title: Development of a quantitative PCR assay for monitoring Streptococcus agalactiae colonization and tissue tropism in experimentally infected tilapia.
abstract: Streptococcus agalactiae has become one of the most important emerging pathogens in the aquaculture industry and has resulted in large economic losses for tilapia farms in China. In this study, three pairs of specific primers were designed and tested for their specificities and sensitivities in quantitative real-time polymerase chain reactions (qPCRs) after optimization of the annealing temperature. The primer pair IGS-s/IGS-a, which targets the 16S-23S rRNA intergenic spacer region, was finally chosen, having a detection limit of 8.6 copies of S. agalactiae DNA in a 20 μL reaction mixture. Bacterial tissue tropism was demonstrated by qPCR in Oreochromis niloticus 5 days post-injection with a virulent S. agalactiae strain. Bacterial loads were detected at the highest level in brain, followed by moderately high levels in kidney, heart, spleen, intestines, and eye. Significantly lower bacterial loads were observed in muscle, gill and liver. In addition, significantly lower bacterial loads were observed in the brain of convalescent O. niloticus 14 days post-injection with several different S. agalactiae strains. The qPCR for the detection of S. agalactiae developed in this study provides a quantitative tool for investigating bacterial tissue tropism in infected fish, as well as for monitoring bacterial colonization in convalescent fish.
authors: Key Laboratory for Aquatic Products Safety of Ministry of Education/State Key Laboratory of Biocontrol, The School of Life Sciences, Sun Yat-sen University, Haizhu District, Guangzhou, Guangdong Province, China.|Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization, Ministry of Agriculture, South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, Guangdong Province, China.
authors: Y-L
authors: YL
authors: Su
authors: Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization, Ministry of Agriculture, South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, Guangdong Province, China.
authors: J
authors: J
authors: Feng
authors: Key Laboratory for Aquatic Products Safety of Ministry of Education/State Key Laboratory of Biocontrol, The School of Life Sciences, Sun Yat-sen University, Haizhu District, Guangzhou, Guangdong Province, China.
authors: Y-W
authors: YW
authors: Li
authors: Guangzhou Airport Extry-Exit Inspection and Quarantine Bureau, Guangzhou, Guangdong Province, China.
authors: J-S
authors: JS
authors: Bai
authors: Key Laboratory for Aquatic Products Safety of Ministry of Education/State Key Laboratory of Biocontrol, The School of Life Sciences, Sun Yat-sen University, Haizhu District, Guangzhou, Guangdong Province, China.
authors: A-X
authors: AX
authors: Li
mesh_terms: D000818:Animals; D058491:Bacterial Load; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D005393:Fish Diseases / Q000175:diagnosis / Q000382:microbiology*; D060888:Real-Time Polymerase Chain Reaction / Q000592:standards / Q000662:veterinary*; D012680:Sensitivity and Specificity; D013290:Streptococcal Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D013292:Streptococcus agalactiae / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D017210:Tilapia; D056189:Viral Tropism
chemical_list: D021903:DNA, Ribosomal Spacer
keywords: Oreochromis niloticus; Streptococcus agalactiae; bacterial colonization; quantitative real-time PCR