Electronic individual identification of zebrafish using radio frequency identification (RFID) microtags

Abstract

SUMMARYAlthough individual electronic tagging using passive integrated acoustic (PIT) tags is established, it is mainly for fish >60mm in length and is unsuitable for fish of <30mm, like zebrafish. We used radio frequency identification (RFID) microtags (1mm in diameter and 6mm in length, with a mass of ~10mg) to individually identify juvenile zebrafish (length 16-42mm, mass 138-776mg) for the first time, and studied the effects of intracoelomic implantation on fish survival and microtag loss, growth, spawning and exploratory behaviour. After 5.5months, both high survival (82%) and low microtag loss (11%) were achieved. The smallest surviving fish weighed 178mg, and success in microtag reading was 73% for the size class 350-450mg (26mm). Greater success was achieved when fish were larger at the time of tagging but no negative effects on growth were observed for any size class and some tagged fish spawned. No significant differences in behavioural responses could be detected between tagged fish and untagged controls after 2months. Overall, the results suggest that the tagging method is highly suitable for fish as small as zebrafish juveniles. We think this method will provide significant advances for researchers of the ever-growing fish model community and more generally for all small-fish users. Tagging is essential when one needs to identify fish (e.g. particular genotypes with no external cue), to run longitudinal monitoring of individual biological traits (e.g. growth) or to repeat assays with the same individual at discrete points in time (e.g. behaviour studies). Such a method will find applications in physiology, genetics, behaviour and (eco)toxicology fields. Supplementary material available online at

Evaluation of visible implant elastomer tags in zebrafish (Danio rerio)

“…The injection force of skin tattooing, a less invasive marking method, killed channel catfish fry the size of zebrafish (unpublished data). Recently, RFID microtags have been evaluated in zebrafish (Cousin et al, 2012). These tags although much smaller than regular PIT tags are rather invasive and costly when compared to the VIE tagging method.…”

“…Multiple colors are available, and they are easily discerned by the unaided eye. Visible implant elastomer (VIE) tags have been successfully used for many aquatic species (Northwest Marine Technology, Inc., 2008, ‘Visible implant elastomer tag project manual’, http://www.nmt.us/support/appnotes/ape06.pdf) (Blackburn et al, 2011; Cousin et al, 2012) but the use in zebrafish to date is sparse. VIE tags need to be validated for each species under study (Astorga et al, 2005) and in this study we evaluated batch tagging of wild-type and rag1 −/− zebrafish.…”

Individual Identification of Goldfish from Eye Morphology: The Eye Mark Method

“…Internal tagging using PIT tags and RFID tags has also recently been developed and can be used to identify large numbers of individuals (Sakalski et al, 1998;Sato et al, 2006;Cousin et al, 2012). External and internal tagging are invasive methods, and effects on the behavior and physiology of the fish are significant, especially when the subjects are small (Carline and Brynildson, 1972;Heugel et al, 1977;Moodie and Salfert, 1982;Columbia Basin Fish and Wildlife Authority PIT Tag Steering Committee, 1999).…”

“…However, tags may be lost, scars may fade, and scarred areas may regenerate (Welch and Mills, 1981). Recently, implantation of passive integrated transponder (PIT) tags or radiofrequency identification (RFID) microtags has been used to identify large numbers of individual fish (Skalski et al, 1998;Sato et al, 2006;Cousin et al, 2012). However, all external and internal tagging methods require invasive surgical procedures, and may cause significant stress and have physiological effects, especially in small individual fish (Carline and Brynildson, 1972;Heugel et al, 1977;Moodie and Salfert, 1982;Columbia Basin Fish and Wildlife Authority PIT Tag Steering Committee, 1999).…”

A method for the automated long‐term monitoring of three‐spined stickleback Gasterosteus aculeatus shoal dynamics

“…Furthermore, the possibility of automatic monitoring using these markers is often not possible. Remote tagging using acoustic, radio or passive inductive transponder (PIT) tags has also been used to monitor the identity and location of individual fishes (Cousin et al , ), although these techniques are invariably invasive and a high spatial and temporal resolution is not always guaranteed. Therefore, reliably monitoring multiple fishes over an extended period of time is still a great challenge.…”