Rapid strand replacement primer thermostat visual sensor based on Bst DNA polymerase and pyrophosphatase for detecting Vibrio parahaemolyticus

•A strand replacement primer thermostat phosphate (SRPP) sensor is designed.•SRP facilitates chain substitution and opens a hairpin with it added at its 3′ end.•The amplification is within 35 min in the manner of thermostat amplification.•Phosphate from the by-product PPi triggers phosphomolybdate blue visualization.•A new detection way of V. parahaemolyticus without extra equipment is proposed. Vibrio parahaemolyticus is a major hidden danger of food safety. To develop a rapid, sensitive and on-site detecting method of Vibrio parahaemolyticus (V. parahaemolyticus), a strand replacement primer thermostat phosphate (SRPP) visual sensor was proposed, based on Bst DNA polymerase and pyrophosphatase. The novel strand replacement primer (SRP) facilitates chain substitution and to open a self-folding hairpin by adding region at its 3′ end. Under the action of the SRP, a pair of external primers and two inner primers, target DNA is specifically amplified at 63 °C relies mainly on the hairpin. Many pyrophosphates (PPi) are simultaneously generated as by-products, which can be converted into phosphates (Pi) by pyrophosphatase for phosphomolybdate blue visual detection within 5 min. The proposed biosensor can detect 1.29 × 103 copies of V. parahaemolyticus within 35 min.