The adaptative responses and divergent evolution shown in the environments habited by the Cichlidae family allow to understand different biological properties, including fish genetic diversity and structure studies. In a zone that has been historically submitted to different anthropogenic pressures, this study assessed the genetic diversity and population structure of cichlid Caquetaia kraussii , a sedentary species with parental care that has a significant ecological role for its contribution to redistribution and maintenance of sedimentologic processes in its distribution area. This study developed de novo 16 highly polymorphic species-specific microsatellite loci that allowed the estimation of the genetic diversity and differentiation in 319 individuals from natural populations in the area influenced by the Ituango hydroelectric project in the Colombian Cauca River. Caquetaia kraussii exhibits high genetic diversity levels (Ho: 0.562–0.885; He: 0.583–0.884) in relation to the average neotropical cichlids and a three group-spatial structure: two natural groups upstream and downstream the Nechí River mouth, and one group of individuals with high relatedness degree, possibly independently formed by founder effect in the dam zone. The three genetic groups show recent bottlenecks, but only the two natural groups have effective population size that suggest their long-term permanence. The information generated is relevant not only for management programs and species conservation purposes, but also for broadening the available knowledge on the factors influencing neotropical cichlids population genetics.