A Two-Step PCR Workflow Reveals Predominance of Flavobacterium covae in Ornamental Fish With Clinical Signs of...

Columnaris is an important bacterial disease of freshwater fish and is increasingly recognised as being caused by a species complex historically referred to as Flavobacterium columnare. A two-step molecular workflow was applied to ornamental fish with clinical signs of columnaris disease from a multispecies commercial distributor operating a shared recirculating system in São Paulo, Brazil. Skin and fin lesion scrapings were Gram stained, and lesion-derived tissue DNA was screened by ISR-PCR for the columnaris disease complex. ISR-PCR yielded the expected amplicon in 61/123 samples (49.6%) from 28 ornamental fish species. All ISR-PCR-positive samples were then analysed by multiplex PCR for species assignment. Group-specific amplicons were obtained from 29/61 samples (47.5%): Flavobacterium covae in 27/61 (44.3%) and F. davisii in 2/61 (3.3%); no GG1 or GG4 amplicons were detected. The remaining 32/61 samples (52.5%) were not typeable by multiplex PCR. These findings provide baseline species-level data and support combined molecular surveillance in multispecies ornamental fish systems.