台灣魚隻分離之無乳鏈球菌溶血型表現與其基因分析

林孟謙

doi:10.6342/NTU.2015.02336

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台灣魚隻分離之無乳鏈球菌溶血型表現與其基因分析

Hemolysis Phenotype and Genetic Analysis of Streptococcus agalactiae Isolated from Diseased Fish in Taiwan

林孟謙 , Masters　　Advisor：陳媺玫　　

繁體中文 DOI： 10.6342/NTU.2015.02336

無乳鏈球菌；溶血； Streptococcus agalactiae ； hemolysis

Abstract │ Reference (167) │ Altmetrics

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Streptococcus agalactiae are Gram positive cocci, also known as Group B Streptococcus (GBS). S. agalactiae can infect numerous fish species, including tilapia, rainbow trout, mullet, and hybrid striped seabass, and cause economic losses during cultivation. From 2009 to 2014, it was found that all S. agalactiae strains isolated from cultured fishes, such as tilapia and seabass, had different hemolytic phenotypes including α, β and γ. GBS hemolysin is responsible for hemolysis, and it is encoded by cyl operon. cyl operon is mainly regulated by the two-component regulatory systems (TCS), covS/R (control of virulence, sensor and regulator). Besides hemolysis, covS/R can also regulate encapsulation. The objective of the present study was to compare phenotype differences among three hemolytic variant strains and investigate genetic composition of hemolysin genes. α and β hemolytic strains showed no difference on encapsulation and growth rate, and these two strains grew into stationary phase after 18 hours culture. γ hemolytic strain showed higher encapsulation by Indian ink staining and slower growth rate with the stationary phase of 48 hours. Genetic analysis of cyl operon among GBS strains by PCR and sequencing found a 1,252 bp insertion sequence (IS), located in cylF region of α hemolytic strains. The IS had two open reading frames, both encoding transposase. The IS interrupted cylF gene by insertion in 23 bp downstream of the ATG start codon of cylF region, causing incomplete mRNA transcription of cylF. β hemolytic strain showed no mutation in cyl operon. γ hemolytic strain lost whole cyl operon with replacement of 14 kb genomic island containing genes of DNA recombinase and septum formation protein, Maf, etc. No mutation of covS/R regulatory gene was found among all three hemolytic variant strains. The results of the present study indicated that the difference of hemolysin gene between α and β hemolytic strain was due to the IS inserted in cylF region, while in the case of γ hemolytic strain, whole cyl operon was deleted and replaced. Differences of encapsulation were found among three hemolytic strains, with no genetic difference of covS/R regulatory gene.

Reference ( 167 ) 〈TOP〉

Abuseliana A, Daud H, Abdul -Aziz S, Khairani -Bejo S, Alsaid M (2010) Streptococcus agalactiae the etiological agent of mass mortality in farmed red tilapia (Oreochromis sp.) J Anim Vet Adv 9, 2640-2646.
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Bartoli A & Marconi P (2011) Group B streptococcus (GBS) disrupts by calpain activation the actin and microtubule cytoskeleton of macrophages. Cell Microbiol 13: 859–884.
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