Aeromonas detection and characterization using genus-specific PCR and single-strand conformation polymorphism (SSCP).
Based on sequence alignment, oligonucleotide primers targeting the Aeromonas extracellular lipase gene were developed for PCR detection of member of the genus. A pair of primers designed for conserved regions of the gene amplified a 276 bp sequence in all Aeromonas species and tested strains, but did not have a positive result with other Gram-positive and Gram-negative bacteria, showing high specificity and sensitivity. Selective enrichment in alkaline peptone water, followed by centrifugation, and direct usage of cells suspension as template, detected initial populations of 10 c.f.u. ml⁻¹. Single-strand conformation polymorphism analysis of the PCR products allowed the characterization of Aeromonas strains with a high discriminatory power (Simpson's index = 0.988). The method presented here provides a useful tool for the rapid detection of Aeromonas and the characterization of Aeromonas isolates.
Institute of Biotechnology, University of Caxias do Sul, R. Francisco G. Vargas 1130, Caxias do Sul, Rio Grande do Sul 95001-970, Brazil. aplongar@yahoo.com
Ana Paula Longaray
AP
Delamare
Roberto Francisco
RF
Lucena
Guilherme
G
Thomazi
Shana
S
Ferrarini
Jucimar
J
Zacaria
Sergio
S
Echeverrigaray
D000333:Aeromonas / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics; D005798:Genes, Bacterial; D008049:Lipase / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D018807:Polymorphism, Single-Stranded Conformational; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012680:Sensitivity and Specificity; D016415:Sequence Alignment / Q000379:methods; D017422:Sequence Analysis, DNA
D017931:DNA Primers; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D008049:Lipase