Objective:　To　provide　a　new　idea　for　the　development　of　Bacillus　subtilis　bio-preservative　based　on　biofilm　forming　ability.　Methods:　The　hydrophobic　ability,　self-agglutination　rate,　co-agglutination　rate　and　biofilm　forming　ability　of　Bacillus　subtilis　were　measured.　The　freshness　and　volatile　flavor　of　tilapia　fillets　were　measured　by　Bacillus　subtilis.　Results:　The　stronger　the　biofilm　forming　ability　of　Bacillus　subtilis,　the　best　preservation　effect　of　tilapia　fish.　Bacillus　subtilis　can　significantly　reduce　the　content　of　volatile　basic　nitrogen　in　fish　meat,　so　that　the　TVB-N　content　of　fish　was　24.43　mg/100　g　on　the　third　day　of　storage,　which　was　lower　than　the　control　group　(34.58　mg/100　g).　Moreover,　the　production　of　alcohol,　phenols,　esters　and　acids,　such　as　isoamyl　alcohol,　phenol,　indole,　acetic　acid　and　other　volatile　flavor　components　in　fish　was　significantly　inhibited.　At　the　same　time,　compared　with　the　biofilm　forming　ability,　the　bacteriostatic　ability　of　Bacillus　subtilis　was　not　correlated　with　the　fresh-keeping　ability.　The　results　showed　that　the　autoagglutination　and　biofilm　forming　ability　of　Bacillus　subtilis　were　most　correlated　with　the　preservation　ability　of　tilapia　fillets,　and　could　be　used　as　a　rapid　index　for　screening　Bacillus　subtilis　for　preservation.　©　2023　Chinese　Institute　of　Food　Science　and　Technology.　All　rights　reserved.