在固態及液態培養基中都會產生第一型線毛的鼠傷寒沙門氏菌ubiB突變株特性分析

Characterization of a Salmonella enterica serotype Typhimurium ubiB mutant that constitutively produces type 1 fimbriae both in static broth and on solid agar culture conditions

張立寰(Li-Huan Chang)

指導教授 ： 葉光勝

臺北醫學大學/醫學院/醫學科學研究所/碩士(2008年)

https://doi.org/10.6831/TMU.2008.00024

摘要

鼠傷寒沙門氏菌(S. Typhimurium)第一型線毛的fim 基因組會受到環境的影響而進行表現與否。本實驗設計兩種培養環境，靜置培養液與固態培養基，當野生株鼠傷寒沙門氏菌在此兩種環境下生長時，在靜置培養液的菌株會表現第一型線毛；反之，在固態培養基上則沒有表現。目前，除了調控第一型線毛已知的基因fimZYWU 和構成線毛的基因fimAICDFH 外，在本實驗室先前的研究結果發現，突變株S. Typhimurium ubiB 也會影響到第一型線毛的表現，在兩種培養環境下皆會產生第一型線毛。UbiB 是一NADP(H) reductase，參與coenzyme B12 的合成。利用回復試驗所建立的S. Typhimurium ubiB(pTA-ubiB)，可以確定ubiB 會影響第一型線毛的表現。接著，使用RT-PCR 來個別增幅ubiB 影響調控基因fimZYW，結果發現當S. Typhimurium ubiB 生長在agar 上，正向調控第一型線毛的fimZ 有被表現出來，而fimY 和fimW 則沒有表現。在電子顯微鏡觀察下，S. Typhimurium ubiB 在固態及液態培養基均表現第一型線毛。利用震盪培養來觀察S. Typhimurium ubiB 生長曲線，發現ubiB 突變株之生長速率較野生株與回復株緩慢許多。在之前的研究發現，沙門氏菌存活在嚴峻的環境下，傾向會表現出線毛以適應環境。我們的研究顯示ubiB會影響第一型線毛的表現，更深入的研究值得進一步探討。

關鍵字

鼠傷寒沙門氏菌 ； 第一型線毛 ； ubiB

並列摘要

Salmonella is an important food-borne pathogen of public health concern. Sal-monella enterica contains more than 2,500 serotypes among which Typhimurium is an important etiology of gastroenteritis. Adhesion of bacteria to the host epithelial cells is a prerequisite step in establishing infection, while fimbriae are the primary structures implicated in such an adherent event. The type 1 fimbriae are the most commonly found fimbrial appendages on the outer membrane of Salmonella enterica serotype Typhimurium. The strongly fimbriate-phase bacteria can be isolated following serial passage every 48 hours in static broth culture, while non-fimbriate bacteria are obtained by growth on solid agar media. The phenotypic expression of type 1 fimbriae in S. Typhimurium is the result of the interaction and cooperation of several genes in the fim gene cluster. A transposon insertion mutagenesis has revealed that several genes outside the fim gene cluster may also involve in the expression of type 1 fimbriae both in static broth and on solid agar culture conditions. One S. Ty-phimurium mutant ubiB constitutively produced type 1 fimbriae in both culture condi-tions. The ubiB gene could encode a NAD(P)H-flavin reductase. Southern hybri-dization indicated that the transposition event occurred once in this mutant strain. Transforming the plasmid pTA-ubiB that contained the coding sequence of ubiB con-ferred the ubiB mutant back to the type 1 fimbrial phenotype as seen in the parental strain as indicated by yeast agglutination and electron microscopy. Reverse tran-scription polymerase chain reaction (RT-PCR) indicated that the fimA expression of the ubiB mutant strain obtained from the static broth was about 1.2% of that obtained from the solid agar culture. When ubiB strain harbored the plasmid pTA- ubiB, the fimA expression from broth was about 34 fold of that from the solid agar. As a con-trol, 16S rRNA was constantly expressed in all the strains tested. Growth curve analysis suggested that ubiB mutant had a slower growth rate in the log phase than the parental strain and the ubiB (pTA- ubiB) strain. How ubiB gene product affects type 1 fimbrial expression in S. Typhimurium is of interest and warrants further investiga-tion.

並列關鍵字

Salmonella enterica serotype Typhimurium ； type 1 fimbriae ； ubiB

參考文獻

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