The bacterial microbiome is an important component of any aquaculture environment. The association and interaction among different microorganisms in finfish aquaculture environments during infections are rarely explored. However, the interaction between the bacterial microbiome and other microorganisms (e.g. parasites, viruses, or other bacteria) in aquaculture can prevent or contribute to disease outbreaks. This study characterised the bacterial composition associated with the abundance of a ciliated protozoan parasite, Chilodonella hexasticha, and their link with mortalities in a freshwater barramundi, Lates calcarifer, farm situated in tropical Queensland, Australia over one year. An environmental DNA (eDNA) approach was used to estimate the abundance of C. hexasticha (copies/µl) in water through SSU-rDNA gene qPCR and the relative abundance of bacterial species in water and fish gills through 16S rRNA V3 and V4 metabarcoding. The overall bacterial community diversity, dominated by Actinobacteria (42%), Proteobacteria (28%), Bacteroidetes (10%) and Cyanobacteria (6%), was stable among ponds over the study period (P > 0.05). Of those that could be identified to species level, Flavobacterium columnare, Veillonella dispar and Bdellovibrio bacteriovorus abundance correlated with both high C. hexasticha levels in pond water and high observed fish mortalities (P < 0.05). Results also revealed significantly higher levels of F. columnare, B. bacteriovorus, Plesiomonas shigelloides, Prostecobactor debontii and Oxalobacter formigenes (P < 0.05) in gills of fish with high infection levels of C. hexasticha compared to fish with no detected parasite infection. This study demonstrated, for the first time, a link between increased parasitic ciliate abundance, bacterial composition and fish mortalities in a freshwater aquaculture environment and the application of eDNA to investigate pathogen, host and environment interactions.