An H6N1 virus, A/teal/Hong Kong/W312/97 (W312), was isolated during the ´bird flu´ incident of Hong Kong in 1997. Genetic analysis suggested that this virus might be the progenitor of the A/Hong Kong/156/97 (HK/97) H5N1 virus as seven of eight gene segments of those viruses had a common source. Continuing surveillance in Hong Kong showed that a W312-like virus was prevalent in quail and pheasant in 1999; however, the further development of H6N1 viruses has not been investigated since 2001. Here we report influenza surveillance data from 2000 to 2005 in southern China that shows H6N1 viruses have become established and endemic in minor poultry, and mainly replicated in the respiratory tract. Phylogenetic analysis indicates that all H6N1 isolates had W312-like hemagglutinin and neuraminidase genes. However, reassortment of internal genes between different subtype virus lineages, including H5N1, H9N2 and other avian viruses, generated multiple novel H6N1 genotypes in different types of poultry. These novel H6N1/N2 viruses are double, triple or even quadruple reassortants. Reassortment between a W312-like H6N1 virus and an A/quail/Hong Kong/G1/97-like H9N2 virus simultaneously generated novel H6N2 subtype viruses that were persistent in poultry. Molecular analyses suggest that W312-like viruses may not be a precursor of HK/97 virus but a reassortant from HK/97-like virus and another unidentified H6 subtype virus. These results provide further evidence of the pivotal role of the live-poultry market system of southern China, in generating increased genetic diversity of influenza viruses in this region.