其ｒｐｓＬ基因４３位密码子突变所致，采用ＰＣＲ－ＳＳＣＰ、ＰＣＲ－ＲＦＬＰ和ＰＣＲ－ＤＳ方法可快速测定部分结核分支杆菌ＳＭ耐药基因型。
OBJECTIVE: To investigate the mutation of rpsL gene in Streptomycin (SM) isolates of Mycobacterium tuberculosis in China and to establish a molecular susceptibility testing method for rapidly detecting the genotype of Mycobacterium tuberculosis. Methods: Polymerase chain reaction (PCR) - single strand polymorphism polymorphism (SSCP), restriction fragment length polymorphism (RFLP) and PCR - direct sequencing DS) was used to analyze the rpsL gene of 38 Mycobacterium tuberculosis isolates resistant to SM. Results Thirty-five strains of SM-resistant isolates were abnormal and were not digested by MboⅡ. DS analysis of the 43-codon AAG → AGG mutation; one SSCP-type abnormality was digested by MboⅡ. The 33-codon GTA → ATA mutation ; 12 SSCP normal, can be digested by Mbo Ⅱ, DS analysis showed no abnormalities; no 88 codon mutation was found. CONCLUSIONS: Most Mycobacterium tuberculosis resistance to SM is due to the mutation at codon 43 of rpsL gene. The resistance of some M. tuberculosis strains to SM can be rapidly determined by PCR-SSCP, PCR-RFLP and PCR-DS.