Comparative genomics analysis of Streptococcus iniae isolated from Trachinotus ovatus: novel insight into antimicrobial resistance and virulence differentiation.

BACKGROUND: Streptococcus iniae is an important fish pathogen that cause significant economic losses to the global aquaculture industry every year. Although there have some reports on the genotype of S.iniae and its relationship with virulence, no genome-scale comparative analysis has been performed so far. In our previous work, we characterized 17 isolates of S.iniae from Trachinotus ovatus and divided them into two genotypes using RAPD and rep-PCR methods. Among them, BH15-2 was classified as designated genotype A (in RAPD) and genotype 1 (in rep-PCR), while BH16-24 was classified as genotype B and genotype 2. Herein, we compared the differences in growth, drug resistance, virulence, and genome between BH15-2 and BH16-24. RESULTS: The results showed that the growth ability of BH16-24 was significantly faster than that of BH15-2 at the exponential stage. Antimicrobial tests revealed that BH15-2 was susceptible to most of the tested antibiotics except neomycin and gentamycin. In contrast, BH16-24 was resistant to 7 antibiotics including penicillin, sulfasomizole, compound sulfamethoxazole tablets, polymyxin B, spectinomycin, rifampin and ceftazidime. Intraperitoneal challenge of T.ovatus, showed that the LD50 value of BH15-2 was 4.0 × 102 CFU/g, while that of BH16-24 was 1.2 × 105 CFU/g. The genome of S.iniae BH15-2 was 2,175,659 bp with a GC content of 36.80%. Meanwhile, the genome of BH16-24 was 2,153,918 bp with a GC content of 36.83%. Comparative genome analysis indicated that compared with BH15-2, BH16-24 genome had a large-scale genomic inversion fragment, at the location from 502,513 bp to 1,788,813 bp, resulting in many of virulence and resistance genes differentially expression. In addition, there was a 46 kb length, intact phage sequence in BH15-2 genome, which was absent in BH16-24. CONCLUSION: Comparative genomic studies of BH15-2 and BH16-24 showed that the main difference is a 1.28 Mbp inversion fragment. The inversion fragment may lead to abnormal expression of drug resistant and virulence genes, which is believed to be the main reason for the multiple resistance and weakened virulence of BH16-24. Our study revealed the potential mechanisms in underlying the differences of multidrug resistance and virulence among different genotypes of S.iniae.

Efficacy and transcriptome analysis of golden pompano (Trachinotus ovatus) immunized with a formalin-inactived vaccine against Streptococcus iniae.

Streptococcus iniae is a worldwide fish pathogen that cause tremendous economic losses to the global aquaculture industry. Vaccination is regarded as the most effective and safe way to control fish diseases. In our study, we developed a formalin-inactivated vaccine against S. iniae and evaluated its effect in golden pompano (Trachinotus ovatus). In addition, in order to clarify the molecular mechanisms underlying the vaccine protection, we compared the spleen transcriptomes of vaccinated and unvaccinated golden pompano at 1, 2 and 7 d post vaccination using the RNA-seq technology. The relative percentage survival (RPS) reached 71.1% at 28 days post-vaccination which suggested that the vaccine provided highly protection against S. iniae. KEGG pathway analysis revealed that phagosome, cytokine-cytokine receptor interaction, MAPK signaling pathway, and CAMs were activated by the vaccine. The most of strongly up-regulated genes in golden pompano spleen are involving in innate immunity. For adaptive immunity, the vaccine evoked a CD8+ CTL-mediated response by MHC Ⅰ pathway to achieve immune protection.

Molecular characterization and expression of interleukin-10 and interleukin-22 in golden pompano (Trachinotus ovatus) in response to Streptococcus agalactiae stimulus.

In the present study, members of the interleukin (IL)-10 family of cytokines, including IL-10 (TOIL-10) and IL-22 (TOIL-22) of golden pompano (Trachinotus ovatus), were cloned for the first time, and their expression patterns and 3D structures analyzed. The full-length cDNA sequences of TOIL-10 and TOIL-22 contained open reading frames of 564 and 567 bp, respectively. TOIL-10 and TOIL-22 shared higher homology (78%-89%) with the corresponding genes from various fish relative to other species (25%-34%) and contained the IL-10 family signature and four cysteine residues that are well conserved in other vertebrate IL-10 members. Phylogenetic tree analysis of our sequences alongside other IL-10 family proteins revealed that TOIL-10 and TOIL-22 cluster together with other teleost IL-10 and IL-22 molecules. Expression of TOIL-10 and TOIL-22 genes was ubiquitous in all tissues examined. The TOIL-10 gene was also highly expressed in skin, heart, gill, spleen, kidney, brain and liver, and lower levels were detected in intestine and muscle. High expression of the TOIL-22 gene was observed in gill, intestine, kidney, spleen, with the lowest levels in liver. TOIL-10 and TOIL-22 were rapidly activated after SAΔphoB immunization and significantly increased to peak levels at 12 h and 4 d in golden pompano kidney and spleen respectively following challenge. Expression in the brain reached peak levels at 4 d and 3 d respectively after post-immunization. Our results collectively indicate that TOIL-10 and TOIL-22 participate in the host immune response to bacterial infection. Moreover, TOIL-22 plays a potentially important role in mucosal immunity.