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Streptococcus parauberis is an alpha-hemolytic gram positive coccoid bacterium belonging to the Streptococcaceae family. This bacterium cause streptococcosis is a major disease in cultured fish due to the intensification of aquaculture and causes significant economic losses in fish farm industry. In our study, we isolated a total of 37 lactic acid bacteria from wild fish (Sander lucioperca, Carassius gibelio, Cyprinus carpio) of Lake Eğirdir and cultured fish (Oncorhynchus mykiss, Walbaum) of farm in Turkey. For the isolation of bacteria phenotypic and biochemical characteristics of the 37 isolates obtained from the colonies grown on MRS, TSA and M17 agar characterized by determining colony morphology, cell morphology, motility, gram staining and the production of cytochrome oxidase and catalase. Further biochemical characteristics were determined using conventional tests according to Bergey’s Manual of Systematic Bacteriology. From the isolated lactic acid bacteria 25 of were identified as S. parauberis, 2 of as Vagococcus sp., 2 of as Lactococcus garvieae and 5 of as Lactococcus lactis by culture-based, biochemical test and 16Sr RNA gene sequencing techniques but 3 of them couldn’t (F65, F49 and F50) identified. In this study, the main objective of us is to identify S. parauberis with conventional and 16Sr RNA gene sequencing techniques from wild fish in Lake Eğirdir and in cultured fish in fish farm of Egirdir-Turkey.

This article provides efficacy and safety data of ceftiofur antibiotics against streptococcal infection in starry flounder. Ceftiofur, which is a veterinary antibiotics, is effective against fishery bacteria. Ceftiofur can be prescribed and sold by veterinarians. However, it is illegal in South Korea for fishery disease managers to prescribe and sell ceftiofur. Therefore, in order to utilize available antibiotics and prevent illegal use of veterinary antibiotics, it is necessary to perform research to determine the recommended effective dose and administration methods of antibiotics for fisheries. In this article, the appropriate concentration and injection method of antibiotics to treat starry flounder infected with S. parauberis PH0710 were provided. In addition, histopathological examination results were provided to confirm the effect of antibiotics on the host tissue. Accordingly, these data could be used as basic data for the application of ceftiofur antibiotics in disease management for fisheries. Keywords: Antibiotics, Ceftiofur, Starry flounder, Streptococcus parauberis PH0710, Efficacy, Safety

Streptococcus parauberis is utilized as an oral vaccine by first inactivating the cells with formalin to produce formalin- killed cells (FKC) and then encapsulating them with polymer beads consisting of a cross-linked alginate-Ca2+ network. The encapsulation efficiency and media-dependent release are controlled by pre-treating the FKC with two types of clay nanoparticles: kaolinite (KA) and layered double hydroxide (LDH). The addition of LDH induced large agglomerates of FKC, and the KA enhanced the dispersion of FKC. The differences in the dispersibility of the FKC upon the use of clay nanoparticles was determined to strongly affect the encapsulation efficiency and release properties. The FKC + LDH mixture exhibited a slightly reduced encapsulation efficiency compared to the FKC alone. However, FKC + KA exhibited a dramatically improved encapsulation efficiency. In terms of the media-dependent release, the alginate beads were found to be fairly stable under gastric conditions and in deionized water with or without clay nanoparticles, preserving most of the encapsulated FKC. The intestine was the final target organ for FKC vaccination, and release at the site varied according to the use of clay nanoparticles. Both clays seemed to enhance the release of FKC, the cumulative amount being 3.6 times and 1.3 times larger for LDH and KA, respectively, than was shown with only FKC encapsulated beads.

This chapter also summarized eight different fish diseases from saline water, viz. 1) Saltwater Streptococcosis (Streptococcus Dysgalactiae, Streptococcus Iniae, Streptococcus parauberis), 2) Nocardiosis, 3) Mycobacterial disease, 4) Pasteurellosis, 5) Vibriosis, 6) Tenacibaculosis, 7) Edwardsiellosis (Edwardsiella tarda), and 8) bacterial hemolytic jaundice. The characteristics of disease agent, and pathogenesis, histopathological interest, diagnostic method, chemotherapy and disease control were introduced., E-book edition, E-book: 978-1-78021-040-7, Print: 978-1-78021-540-2, Authors, keywords, contents, Summary [p. 190-191], 1. Saltwater streptococcosis / Terutoyo Yoshida [p. 191-213], 2. Nocardiosis / Masahiro Sakai [p. 213-217], 3. Mycobacterial disease / Kim D. Thompson and Alexandra Adams [p. 217-221], 4. Pasteurellosis / Tae-Sung Jung and Takashi Aoki [p. 221-229], 5. Vibriosis / Yukinori Takahashi and Jun-ichi Hikima [p. 229-238], 6. Tenacibaculosis / Hisatsugu Wakabayashi [p. 239-244], 7. Edwardsiellosis (edwardsiella tarda) / Tomokazu Takano [p. 245-251], 8. Bacterial hemolytic jaundice / Takaji Iida [p. 251-254], Bibliography, biographical sketches [p. 254-277]

Streptococcus parauberis is a pathogen of streptococcosis in turbot Scophthalmus maximus and the Japanese flounder Paralichthys olivaceus. S. parauberis isolates from diseased Japanese flounder have been classified into several serological phenotypes. In this study we analyzed the DNA sequence of the genetic locus for capsular polysaccharide (CPS) biosynthesis of S. parauberis KRS02083 (subserotype Ia), NUF1003 (subserotype Ib), NUF1071 (subserotype Ic), NUF1032 (serotype II), 2007-1 (nontypeable/PFGE cluster I) and NUF1095 (nontypeable/PFGE cluster III) to elucidate the genetic basis for serological diversity. As a result, three kinds of cps locus were revealed among serotypes and subserotypes, namely the loci for subserotype Ia, subserotypes Ib/Ic and serotype II. The genetic structure of cps loci suggests that the capsules of S. parauberis are synthesized through the Wzy-dependent pathway. Subserotypes Ib and Ic possessed the same genetic structure, although single-base substitution at several regions or insertion of an IS (insertion sequence) element was found in subserotype Ic. The nontypeable strains, which agglutinated with both serotypes I and II antisera, possessed the same genetic structure as subserotype Ib/Ic or serotype II with single-base substitution at several regions., Fish Pathology, 50(4), pp.192-199; 2015

Streptococcus parauberis is the etiologic agent of streptococcosis in Japanese flounder Paralichthys olivaceus. Two serotypes, termed serotypes I and II, are known among the Japanese S. parauberis isolates. In the course of serodiagnosis, we found several strains that did not agglutinate with rabbit anti-serotype I or II sera. In this study, we investigated the serological and genetic relationships among the stocked S. parauberis strains including the non-agglutinating ones using a newly prepared rabbit antiserum against a non-agglutinating strain (NUF1071) as well as previously prepared antiserotype I and II sera, and pulsed-field gel electrophoresis (PFGE). An antiserum cross-absorption test and microtiter agglutination test revealed that the serotype I was divided into three subserotypes, tentatively designated Ia, Ib and Ic. The non-agglutinating strains belonged to the subserotype Ic. Of the 104 serotype I strains, 6, 91 and 7 strains belonged to subserotypes Ia, Ib and Ic, respectively. Formalin-killed cells of subserotype Ia and Ic strains were agglutinated with the anti-subserotype Ia serum (so far being used as an anti-serotype I serum) and Ic serum, respectively. Subserotype Ib strains were agglutinated with both sera. In PFGE analysis, the stocked 188 S. parauberis strains were divided into three clusters corresponding to subserotypes Ib/Ic, Ia and serotype II., 魚病研究, 50(2), pp.75-80; 2015

A 12-year-old boy presented with trauma to left eye with a wooden stick. On examination, there was full thickness corneal laceration with cataractous lens behind the laceration. The laceration was sutured, and intravitreal injections of vancomycin, ceftazidime and clindamycin were administered. Vitreous tap grew Streptococcus parauberis. The isolate was sensitive to amoxicillin, erythromycin and vancomycin, and topical vancomycin was used to treat the infection. We present the first case of human post-traumatic infective endophthalmitis caused by the rare agent S. parauberis.

Streptococcus parauberis is the etiologic agent of streptococcosis in Japanese flounder Paralichthys olivaceus. Two serotypes, termed serotypes I and II, are known among the Japanese S. parauberis isolates. In the course of serodiagnosis, we found several strains that did not agglutinate with rabbit anti-serotype I or II sera. In this study, we investigated the serological and genetic relation- ships among the stocked S. parauberis strains including the non-agglutinating ones using a newly pre- pared rabbit antiserum against a non-agglutinating strain (NUF1071) as well as previously prepared anti- serotype I and II sera, and pulsed-field gel electrophoresis (PFGE). An antiserum cross-absorption test and microtiter agglutination test revealed that the serotype I was divided into three subserotypes, tenta- tively designated Ia, Ib and Ic. The non-agglutinating strains belonged to the subserotype Ic. Of the 104 serotype I strains, 6, 91 and 7 strains belonged to subserotypes Ia, Ib and Ic, respectively. Formalin-killed cells of subserotype Ia and Ic strains were agglutinated with the anti-subserotype Ia serum (so far being used as an anti-serotype I serum) and Ic serum, respectively. Subserotype Ib strains were agglutinated with both sera. In PFGE analysis, the stocked 188 S. parauberis strains were divided into three clusters corresponding to subserotypes Ib/Ic, Ia and serotype II.

This study was conducted to select the media for the formalin killed vaccine (FKC) production of Vibrio harveyi and its application for olive flounder, Paralichthys olivaceus. For this, we have investigated the immune effects of Vibrio harveyi FKC vaccines grown in 3 different media i.e. Tryptic Soy Broth (TSB), TSB containing 2-2"-dipyridyl (TSB-D), Brain Heart Infusion Broth (BHIB) on the production of agglutinating antibody and protection against Vibrio harveyi in olive flounder. Additionally, a dual vaccine was prepared by combining Streptococcus parauberis vaccine to V. harveyi vaccine and its efficacy was also analyzed with the determination of optimal administration dosage. Consequently, olive flounder immunized with FKC grown in TSB-D showed the same protection with the vaccine grown in BHIB and the optimal dose of the vaccine was 10mg/kg of body weight. Indeed the dual vaccine showed higher agglutination titer and protection than control fish. The optimal dose for dual vaccine was 10mg/kg body weight for each of two vaccines.

Loop-mediated isothermal amplification (LAMP) technique relies on autocycling strand displacement DNA sysnthesis without template denaturation steps under isothermal conditions. LAMP has more advantages than modern PCR, as it requires only basic laboratory equipment like an isothermal water bath, oven, and heating cabinet. Hence, in this study, five random primers were designed with Streptococcus parauberis, shikimate kinase Arok gene sequences (Genbank accession number: CP0024711). Two primers were selected based on the ladder pattern. Other optimum reaction conditions like temperature, time, and sensitivity were established and confirmed with conventional SYBR-green PCR. Results confirmed that the designed random primers were species specific, without any non-target DNA amplification under isothermal conditions. Hence, this study suggests that LAMP techniques could be used in the diagnosis of fish pathogen, specifically S. parauberis.

A multiplex PCR assay for differentiation of Streptococcus parauberis serotypes was developed. The three primer pairs for subserotypes Ia and Ib/Ic and serotype II were designed from the serotype-specific sequences of the wzy gene in the loci for capsular polysaccharide biosynthesis. All of 188 S. parauberis isolates from Japanese flounder showed positive reaction with the expected size of PCR product for each serotype, which was consistent with the results of agglutination test using rabbit antisera. The nontypeable isolates, which are not differentiated by agglutination test, could be identified as subzero type Ib/Ic or serotype II. Other streptococci including S. parauberis derived from cow and major bacterial fish pathogens showed negative reaction., Fish Pathology, 50(4), pp.213-215; 2015

Streptococcus parauberis strain SP-llh was isolated from cows with mastitis in western China in 2015. The 2,522,235-bp genome sequence consists of 46 large contigs in 14 scaffolds and contains 2,620 predicted protein-coding genes, with a G+C content of 35.3%.

Streptococcus parauberis is a pathogen of cattle and fish, closely related Streptococcus uberis and Streptococcus iniae . We report the genomes of seven S. parauberis strains recovered from striped bass ( Morone saxatilis ) in the Chesapeake Bay. The availability of these genomes will allow comparative genomic analysis of Chesapeake Bay S. parauberis strains versus S. parauberis cultured from other animal hosts and geographic regions.

A multiplex PCR assay for differentiation of Streptococcus parauberis serotypes was developed. The three primer pairs for subserotypes Ia and Ib/Ic and sero- type II were designed from the serotype-specific sequences of the wzy gene in the loci for capsular polysaccharide biosynthesis. All of 188 S. parauberis isolates from Japanese flounder showed positive reaction with the expected size of PCR product for each serotype, which was consistent with the results of agglutination test using rabbit antisera. The nontypeable isolates, which are not differen- tiated by agglutination test, could be identified as subsero- type Ib/Ic or serotype II. Other streptococci including S. parauberis derived from cow and major bacterial fish patho- gens showed negative reaction.

Starry flounder, which are recently increasingly cultured in Korea, are known to highly vulnerable to Streptococcus parauberis infection. Five groups of starry flounder (n=30 for each group) were vaccinated with S. parauberis formalin-killed whole cells by intraperitoneal injection at a final concentration of 0, 0.01, 0.1, 1 and 10 mg fish -1 . Specific antibody production of 1 and 10 mg fish -1 administered groups significantly increased at four weeks post immunization. All vaccinated groups showed higher survival rates than a control group when five groups of fish were challenged with S. parauberis at a dose of 1.14×10 4 cfu fish -1 and 1.14×10 2 cfu fish -1 , respectively. In particular, 0.1 or higher concentrations of formalin killed bacterial cells are able to confer the fish high protection against S. parauberis infection.

In Japan, mass mortalities of cultured marine fish due to bacterial diseases have been often reported. Among diseases caused by Gram-positive bacteria, Lactococcus garvieae infection in yellowtail, amberjack, jack mackerel and striped jack, Streptococcus iniae infection in yellowtail, Japanese flounder and filefish, S. parauberis infection in Japanese flounder, Nocardia seriolae infection in yellowtail. Among diseases caused by Gram-negative bacteria, Edwardsiella tarda infection in Japanese flounder and red sea bream, Photobacterium damsela subsp. piscicida infection in yellowtail, red sea bream, striped jack and Japanese flounder, Tenacibaculum maritimum infection in Japanese flounder, red sea bream and striped jack, Vibrio spp. infection in various marine fishes cause serious damages every year. A commercially available L. garvieae and V. anguillarum vaccine is used in yellowtail and amberjack by oral administration or injection. S. iniae vaccine is used in Japanese flounder by injection. Recently, oil-based bivalent vaccine against P. damsela and L. garvieae infection was developed in yellowtail and amberjack. We have developed pentavalent vaccine against streptococcicosis and edwardsiellosis in Japanese flounder. Efficacy of pentavalent vaccine with formalin-killed S. parauberis (serotype I and Ⅱ strain), S. iniae and E. tarda (motile and non-motile strain) were tested by vaccination followed by intramuscular challenge with the each pathogen. The Japanese flounder vaccined with pentavalent vaccine showed higher resistance against the each pathogen on two weeks post-vaccination. These results revealed that pentavalent vaccine was effective in prevention against streptococcicosis and edwardsiellosis in cultured Japanese flounder.

The protective effect of dietary L-carnitine supplementation was accessed in turbot (Scophthalmusmaximus) challenged with Streptococcus parauberis. Infected fish fed 240mg L-carnitine showed significantly (P < 0.01) higher survival (78%) than infected fish fed 40mg L-carnitine (11%). When dietary L-carnitine supplementation was suspended, a significant increase in mortality was observed in infected fish. The present report documents that dietary L-carnitine fed on a continuous basis may act as an immunostimulant in turbot.

We evaluated the protective efficacy of a combined vaccine containing formalin-inactivated Edwardsiella tarda, Streptococcus iniae, and S. parauberis using olive flounder Paralichthys olivaceus. The fish were injected intraperitoneally with 0.1 mL of the combined vaccine. The fish were then challenged by intraperitoneal injection of the three bacterial strains. Mortality was significantly lower in the vaccinated groups compared with the control groups 3 wk post vaccination. Furthermore, there was a significant increase in antibody titer up to 16 wk after vaccination. Our results suggest that the combined vaccine successfully induced humoral and protective immunity against the three pathogens in olive flounder.

ヒラメの腹腔内，血管内，体側筋肉内，背鰭基部挙屈筋内，背鰭基部皮下または前鰓蓋外側皮下にS. parauberisを接種したところ，腹腔内接種では保菌状態になるものの，死亡個体が出現しない場合があった。腹腔内以外の接種法では比較的高い累積死亡率が得られたが，接種菌量と死亡率に正の関係が認められたのは皮下接種法だけであった。皮下接種感染魚の肉眼症状，病理組織ともに自然感染魚と異なる像は観察されなかったことから，皮下組織は感染実験の接種部位として妥当と思われる。

AIMS: To screen for the existence and determine the structure of Tn916-like element in Streptococcus parauberis serotype II strains isolated from cultured Japanese flounder in western Japan. METHODS AND RESULTS: In this study, the structure of Tn916-like element and the flanking regions were characterized by polymerase chain reaction (PCR) and inverse PCR, followed by cloning and sequencing. The Tn916-like element is 18 031 bp in length and composed of 22 ORFs. Southern blot hybridization analysis showed that the HincII-digested internal structures of Tn916-like elements yielded two patterns among S. parauberis serotype II strains. The flanking sequences were identical with the corresponding region of S. parauberis serotype I strain except for the addition of 6-bp coupling sequence (ATCATA) being adjacent to the upstream of the element. CONCLUSION: The Tn916-like element exhibited high homology (more than 99%) with Tn916 observed in other streptococci and enterococci and was integrated in the same site of chromosome for all of the tested S. parauberis serotype II strains. SIGNIFICANCE AND IMPACT OF THE STUDY: The results indicate that the Tn916-like element encoding tet(M) gene is present in all of the tested S. parauberis serotype II strains, which are disseminated in the flounder-culturing areas in western Japan., The definitive version is available at www.blackwell-synergy.com, Letters in applied microbiology, 48(6), pp.770-776; 2009

わが国のヒラメ由来 Streptococcus parauberis の血清学的性状を調べた結果，ウサギ抗血清を用いた凝集試験で 2 つの血清型に分けられた。各血清型のオートクレーブ抽出菌体抗原はそれぞれの血清型抗血清とゲル内沈降反応を示した。培養菌体からは酸性多糖が検出され，それぞれの血清型抗血清で前処理した培養菌体には透過型電顕で莢膜構造が観察された。以上のことから，莢膜抗原が血清型決定因子となっていることが示唆された。

Streptococcus parauberis strain SK-417 was isolated from the brain of a diseased Sebastes ventricosus , collected from an aquaculture farm in April 2013 in Kagoshima Prefecture, Japan. The draft genome sequence, obtained with a 454 GS Junior sequencing system, consists of 33 large contigs of >500 bp, totaling 1,958,836 bp, and has a G+C content of 35.4%.

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.

Several strains of Garm-positive short rod (coccibacilli)-shaped bacteria were isolated from diseased cultured turbot, Scophthalmus maximus (L.), in the North of Spain with lesions and signs of Spain with lesions and signs of streptocecosis. The α-haemolytic streptoceoccus-like organisms from diseased turbot were identified by physiological, biochemical and 16S rRNA gene sequence analysis as Streptococcus parauberis. This is the first report of S. parauberis associated with fish disease.

Streptococcosis is a common cause of pathology and mortality in fishes resulting in significant economic losses for the aquaculture industry. One etiologic agent of the disease, Streptococcus parauberis, has been associated with fish mortalities in Spain and Korea. Here we report the first identification of S. parauberis in wild finfish in Chesapeake Bay, USA. Gram-positive cocci were isolated from the spleens of striped bass, Morone saxatilis, and identified via species-specific primers and 16S rRNA gene sequencing. Biochemical characterization and antibiotic susceptibility tests were used to compare local isolates to isolates infecting aquacultured fishes and dairy cattle. This is also the first report of a plasmid in S. parauberis from any host.

The nucleotide sequences of 16S ribosomal-RNA of Streptococcus uberis types I and II were determined by reverse transcriptase. Comparative analysis of the sequence data showed that the two types are phylogenetically distinct and worthy of separate species status. A new species Streptococcus parauberis is proposed for the type II strains. The type strain of Strep. parauberis is NCDO 2020. and accepted 24 August 1989

Streptococcus sp. is gram-positive coccus that causes streptococcal infections in fish due to intensification of aquaculture and caused significant economic losses in fish farm industry. A streptococcal infection occurred from cultured diseased olive flounder (Paralichthys olivaceus) in May, 2005 at a fish farm in Jeju Island, Korea. The diseased flounder exhibited bilateral exophthalmic eyes and rotten gills; water temperature was 16-18 degrees C when samples were collected. Of the 22 fish samples collected, 3 samples were identified as Lactococcus garvieae and 18 samples were identified as Streptococcus parauberis by culture-based, biochemical test. Serological methods such as slide agglutination, hemolysis and antimicrobial susceptibility test were also used as well as multiplex PCR-based method to simultaneously detect and confirm the pathogens involved in the infection. S. parauberis and L. garvieae have a target region of 700 and 1100 bp., respectively. One fish sample was not identified because of the difference in the different biochemical and serological tests and was negative in PCR assay. In the present study, it showed that S. parauberis was the dominant species that caused streptococcosis in the cultured diseased flounder.