Considerations for incorporating real-time PCR assays into routine marine biosecurity surveillance programmes: a case study targeting the Mediterranean fanworm () and club tunicate () .

Molecular techniques may provide effective tools to enhance marine biosecurity surveillance. Prior to routine implementation, evidence-based consideration of their benefits and limitations is needed. In this study, we assessed the efficiency and practicality of visual diver surveys and real-time PCR assays (targeting DNA and RNA) for detecting two marine invasive species whose infestation levels varied between species and location: and . Filtered water samples ( = 171) were collected in parallel with dive surveys at two locations as part of the New Zealand Marine High Risk Site Surveillance programme: Nelson Harbour (27 sites) and Waitemata Harbour (30 sites). Diver surveys resulted in a greater number of detections compared to real-time PCR: - 21 versus 5 sites in Nelson, 6 versus 1 in Auckland; - 18 versus 10 in Auckland, no detections in Nelson. Occupancy modelling derived detection probabilities for the real-time PCR for were low (14%), compared to (66%). This could be related to abundances, or species-specific differences in DNA shedding. Only one RNA sample was positive, suggesting that most detections were from extracellular DNA or non-viable fragments. While molecular methods cannot yet replace visual observations, this study shows they provide useful complementary information.