Anisakis pegreffii: A quantitative fluorescence PCR assay for detection in situ

• Published in: Experimental parasitology Vol. 127; no. 2; pp. 587 - 592
• Main Authors: Fang, Wenzhen, Liu, Fan, Zhang, Shaolei, Lin, Junhua, Xu, Shisan, Luo, Damin
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier Inc 01-02-2011; Elsevier
• Subjects: Animals; Anisakiasis - diagnosis; Anisakiasis - parasitology; Anisakiasis - veterinary; Anisakis; Anisakis - genetics; Anisakis - isolation & purification; Anisakis pegreffii; Biological and medical sciences; DNA Primers; DNA Probes; DNA, Ribosomal; DNA, Ribosomal Spacer - genetics; Fish Diseases - diagnosis; Fish Diseases - parasitology; Fishes; Fluorescent Dyes; Food-borne parasitic zoonosis; Fundamental and applied biological sciences. Psychology; In situ; Invertebrates; Life cycle. Host-agent relationship. Pathogenesis; Marine; Nemathelminthia. Plathelmintha; Nematode; Polymerase Chain Reaction - methods; Protozoa; Reproducibility of Results; Second internal transcribed spacer (ITS-2); Sensitivity and Specificity; TaqMan real-time PCR; ISEW, Pacific, Taiwan Strait; ISEW, China, People's Rep., Minnan-Taiwan Bank; Anisakis pegreffii; Nematode; Second internal transcribed spacer (ITS-2); TaqMan real-time PCR; Food-borne parasitic zoonosis; In situ; Fluorescence; Parasite; Zoonosis; Infection; Polymerase chain reaction; Helmintha; Nemathelminthia; Invertebrata; Detection; Nematoda
• ISSN: 0014-4894; 1090-2449
• DOI: 10.1016/j.exppara.2010.11.008

[Display omitted]
► A real-time PCR method for the detection in situ of Anisakis pegreffii was developed. ► The ITS-2 of rDNA of A. pegreffii was used as molecular marker. ► The assay was capable of detecting 1/3 L3 in 30mg of fish tissue.
To facilitate improved diagnosis and detection of the third stage larva (L3) of Anisakis pegreffii from the Minnan-Taiwan bank fishing ground in Taiwan Strait, a real-time PCR method for the detection in situ and differentiation was developed to amplify a region of the second internal transcribed spacer (ITS-2) of this parasite. The real-time PCR assay was capable of detecting 1/3 of a single L3 in 30mg of marine fish tissue, and also exhibited a high level of specificity for A. pegreffii, no fluorescence signals were observed in other five major larval anisakid species found in commercial marine fishes caught in this fishing ground.