title: Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of lymphocystis disease virus.
abstract: A loop-mediated isothermal amplification (LAMP) assay was developed for the detection of lymphocystis disease virus (LCDV). A set of five specific primers, two inner and two outer primers and a loop primer, were designed on the basis of the major capsid protein gene of LCDV. The reaction time and temperatures were optimized for 60 min at 63 degrees C, respectively. LAMP amplification products were detected by a ladder-like appearance on agarose gel electrophoresis or a naked-eye inspection of a color change in the reaction tube by addition of SYBR Green I. The assay was specific for LCDV, and there was no cross-reactivity with white spot syndrome virus (WSSV) or six other Iridoviridae viruses (epizootic hematopoietic necrosis virus, EHNV; tiger frog virus, TFV; Bohle iridovirus, BIV; soft-shelled turtle iridovirus, STIV; infectious spleen and kidney necrosis virus, ISKNV; red sea bream iridovirus, RSIV). The detection limit of the LAMP assay was 15 fg, which was similar to that of real-time quantitative polymerase chain reaction (PCR) and 10-fold higher than the conventional PCR. The LAMP assay was evaluated using 109 clinical samples, and the results indicated the suitability and simplicity of the test as a rapid, field diagnostic tool for detection of LCDV. The LCDV LAMP assay has potential for early diagnosis of LCDV infection.
authors.affiliation: Technical Center of Shandong Entry-Exit Inspection and Quarantine Bureau, No. 70 Qutangxia Road, Qingdao 266002, China.
authors.forename: Qiong
authors.initials: Q
authors.lastname: Li
authors.forename: Zhiqin
authors.initials: Z
authors.lastname: Yue
authors.forename: Hong
authors.initials: H
authors.lastname: Liu
authors.forename: Chengzhu
authors.initials: C
authors.lastname: Liang
authors.forename: Xiaolong
authors.initials: X
authors.lastname: Zheng
authors.forename: Yuran
authors.initials: Y
authors.lastname: Zhao
authors.forename: Xiao
authors.initials: X
authors.lastname: Chen
authors.forename: Xizhi
authors.initials: X
authors.lastname: Xiao
authors.forename: Changfu
authors.initials: C
authors.lastname: Chen
mesh_terms: D000818:Animals; D052160:Benzothiazoles; D036022:Capsid Proteins / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004266:DNA Virus Infections / Q000175:diagnosis / Q000662:veterinary*; D003959:Diamines; D004587:Electrophoresis, Agar Gel; D005393:Fish Diseases / Q000175:diagnosis* / Q000821:virology; D005412:Flatfishes / Q000821:virology*; D005456:Fluorescent Dyes / Q000494:pharmacology; D007497:Iridoviridae / Q000235:genetics / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009930:Organic Chemicals / Q000494:pharmacology; D011804:Quinolines; D012680:Sensitivity and Specificity; D013194:Staining and Labeling / Q000379:methods; D013696:Temperature; D013997:Time Factors
chemical_list: D052160:Benzothiazoles; D036022:Capsid Proteins; D017931:DNA Primers; D003959:Diamines; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I