Diseases of Aquatic Organisms | |
Molecular detection methods developed for a systemic rickettsia-like bacterium (RLB) in Penaeus monodon (Decapoda: Crustacea) | |
Donald V. Lightner1  Bonnie Poulos1  Marc Le Groumellec1  Rita Redman1  Linda M. Nunan1  | |
关键词: Rickettsia; Penaeus monodon; Shrimp; Crustacea; | |
DOI : 10.3354/dao053015 | |
学科分类:生物科学(综合) | |
来源: Inter-Research | |
【 摘 要 】
ABSTRACT: Molecular detection methods were developed to aid in the diagnosis of a rickettsia-like bacterium (RLB) which caused severe mortalities of farm-raised Penaeus monodon in Madagascar. Using primers derived from the 16S rRNA gene ofbacteria, a PCR assay was optimized to amplify this region of the genome of the RLB, using extracted DNA from infected P. monodon tissue as the template. The resulting amplified PCR product was sequenced and 2 novel primers were selected from thevariable region of the gene. These primers amplified a 532 bp fragment of DNA originating from the rickettsia-infected samples. The PCR assay was optimized and tested on DNA extracted from specific pathogen-free (SPF) P. vannamei tissue and severalother strains of bacteria. The PCR assay with the rickettsia-specific primers was specific for this RLB and did not amplify the other DNA samples tested. The 532 bp PCR-amplified fragment was labeled with digoxigenin (DIG) for in situhybridization assays. This probe was tested on SPF, RLB and bacteria-infected shrimp specimens preserved in Davidson¹s fixative. The probe was specific for both natural and experimental rickettsial infections. Hybridization with this probe required astringent temperature of 65°C, otherwise cross-reactivity was observed with other types of bacteria.
【 授权许可】
Unknown
【 预 览 】
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RO201911300203469ZK.pdf | 940KB | download |