Sensitive and rapid detection of Vibrio corallilyticus by loop-mediated isothermal amplification targeted to the alpha subunit gene of RNA polymerase.
OBJECTIVE
A diagnostic protocol was developed for rapid detection of Vibrio corallilyticus by method of loop-mediated isothermal amplification (LAMP).


RESULTS
For cloning and sequencing of rpo A gene of V. corallilyticus, a set of four LAMP primers were designed by targeting the rpoA gene. With Bst DNA polymerase, the reaction time and temperature were optimized for 70 min at 65 degrees C, respectively. The amplification products were detected by electrophoresis. The detection limit of V. corallilyticus by LAMP was 3.6 x 10(3) CFU ml(-1) (8 CFU per reaction), but PCR could detect up to 3.6 x 10(4) CFU ml(-1) (72 CFU per reaction). The LAMP method was ninefold more sensitive than conventional PCR. The results also indicated that the LAMP reaction was highly specific to V. corallilyticus.


CONCLUSIONS
The LAMP assay was a sensitive, specific and cost-effective method for the rapid detection of V. corallilyticus.


CONCLUSIONS
This LAMP method provides an important diagnostic tool for the detection of V. corallilyticus infection. It can replace laborious biochemical tests for the identification of V. corallilyticus.
Liu
G F
GF
School of Life Sciences, Sun Yat-sen University, Guangzhou, China.
Wang
J Y
JY
Xu
L W
LW
Ding
X
X
Zhou
S N
SN
D001426:Bacterial Proteins / Q000235:genetics*; D001431:Bacteriological Techniques / Q000191:economics / Q000379:methods*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D004586:Electrophoresis; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000379:methods*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013696:Temperature; D013997:Time Factors; D014733:Vibrio / Q000235:genetics / Q000302:isolation & purification*; D014735:Vibrio Infections / Q000175:diagnosis*
D001426:Bacterial Proteins; D017931:DNA Primers; D004269:DNA, Bacterial; D012321:DNA-Directed RNA Polymerases; C113275:RNA polymerase alpha subunit