This paper presents the world's first field trial of a 40-km reach and over 512-split symmetric-rate 40-Gbit/s λ-tunable time and wavelength division multiplexing passive optical network (TWDM-PON) that utilizes our newly developed high-speed λ-tunable optical network unit transceivers and automatic gain controlled semiconductor optical amplifier-based PON extender for repeater and central office use. The upstream signals for C-band and downstream signals for L+-band in our developed λ-tunable TWDM-PON comply with the wavelength plan of ITU-T G.989 series. In a field trial, we successfully demonstrate error free λ-tunable transmission for the 40-km reach and 1024-split (maximum number) symmetric-rate 40-Gbit/s λ-tunable TWDM-PON over installed standard single-mode fiber links, and advanced network functions such as dynamic load balancing, incremental system upgrade, optical subscriber unit protection, and OLT equipment power saving are also confirmed.