Le Hao

LuxS/AI-2 in Streptococcus agalactiae reveals a key role in acid tolerance and virulence

LuxS-mediated autoinducer-2 (AI-2) directly or indirectly regulates important physiologic function in a variety of bacteria. We found a luxS homologue in the genome of Streptococcus agalactiae, an important pathogen of tilapia. To investigate the relationship between luxS/AI-2 and pathogenicity for tilapia, its bioluminescent activity, acid resistance, cell adherence, virulence, and regulation of virulence gene were evaluated. Compared with the wild-type strain, the bioluminescent activity lost in the luxS mutant, its resistance to acid (pH2.8) was significantly decreased 33.8 times, and furthermore, its adherence to the NGF-2 cell line was dramatically reduced 3 times in the mutant strain. The virulence of the mutant strain was decreased in the tilapia infection model, exogenous AI-2 molecule (7.4nM) and luxS gene complementation with plasmid could complement the deficiencies of function in the luxS mutant strain. These results showed that inactivation of luxS gene caused a significant decrease of bioluminance, acid resistance, cell adhesion, virulence to tilapia and transcription levels of many virulence genes in S. agalactiae. Expression of the known stress resistance factors DnaK and GroEL, relative regulator factors CovR/CovS and virulence factor cpsE verified above results. These findings suggest that luxS may be involved in the interruption of bacterial virulence and resistance to environmental factors.

Multiple Evolutionary Selections Involved in Synonymous Codon Usages in the Streptococcus agalactiae Genome

Streptococcus agalactiae is an important human and animal pathogen. To better understand the genetic features and evolution of S. agalactiae, multiple factors influencing synonymous codon usage patterns in S. agalactiae were analyzed in this study. A- and U-ending rich codons were used in S. agalactiae function genes through the overall codon usage analysis, indicating that Adenine (A)/Thymine (T) compositional constraints might contribute an important role to the synonymous codon usage pattern. The GC3% against the effective number of codon (ENC) value suggested that translational selection was the important factor for codon bias in the microorganism. Principal component analysis (PCA) showed that (i) mutational pressure was the most important factor in shaping codon usage of all open reading frames (ORFs) in the S. agalactiae genome; (ii) strand specific mutational bias was not capable of influencing the codon usage bias in the leading and lagging strands; and (iii) gene length was not the important factor in synonymous codon usage pattern in this organism. Additionally, the high correlation between tRNA adaptation index (tAI) value and codon adaptation index (CAI), frequency of optimal codons (Fop) value, reinforced the role of natural selection for efficient translation in S. agalactiae. Comparison of synonymous codon usage pattern between S. agalactiae and susceptible hosts (human and tilapia) showed that synonymous codon usage of S. agalactiae was independent of the synonymous codon usage of susceptible hosts. The study of codon usage in S. agalactiae may provide evidence about the molecular evolution of the bacterium and a greater understanding of evolutionary relationships between S. agalactiae and its hosts.

Protective efficacy of cationic-PLGA microspheres loaded with DNA vaccine encoding the sip gene of Streptococcus agalactiae in tilapia.

Abstract Streptococcus agalactiae (S. agalactiae) is an important fish pathogen, which has received more attention in the past decade due to the increasing economic losses in the tilapia industry worldwide. As existing effective vaccines of S. agalactiae in fish have obvious disadvantage, to select immunoprotective antigens and package materials would undoubtedly contribute to the development of novel oral vaccines. In the present study, surface immunogenic protein (sip) was selected from the S. agalactiae serovar I a genomes as immunogenic protein in DNA vaccine form with cationic chitosan and biodegradable and biocompatible PLGA. The pcSip plasmid in cationic‐PLGA was successfully expressed in tissues of immunized tilapia and the immunogenicity was assessed in tilapia challenge model. A significant increase was observed in the cytokine levels of IL‐1&bgr;, TNF‐&agr;, CC1, CC2 in spleen and kidney tissues. Furthermore, immunized tilapia conferred different levels of protection against challenge with a lethal dose of highly virulent serovar I a S. agalactiae. Our results indicated that the pcSip plasmid in cationic‐PLGA induced high level of antibodies and protection against S. agalactiae infection, could be effective oral DNA vaccine candidates. HighlightsSurface immunogenic protein (sip) was selected as immunogenic protein for DNA vaccine.DNA vaccine loaded in cationic chitosan and biodegradable and biocompatible PLGA.The immunogenicity of DNA vaccine was assessed in tilapia challenge model.The DNA vaccine induced high level of antibodies and protection against S. agalactiae infection.