Fernebro Jenny, Andersson Ingrid, Sublett Jack, Morfeldt Eva, Novak Rodger, Tuomanen Elaine, Normark Staffan, Normark Birgitta Henriques
Department of Molecular Epidemiology and Biotechnology, Smittskyddsinstitutet, Solna, Stockholm, Sweden.
J Infect Dis. 2004 Jan 15;189(2):328-38. doi: 10.1086/380564. Epub 2003 Dec 30.
Penicillin and vancomycin induce a lytic response in Streptococcus pneumoniae that requires the N-acetylmuramyl-l-alanine amidase LytA. We show that clinical isolates of pneumococci of capsular serotypes 1, 4, 6B, and 23F were generally less lytic to penicillin than pneumococci of serotypes 14 and 3. In addition, most 9V isolates were less lytic to vancomycin, compared with isolates of other serotypes. Parent-mutant pairs expressing and not expressing capsular serotypes 2, 4, and 9V were compared for antibiotic-induced lysis. The nonencapsulated variants were considerably more lytic after beta-lactam and/or vancomycin treatment, and antibiotic tolerance was seen only in the context of capsule expression. Conversion from a nonlytic to a lytic phenotype, after loss of capsule expression, required an intact lytA autolysin gene. Exogenous addition of purified LytA gave a lower lytic response in capsulated strains, compared with that in nonencapsulated mutants. Spontaneous autolysis in stationary phase also was negatively affected by capsule expression in an autolysin-dependent manner. Long-term starvation in the stationary phase of the vancomycin- and penicillin-tolerant isolate I95 yielded nonencapsulated mutants that had lost antibiotic tolerance and were lytic to penicillin and vancomycin. The 9V capsular locus of I95 and one of these stationary phase-selected mutants were completely sequenced. The only difference found was a 1-bp frameshift deletion in the cps9vE gene of the lytic mutant, encoding a uridine diphosphate-glucosyl-1-phosphate transferase. Two additional independently isolated lytic mutants of I95 from the stationary phase also contained mutations in the same region of cps9vE, which identified it as a mutational hot spot. This report demonstrates that capsular polysaccharides negatively influence the lytic process and contribute to antibiotic tolerance in clinical isolates of pneumococci.
青霉素和万古霉素可诱导肺炎链球菌产生溶菌反应，这一过程需要N - 乙酰胞壁酰 - L - 丙氨酸酰胺酶LytA的参与。我们发现，与14型和3型肺炎链球菌相比，1、4、6B和23F荚膜血清型肺炎链球菌的临床分离株对青霉素的溶菌作用通常较弱。此外，与其他血清型的分离株相比，大多数9V分离株对万古霉素的溶菌作用也较弱。对表达和不表达2、4和9V荚膜血清型的亲本 - 突变体对进行抗生素诱导的溶菌作用比较。非荚膜变体在β - 内酰胺和/或万古霉素处理后溶菌作用明显增强，且仅在荚膜表达的情况下才观察到抗生素耐受性。在荚膜表达缺失后，从非溶菌表型转变为溶菌表型需要完整的lytA自溶素基因。与非荚膜突变体相比，在荚膜菌株中外源添加纯化的LytA产生的溶菌反应较低。稳定期的自发自溶也以自溶素依赖的方式受到荚膜表达的负面影响。在万古霉素和青霉素耐受的分离株I95的稳定期进行长期饥饿培养，产生了非荚膜突变体，这些突变体失去了抗生素耐受性，并且对青霉素和万古霉素具有溶菌作用。对I95的9V荚膜基因座和其中一个稳定期选择的突变体进行了全序列测定。发现的唯一差异是溶菌突变体的cps9vE基因中一个1个碱基的移码缺失，该基因编码尿苷二磷酸 - 葡萄糖基 - 1 - 磷酸转移酶。另外两个从稳定期独立分离的I95溶菌突变体在cps9vE的同一区域也含有突变，这表明该区域是一个突变热点。本报告表明，荚膜多糖对溶菌过程有负面影响，并导致肺炎链球菌临床分离株产生抗生素耐受性。
