Molecular identification of Vermamoeba vermiformis from freshwater fish in lake Taal, Philippines

Abstract

Free Living Amoebae (FLA) are considered ubiquitous. FLAs may infect various biological organisms which act as reservoir hosts. Infected freshwater fishes can pose a public health concern due to possible human consumption. This study aims to identify possible pathogenic FLAs present in freshwater fishes. Seventy five (75) Oreochromis niloticus were studied for the presence of FLAs. Fish organs were suspended in physiologic saline pelleted and cultured in non-nutrient agar (NNA) lawned with Escherichia coli and were incubated in 33 °C for 14 days. Eighteen (18) fish gills and nineteen (19) fish intestine samples presented with positive growth. Trophozoites and cystic stages of FLAs were subcultured until homogenous growth was achieved. Cells were harvested from cultured plates and DNA was extracted using Chelex resin. DNA was subjected to polymerase chain reaction using universal forward primer EukA and reverse primer EukB targeting the 18s RNA. Of the 37 plates that presented with positive amoebic growth, 9 samples showed the presence of DNAs and were sent for further purification and sequencing. Basic Local Alignment Search Tool (BLAST) results showed that protists isolated from fish organs in Lake Taal include: Eocercomonas (HM536152), Colpoda steinii (KJ607915) and Vermamoeba vermiformis (KC161965). The results showed that fresh-water fishes can harbour FLAs in the gut. It is proposed that freshwater reservoirs utilized for aquaculture be monitored for the presence of FLAs and extensive study be conducted on the pathogenicity of bacterial endosymbionts and infecting viruses to its mammalian and non-mammalian host.