A multi-piece locking fastener assembly including a thickened base layer having a first wider diameter passageway exhibiting a first interiorly threaded orientation and a second narrower diameter passageway exhibiting a second opposite threaded orientation and extending a distance in communication with and beyond a bottom end wall of the first wider passageway. A first bolt fastener has a first arrangement of exterior threads engageable with the wider diameter passageway of the base layer. The first fastener further includes an interior passageway. Following installation of the wider fastener into the base layer, a second narrower diameter bolt fastener having at least a partially threaded stem is threadably inter-engaged with the second passageway in the base layer to prevent loosening of either fastener via their counter-threaded orientations.