| The expanding recurrence of invasive fungal infection and the high mortality associated with dispersed fungal disease have highlighted the requirement for fast recognizable proof of infectious molds. The current study was conducted to identify the most prevalent mycotic diseases among cultured freshwater fishes with special focus on isolation, molecular identification and antimicrobial sensitivity testing. A total number of 300 examined cultured freshwater fishes; 200 Oreochromis niloticus (120 apparently healthy and 80 showing clinical signs) and 100 Clarias gariepinus (30 apparently healthy and 70 showing clinical signs), were collected alive from five freshwater fish farms at Kafr El Sheikh Governorate all over the year 2018. Clinically, examined Oreochromis niloticus revealed scale detachment, fins erosion, pale body coloration and eye opacity particularly in fish naturally infected with Aphanomycess, Aspergillus, Pencillium and Fusarium; however, infected Clarias gariepinus showed scattered hemorrhagic patches on the ventral abdomen. Mycological examination revealed the isolation of 900 fungal isolates; 570 fungal isolates from Oreochromis niloticus and 330 isolates from C. gariepinus. Aphanomyces invadans was the predominant fungi among diseased fishes mainly from skin, fins and gills. However, Aspergillus flavus, Aspergillus niger and Alternaria sp. were the most predominant fungi isolated from apparently healthy fishes mainly from internal organs. Molecular identification was performed using inter-transcriped spacer (ITS) gene for the most prevalent isolates among either diseased or apparently healthy fishes. Developed PCR assay specific primer detect not only Aphanomyces invadans showing clear bands at 569 bp molecular weight, but also Asperigillus flavus and Asperigillus niger showing clear bands at 595 and 600 bp molecular weight, respectively. Anti-fungal sensitivity test revealed that all mould isolates are sensitive to Nystatin and the majority was resistant to Voriconazole. Briefly, being normal mycoflora, most fungi still can cause infections to fishes. Consequently, proper health management practices should be adapted while fish rearing.Citation | El-Tawab AAA, El-Hofy FI, Moustafa EM, Halawa MR (2020). Insight into isolation, identification and antimicrobial sensitivity of some moulds isolated from fresh water fishes. Adv. Anim. Vet. Sci. 8(2): 174-182.