Simultaneous evaluation of four PCR primer sets for the diagnosis of Streptococcus phocae infection

Resultado de la investigación: Article

10 Citas (Scopus)

Resumen

The sensitivity of 4 published primer pairs for the detection of Streptococcus phocae strains was evaluated. Primer sets cpn60-F and -R and sodA-F and -R correctly identified S. phocae. Correct identification was also achieved with the primer pairs cae1-cae2 and PX1-PX2, but using the reverse complementary version of both reverse primers (caeVQ2 and PXVQ2, respectively). Among the 4 PCR protocols with pure and mixed cultures, the primer pair PX1-PXVQ2 provided the highest level of sensitivity for S. phocae (10 2 and 104 cells per PCR tube), and detection was 10- to 100-fold higher than the other 3 primer pairs. When the cae1-caeVQ2 and PX1-PXVQ2 PCR protocols were applied to different seeded Atlantic salmon tissues (spleen, kidney and liver), the detection limit achieved was 5.1 × 105 to 6.4 × 107 CFU g-1, and the lowest sensitivity detected was 1.18 × 106 S. phocae per tube (which corresponds to 6.4 × 107 CFU g-1) in spleen samples using PX1-PXVQ2. In the kidney samples seeded with S. phocae strains, regardless of the primer set used, the PCR sensitivity was the same (7.31 ± 1.5 × 106 CFU g-1). In addition, the nested PCR assay using the primer pair PX1-PXVQ2 improved the sensitivity of detection of S. phocae by at least 100 times compared to the first round PCR, not only in mixed and pure suspensions, but also in experimentally seeded fish tissues. The picked tissues that allowed the easiest detection of S. phocae were the liver, kidney and spleen, respectively. Thus, the nested PCR approach is an important tool for the rapid and reliable diagnosis of streptococcosis due to S. phocae.

Idioma originalEnglish
Páginas (desde-hasta)217-222
Número de páginas6
PublicaciónDiseases of Aquatic Organisms
Volumen82
N.º3
DOI
EstadoPublished - 22 dic 2008

Huella dactilar

Streptococcus phocae
infection
spleen
kidneys
liver
detection
evaluation
assay
mixed culture
fold
Salmo salar
fish
detection limit

ASJC Scopus subject areas

  • Aquatic Science
  • Ecology, Evolution, Behavior and Systematics

Citar esto

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title = "Simultaneous evaluation of four PCR primer sets for the diagnosis of Streptococcus phocae infection",
abstract = "The sensitivity of 4 published primer pairs for the detection of Streptococcus phocae strains was evaluated. Primer sets cpn60-F and -R and sodA-F and -R correctly identified S. phocae. Correct identification was also achieved with the primer pairs cae1-cae2 and PX1-PX2, but using the reverse complementary version of both reverse primers (caeVQ2 and PXVQ2, respectively). Among the 4 PCR protocols with pure and mixed cultures, the primer pair PX1-PXVQ2 provided the highest level of sensitivity for S. phocae (10 2 and 104 cells per PCR tube), and detection was 10- to 100-fold higher than the other 3 primer pairs. When the cae1-caeVQ2 and PX1-PXVQ2 PCR protocols were applied to different seeded Atlantic salmon tissues (spleen, kidney and liver), the detection limit achieved was 5.1 × 105 to 6.4 × 107 CFU g-1, and the lowest sensitivity detected was 1.18 × 106 S. phocae per tube (which corresponds to 6.4 × 107 CFU g-1) in spleen samples using PX1-PXVQ2. In the kidney samples seeded with S. phocae strains, regardless of the primer set used, the PCR sensitivity was the same (7.31 ± 1.5 × 106 CFU g-1). In addition, the nested PCR assay using the primer pair PX1-PXVQ2 improved the sensitivity of detection of S. phocae by at least 100 times compared to the first round PCR, not only in mixed and pure suspensions, but also in experimentally seeded fish tissues. The picked tissues that allowed the easiest detection of S. phocae were the liver, kidney and spleen, respectively. Thus, the nested PCR approach is an important tool for the rapid and reliable diagnosis of streptococcosis due to S. phocae.",
keywords = "Diagnosis, Nested PCR, PCR, Streptococcus phocae",
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TY - JOUR

T1 - Simultaneous evaluation of four PCR primer sets for the diagnosis of Streptococcus phocae infection

AU - Avendaño-Herrera, R.

PY - 2008/12/22

Y1 - 2008/12/22

N2 - The sensitivity of 4 published primer pairs for the detection of Streptococcus phocae strains was evaluated. Primer sets cpn60-F and -R and sodA-F and -R correctly identified S. phocae. Correct identification was also achieved with the primer pairs cae1-cae2 and PX1-PX2, but using the reverse complementary version of both reverse primers (caeVQ2 and PXVQ2, respectively). Among the 4 PCR protocols with pure and mixed cultures, the primer pair PX1-PXVQ2 provided the highest level of sensitivity for S. phocae (10 2 and 104 cells per PCR tube), and detection was 10- to 100-fold higher than the other 3 primer pairs. When the cae1-caeVQ2 and PX1-PXVQ2 PCR protocols were applied to different seeded Atlantic salmon tissues (spleen, kidney and liver), the detection limit achieved was 5.1 × 105 to 6.4 × 107 CFU g-1, and the lowest sensitivity detected was 1.18 × 106 S. phocae per tube (which corresponds to 6.4 × 107 CFU g-1) in spleen samples using PX1-PXVQ2. In the kidney samples seeded with S. phocae strains, regardless of the primer set used, the PCR sensitivity was the same (7.31 ± 1.5 × 106 CFU g-1). In addition, the nested PCR assay using the primer pair PX1-PXVQ2 improved the sensitivity of detection of S. phocae by at least 100 times compared to the first round PCR, not only in mixed and pure suspensions, but also in experimentally seeded fish tissues. The picked tissues that allowed the easiest detection of S. phocae were the liver, kidney and spleen, respectively. Thus, the nested PCR approach is an important tool for the rapid and reliable diagnosis of streptococcosis due to S. phocae.

AB - The sensitivity of 4 published primer pairs for the detection of Streptococcus phocae strains was evaluated. Primer sets cpn60-F and -R and sodA-F and -R correctly identified S. phocae. Correct identification was also achieved with the primer pairs cae1-cae2 and PX1-PX2, but using the reverse complementary version of both reverse primers (caeVQ2 and PXVQ2, respectively). Among the 4 PCR protocols with pure and mixed cultures, the primer pair PX1-PXVQ2 provided the highest level of sensitivity for S. phocae (10 2 and 104 cells per PCR tube), and detection was 10- to 100-fold higher than the other 3 primer pairs. When the cae1-caeVQ2 and PX1-PXVQ2 PCR protocols were applied to different seeded Atlantic salmon tissues (spleen, kidney and liver), the detection limit achieved was 5.1 × 105 to 6.4 × 107 CFU g-1, and the lowest sensitivity detected was 1.18 × 106 S. phocae per tube (which corresponds to 6.4 × 107 CFU g-1) in spleen samples using PX1-PXVQ2. In the kidney samples seeded with S. phocae strains, regardless of the primer set used, the PCR sensitivity was the same (7.31 ± 1.5 × 106 CFU g-1). In addition, the nested PCR assay using the primer pair PX1-PXVQ2 improved the sensitivity of detection of S. phocae by at least 100 times compared to the first round PCR, not only in mixed and pure suspensions, but also in experimentally seeded fish tissues. The picked tissues that allowed the easiest detection of S. phocae were the liver, kidney and spleen, respectively. Thus, the nested PCR approach is an important tool for the rapid and reliable diagnosis of streptococcosis due to S. phocae.

KW - Diagnosis

KW - Nested PCR

KW - PCR

KW - Streptococcus phocae

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C2 - 19244973

AN - SCOPUS:58949094205

VL - 82

SP - 217

EP - 222

JO - Diseases of Aquatic Organisms

JF - Diseases of Aquatic Organisms

SN - 0177-5103

IS - 3

ER -