Biofilm　in　secondary　water　supply　system　(SWSS)　may　reduce　the　biological　safety　of　tap　water.　This　study　focused　on　the　collaborative　effect　of　secondary　chlorination　and　organic　matter　content　on　the　regrowth　of　biofilm　bacteria　and　the　diversity　of　microbial　community　in　lab　scale　SWSS.　Several　biofilms　cultivated　in　the　same　condition　were　used　for　secondary　chlorination　under　different　organic　matter　content　and　chlorination　dose　conditions.　Bacteria　regrowth　yield　under　the　highest　organic　matter　content　and　the　lowest　free　residual　chlorine　(FRC)　condition　was　2.84-3.11　times　of　that　under　the　lowest　organic　matter　content　and　the　highest　FRC　condition.　The　collaborative　effect　ratio　of　organic　matter　content　increase　(per　1.00　mg/L)　and　secondary　chlorination　decrease　(per　0.10　mg/L　as　Cl-2)　was　0.99.　Metagenomic　sequencing　was　used　to　analyze　the　biofilm　microbial　community　diversity　in　this　experiment.　Proteobacteria,　Bacteroidetes,　Actinobacteria　and　Firmicutes　were　the　four　major　phyla　of　biofilm　microbial　communities.　Proteobacteria　presented　significant　increase　after　secondary　chlorination　in　both　high　and　low　organic　matter　contents.　The　phylum　Bacteroidetes　was　found　to　be　dominant　in　beakers　with　high　organic　matter　content　while　Actinobacteria　was　the　most　in　beakers　with　low　organic　matter　content.　Firmicutes　could　almost　be　controlled　by　secondary　chlorination　with　the　dose　more　than　0.10　mg/L　as　Cl-2.