Mutation in cyl operon alters hemolytic phenotypes of Streptococcus agalactiae.

Streptococcus agalactiae infects numerous fish species, causing considerable economic losses during fish cultivation. This study compared the phenotypic differences among S. agalactiae hemolytic variant isolates and investigated the genetic composition of their hemolysin genes. Hemolysin is encoded by the cyl operon and mainly regulated by covS/R, which also regulates encapsulation. In total, 45 S. agalactiae clinical isolates were collected from cultured fishes in Taiwan. Three different hemolytic phenotypes-α, β, and γ-were identified. Of the 45 isolates, 39 were β hemolytic, 3 were α hemolytic, and 3 were γ hemolytic. The γ-hemolytic isolates demonstrated significantly thicker encapsulation and slower growth rates than did the α- and β-hemolytic isolates. However, no isolate had mutations in the regulatory gene covS/R. A 1252-bp insertion sequence (IS) in the cyl operon of α-hemolytic isolates, located at cylF region, was found. This IS interrupted cylF through insertion at 23 bp downstream of starting codon, causing incomplete mRNA transcription. The β-hemolytic isolates showed no mutation in the cyl operon. By contrast, the γ-hemolytic isolates had lost the entire cyl operon; it had been replaced by a 14-kb genomic island containing genes for DNA recombinase and septum formation proteins. In summary, the differences in hemolysin genes between α- and β-hemolytic isolates were due to the IS in the cylF region, whereas in the γ-hemolytic isolates, the entire cyl operon was deleted and replaced. These findings explain different hemolysin expressions of the clinical S. agalactiae isolates taken from fish ponds in Taiwan. IMPORTANCE: Streptococcus agalactiae infects both warm- and cold-blooded animals and causes major aquatic cultivation loss. Pathogenic isolates from the outbreak of fish ponds were examined their cyl operon gene. α-Hemolytic isolate with mutant cyl operon was observed for the first time in aquaculture animals and was compared to intact or entire cyl operon deletion of β- and γ-hemolytic isolates. Hemolysis expression levels of Streptococcus agalactiae are explained.

Chou CC ，Lin MC ，Su FJ ，Chen MM ... -  《-》

Phenotypic and molecular characterization of hyperpigmented group B Streptococci.

Group B Streptococcus (GBS) causes invasive infections in neonates, older adults and patients with comorbidities. β-hemolysin/cytolysin is an important GBS virulence factor. It is encoded by the cyl operon and confers GBS hemolytic activity. Isolates displaying hyperpigmentation are typically hyperhemolytic. Comparison of clonally identical isolates displaying different levels of pigmentation has shown transcriptional dysregulation due to mutations in components of the control of the virulence S/R (CovS/R) regulatory system. In addition, hyperpigmented isolates show decreased CAMP factor and decreased capsule thickness. In analogy to findings in group A Streptococcus, a pivotal role of CovS/R has been proposed in the host-pathogen interaction of invasive GBS infection. However, corresponding investigations on multiple clinical GBS isolates have not been performed. We prospectively collected hyperpigmented isolates found in a diagnostic laboratory and performed phenotypic, molecular and transcriptional analyses. In the period from 2008 to 2012, we found 10 isolates obtained from 10 patients. The isolates reflected both invasive pathogens and colonizers. In three cases, clonally identical but phenotypically different variants were also found. Hence, the analyses included 13 isolates. No capsular serotype was found to be significantly more frequent. Bacterial pigments were analyzed via spectrophotometry and for their hemolytic activity. Data obtained for typical absorbance spectra peaks correlated significantly with hemolytic activity. Molecular analysis of the cyl operon showed that it was conserved in all isolates. The covR sequence displayed mutations in five isolates; in one isolate, the CovR binding site to cylX was abrogated. Our results on clinical isolates support previous findings on CovR-deficient isogenic mutants, but suggest that - at least in some clinical isolates - for β-hemolysin/cytolysin and CAMP factor production, other molecular pathways may be involved.

Lupo A ，Ruppen C ，Hemphill A ，Spellerberg B ，Sendi P ... -  《-》

The cyl genes of Streptococcus agalactiae are involved in the production of pigment.

Spellerberg B ，Martin S ，Brandt C ，Lütticken R ... -  《FEMS MICROBIOLOGY LETTERS》

Genetic Basis Underlying the Hyperhemolytic Phenotype of Streptococcus agalactiae Strain CNCTC10/84.

Streptococcus agalactiae (group B streptococcus [GBS]) is a major cause of infections in newborns, pregnant women, and immunocompromised patients. GBS strain CNCTC10/84 is a clinical isolate that has high virulence in animal models of infection and has been used extensively to study GBS pathogenesis. Two unusual features of this strain are hyperhemolytic activity and hypo-CAMP factor activity. These two phenotypes are typical of GBS strains that are functionally deficient in the CovR-CovS two-component regulatory system. A previous whole-genome sequencing study found that strain CNCTC10/84 has intact covR and covS regulatory genes. We investigated CovR-CovS regulation in CNCTC10/84 and discovered that a single-nucleotide insertion in a homopolymeric tract in the covR promoter region underlies the strong hemolytic activity and weak CAMP activity of this strain. Using isogenic mutant strains, we demonstrate that this single-nucleotide insertion confers significantly decreased expression of covR and covS and altered expression of CovR-CovS-regulated genes, including that of genes encoding β-hemolysin and CAMP factor. This single-nucleotide insertion also confers significantly increased GBS survival in human whole blood ex vivoIMPORTANCE Group B streptococcus (GBS) is the leading cause of neonatal sepsis, pneumonia, and meningitis. GBS strain CNCTC10/84 is a highly virulent blood isolate that has been used extensively to study GBS pathogenesis for over 20 years. Strain CNCTC10/84 has an unusually strong hemolytic activity, but the genetic basis is unknown. In this study, we discovered that a single-nucleotide insertion in an intergenic homopolymeric tract is responsible for the elevated hemolytic activity of CNCTC10/84.

Zhu L ，Beres SB ，Yerramilli P ，Pruitt L ，Cantu CC ，Olsen RJ ，Musser JM ... -  《-》

The transcriptional regulator RovS controls the attachment of Streptococcus agalactiae to human epithelial cells and the expression of virulence genes.

Samen UM ，Eikmanns BJ ，Reinscheid DJ 《-》