A Multiplex PCR Assay for Different-iation of &lt;i&gt;Streptococcus parauberis&lt;/i&gt; Serotypes

Tu, Chuandeng; Suga, K.; Kanai, Kazuaki

doi:10.3147/jsfp.50.213

Cited by 12 publications

(12 citation statements)

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“…Consistent with our findings, the profile of 22 antibiotic resistance genes (ARGs) from effluents of 20 coastal aquaculture were observed in Jeju and South Jeolla province (Jang et al, 2018). In this study, using a previously described PCR‐based method (Tu et al, 2015), we examined the prevalence of all three serotypes (Ia, Ib/Ic and II) of S. parauberis (Kanai et al, 2015) in diseased P. olivaceus , P. stellatus and S. schlegelii . Our finding showed that S. parauberis serotype Ia (74.7%) was more abundant than serotypes Ib/Ic (19.3%) and II (6.0%); this corroborates a previous study where 122 S. parauberis strains showed serotype I (64%) to be more predominant than serotype II (36%) in Korea (Park et al, 2016).…”

Section: Discussionmentioning

confidence: 99%

“…The three capsular polysaccharide serotypes (Ia, Ib/Ic and II) were identified by multiplex PCR using a previously reported method (Tu et al, 2015). Briefly, the forward primer For‐Ia (5′‐ATT GTT AGT CAT TCA GTT GT‐3′), For‐Ib/Ic (5′‐ATT TCT ACC AGG TTA CTT TG‐3′), For‐II (5′‐GAA CTA CTT AGG TTT AGC AT‐3′), the reverse primer Rev‐Ia (5′‐AAT TAT AGT CAA CAG TCC AG‐3′), Rev‐Ib/Ic (5′‐ACA TCT CGA AAC TTC ATA TT‐3′), and Rev‐II (5′‐AAC TTG TAA ATA GGA TTG CT‐3′) were designed to amplify a 213, 303 and 413 bp for S. parauberis of serotype Ia, Ib/Ic and II respectively.…”

Section: Methodsmentioning

confidence: 99%

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Prevalence, antibiotic susceptibility and serotyping of Streptococcus parauberis isolates from diseased marine fish

et al. 2021

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Streptococcus parauberis is the main causative bacterial pathogen for streptococcosis in marine fish, causing a significant economic loss to the mariculture industry. In this study, we aimed to investigate the serotypes and antimicrobial susceptibilities of 83 S. parauberis isolates from diseased farmed fish (Paralichthys olivaceus, Platichthys stellatus and Sebastes schlegelii) in Korea. This study categorized the S. parauberis isolates based on PCR serotyping into serotype I (subtypes Ia and Ib/Ic) and serotype II. Serotype Ia was identified to be significantly more prevalent than serotypes Ib/Ic and II. An antimicrobial susceptibility analysis showed that all of the isolates were susceptible to amoxicillin/clavulanic acid. Overall, the isolates studied showed resistance to penicillin and ampicillin (1.2%), tetracycline (68.7%), oxytetracycline (72.3%), doxycycline (53.0%) and erythromycin (37.3%). Interestingly, we found that serotypes Ia and II showed higher degrees of tetracycline resistance than serotype Ib/Ic. The macrolide resistance gene erm(B) was present in 75.8% of the Ia isolates, whereas the mef(A) gene was not detected in any of the isolates. The tetracycline resistance genes tet(S) and tet(M) were detected in 95.2% and 60% of Ia and II isolates respectively. Moreover, β‐lactam antibiotic‐resistant Ia isolates contained a set of 11 mutations in PBP1A and PBP2X. Our study provides a valuable reference for surveillance of antimicrobial resistance and underscores the strict management of antibiotic use to prevent the emergence of resistant of S. parauberis serotypes.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Prevalence, antibiotic susceptibility and serotyping of Streptococcus parauberis isolates from diseased marine fish

et al. 2021

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“…These limitations added to the recent determination of complete genomic sequences of a wide variety of bacteria have allowed the development of new molecular methods based on polymerase chain reaction (PCR) for bacterial typing. In recent years, different PCR bacterial typing protocols have emerged as an alternative for the use of antisera in pathogenic fish species, such as Streptococcus agalactiae (Demczuk et al 2017;Kannika et al 2017;Shoemaker et al 2017), Flavobacterium psychrophylum (Rochat et al 2017), S. parauberis (Tu et al 2015b), and Lactococcus garvieae (Ohbayashi et al 2017).…”

Section: Introductionmentioning

confidence: 99%

Comparative genomics of Streptococcus parauberis: new target for molecular identification of serotype III

Torres‐Corral

Santos

2020

Appl Microbiol Biotechnol

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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 × 10 2 gene copies (equivalent to 3.8 × 10 −9 ng/μl) using pure bacterial cultures of S. parauberis serotype III and 1.76 × 10 2 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.

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“…Traditionally, serotyping of bacterial isolates was performed for both diagnostic and epidemiological purposes (Buller, ; Shin et al, ), but serotyping has limited ability to discriminate between isolates and the need to obtain antisera from animals would be a major constraint. The identification and typing of microorganisms based on the amplification of bacterial DNA are commonly used in medical and biological research (Hassan, Khan, Abdulmawjood, & Lämmler, ; Jung, Chang, & Kim, ; Mata, Gibello et al, ; Nguyen, Lim, Kim, & Austin, ; Tu, Suga, & Kanai, ). These methods give species‐specific identifications and information on the genetic relatedness of strains, source of infection, and virulence of strains, geographical and host distribution of possible variants.…”

Section: Introductionmentioning

confidence: 99%

“…The identification and typing of microorganisms based on the amplification of bacterial DNA are commonly used in medical and biological research (Hassan, Khan, Abdulmawjood, & Lämmler, 2001;Jung, Chang, & Kim, 2010;Nguyen, Lim, Kim, & Austin, 2016;Tu, Suga, & Kanai, 2015). These methods give speciesspecific identifications and information on the genetic relatedness of strains, source of infection, and virulence of strains, geographical and host distribution of possible variants.…”

mentioning

confidence: 99%

Identification and typing of Vagococcus salmoninarum using genomic and proteomic techniques

Torres‐Corral

Santos

2019

Journal of Fish Diseases

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This study reports on the characterization of Vagococcus salmoninarum using phenotypic, serological, antigenic, genetic and proteomic methods. All strains of V. salmoninarum were resistant to most of the antimicrobials tested, and only 10% of strains were sensitive to florfenicol. Serological analysis demonstrated a high antigenic homogeneity within the species. No cross‐reaction was detected with other fish pathogenic species causing streptococcosis (Lactococcus garvieae, Streptococcus parauberis, Streptococcus iniae, Streptococcus agalactiae, Carnobacterium maltaromaticum) using serum against V. salmoninarum CECT 5810. Electrophoretic analysis of cell surface proteins and immunoblot supported the antigenic homogeneity within V. salmoninarum strains. Moreover, limited diversity was detected using genomic (RAPD, ERIC‐PCR and REP‐PCR) and MALDI‐TOF‐MS analyses. The phenotypic, genomic and proteomic methods tested allowed the rapid differentiation of V. salmoninarum from the other species causing streptococcosis. However, MALDI‐TOF‐MS is the most promising method for typing and characterization of V. salmoninarum.

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A Multiplex PCR Assay for Different-iation of <i>Streptococcus parauberis</i> Serotypes

Cited by 12 publications

References 8 publications

Prevalence, antibiotic susceptibility and serotyping of Streptococcus parauberis isolates from diseased marine fish

Prevalence, antibiotic susceptibility and serotyping of Streptococcus parauberis isolates from diseased marine fish

Comparative genomics of Streptococcus parauberis: new target for molecular identification of serotype III

Identification and typing of Vagococcus salmoninarum using genomic and proteomic techniques

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