Project description:Streptococcus agalactiae (Lancefield’s group B Streptococcus, GBS) is a major bacterial species of genus Streptococcus and has medical and veterinary importance by affecting mainly humans (Maione et al., 2005; Johri et al., 2006), cattle (Keefe, 1997) and fish (Mian et al., 2009). The GBS is the most important pathogen for the Nile tilapia, a global commodity of the aquaculture sector, causing outbreaks of septicemia and meningoencephalitis (Hernández et al., 2009; Mian et al., 2009). This study aimed to evaluate the global abundancy of proteins among the main genotypes of GBS isolated from fish identified in Brazil using a label free shotgun liquid chromatography-ultra definition mass spectrometry (LC-UDMSE) approach and to compare the differential expression of proteins identified between isolates from fish and human.
Project description:Streptococcus uberis and Streptococcus parauberis reference strains and isolates obtained from routine diagnostics were investigated by PCR with oligonucleotide primers designed according to species-specific parts of the 16S rRNA gene, the 23S rRNA gene, and the 16S-23S rRNA intergenic spacer region of both species. All three primer pairs allowed an identification of 67 isolates as S. uberis and 4 isolates as S. parauberis.
Project description:Streptococcus parauberis is an important bacterial fish pathogen that causes streptococcosis in a variety of fish species including the olive flounder. Despite its importance in the aquaculture industry, little is known about the survival strategy of S. parauberis in the host. Therefore, the objective of this study was to produce genome-wide transcriptome data and identify key factors for the survival of S. parauberis SPOF3K in its host. To this end, S. parauberis SPOF3K was incubated in olive flounder serum and nutrient-enriched media as a control. Although S. parauberis SPOF3K proliferated in both culture conditions, the transcriptomic patterns of the two groups were very different. Interestingly, the expression levels of genes responsible for the replication of an S. parauberis plasmid in the presence of olive flounder serum were higher than those in the absence of olive flounder serum, indicating that this plasmid may play an important role in the survival and proliferation of S. parauberis in the host. Several ATP-binding cassette transporters known to transport organic substrates (e.g., biotin and osmoprotectants) that are vital for bacterial survival in the host were significantly up-regulated in S. parauberis cultured in serum. In addition, groEL, dnaK operon, and members of the clp protease family, which are known to play important roles in response to various stressors, were up-regulated in S. parauberis incubated in serum, thus limiting damage and facilitating cellular recovery. Moreover, important virulence factors including the hyaluronic acid capsule (has operon), sortase A (srtA), C5a peptidase (scp), and peptidoglycan O-acetyltransferase (oatA) were significantly upregulated in S. paraubers in serum. These results indicate that S. paraubers can resist and evade the humoral immune responses of fish. The transcriptomic data obtained in this study provide a better understanding of the mode of action of S. parauberis in fish.
Project description:Streptococcus parauberis is a coccoid, nonmotile, alpha-hemolytic, Gram-positive bacterium of the Streptococcaceae family. Streptococcus parauberis strain KCTC11980 was isolated from the kidney of a diseased olive flounder collected from an aquaculture farm on Jeju Island in 2010. The 2.12-Mb genome sequence consists of 44 large contigs in 16 scaffolds and contains 2,214 predicted protein-coding genes, with a G+C content of 35.4%.
Project description:Although Streptococcus parauberis is known as a bacterial pathogen associated with bovine udder mastitis, it has recently become one of the major causative agents of olive flounder (Paralichthys olivaceus) streptococcosis in northeast Asia, causing massive mortality resulting in severe economic losses. S. parauberis contains two serotypes, and it is likely that capsular polysaccharide antigens serve to differentiate the serotypes. In the present study, the complete genome sequence of S. parauberis (serotype I) was determined using the GS-FLX system to investigate its phylogeny, virulence factors, and antigenic proteins. S. parauberis possesses a single chromosome of 2,143,887 bp containing 1,868 predicted coding sequences (CDSs), with an average GC content of 35.6%. Whole-genome dot plot analysis and phylogenetic analysis of a 60-kDa chaperonin-encoding gene and the glyceraldehyde-3-phosphate dehydrogenase (GAPDH)-encoding gene showed that the strain was evolutionarily closely related to Streptococcus uberis. S. parauberis antigenic proteins were analyzed using an immunoproteomic technique. Twenty-one antigenic protein spots were identified in S. parauberis, by reaction with an antiserum obtained from S. parauberis-challenged olive flounder. This work provides the foundation needed to understand more clearly the relationship between pathogen and host and develops new approaches toward prophylactic and therapeutic strategies to deal with streptococcosis in fish. The work also provides a better understanding of the physiology and evolution of a significant representative of the Streptococcaceae.
Project description:This paper describes the predicted structure for the cps loci involved in capsule biosynthesis for Streptococcus parauberis serotypes III, IV, and V. Based on the specific serotype regions I, II, and III, a multiplex PCR protocol (mPCR) was designed to differentiate the main serotypes causing fish diseases. A real-time PCR method (qPCR) is also described to identify S. parauberis of serotype III in bacterial cultures and fish tissues. In silico and in vitro analyses revealed that both methods have a 100% specificity. The mPCR assay was optimized for the detection of S. parauberis strains of subtypes Ia (amplicon size 213 bp), subtypes Ib and Ic (both amplicon size 303 bp), serotype II (amplicon size 403 bp), and serotype III (amplicon size 130 bp) from bacterial cultures. The qPCR assay was optimized for the identification and quantification of S. parauberis serotype III strains in bacterial cultures and fish tissues. This assay achieved a sensitivity of 2.67?×?102 gene copies (equivalent to 3.8?×?10-9 ng/?l) using pure bacterial cultures of S. parauberis serotype III and 1.76?×?102 gene copies in fish tissues experimentally and naturally infected with S. parauberis of the serotype III. The specificity and sensitivity of the protocols described in this study suggest that these methods could be used for diagnostic and/or epidemiological purposes in clinical diagnostic laboratories. KEY POINTS: • Structure of loci cps for S. parauberis of serotypes III, IV and V was described. • mPCR to differentiate S. parauberis serotypes causing disease in fish was optimized. • qPCR assay to quantify strains of S. parauberis serotype III in fish tissues.
Project description:Streptococcus agalactiae (Lancefield’s group B Streptococcus, GBS) is a major bacterial species of genus Streptococcus and has medical and veterinary importance by affecting mainly humans (Maione et al., 2005; Johri et al., 2006), cattle (Keefe, 1997) and fish (Mian et al., 2009). The GBS is the most important pathogen for the Nile tilapia, a global commodity of the aquaculture sector, causing outbreaks of septicemia and meningoencephalitis (Hernández et al., 2009; Mian et al., 2009).
Project description:A multiplex PCR protocol was established to simultaneously detect major bacterial pathogens in olive flounder (Paralichthys olivaceus) including Edwardsiella (E.) tarda, Streptococcus (S.) parauberis, and S. iniae. The PCR assay was able to detect 0.01 ng of E. tarda, 0.1 ng of S. parauberis, and 1 ng of S. iniae genomic DNA. Furthermore, this technique was found to have high specificity when tested with related bacterial species. This method represents a cheaper, faster, and reliable alternative for identifying major bacterial pathogens in olive flounder, the most important farmed fish in Korea.