Summary
We reviewed the literature in an attempt to determine the importance of aseptic technique when implanting electronic tags in fish. Given that there was negligible information on this topic we embarked on a study where bluegill (Lepomis macrochirus) were used as a model to investigate the effects of different aseptic surgical techniques for the intracoelomic implantation of electronic tags in fish. First we tested the effects of water entry into the incision using five treatments: lake, distilled and saline water introduced into the incision, water-free controls, and non-surgery controls. For fish in the water treatments, 1 mL of the sample was introduced into their coelom prior to incision closure. Fish were held for 10 days to monitor survival and at the end of the study, the survivors were blood sampled and euthanized to evaluate condition and health using the health assessment index. In a second experiment, four aseptic treatments were used: non-sterile, field-based, high-grade sterility, and non-surgery controls and fish were monitored as in the first experiment. For both experiments, no differences in physiological status, health or mortality were noted among treatment groups. However, in the aseptic techniques experiment, surgical times were approximately twice as long for fish in the sterile treatment as compared to other groups and the costs of surgical supplies was greater than that of the less-sterile treatments. Although we failed to document any benefit of keeping water out of the incision or using aseptic technique for bluegill, in other situations and for other species, such approaches may be important. As such, we encourage fish surgeons practicing intracoelomic implants to attempt to prevent water entry into the coelom. We also encourage, at least some level of infection control (e.g., non-sterile gloves, clean tags and surgical tools) consistent with good veterinary practices to maintain the welfare status of tagged fish and ensure that the data from tagged fish representative of untagged conspecifics. However, the most prudent and ethical approach would be to work with veterinarians to incorporate formal sterilization procedures, equipment (e.g., sterile gloves) and aseptic technique into field surgical techniques.
Methodology
Study site
All experiments took place at the Queen’s University Biological Station on Lake Opinicon in eastern Ontario, Canada. Bluegill were collected from Lake Opinicon in June 2009 by rod and reel using barbless hooks. We retained all bluegill that were shallowly hooked and were at least 160 mm L T. Fish were temporarily held in 50 L coolers supplied with fresh water during transport by boat to the research station. Fish were then placed into 100 L flow-through outdoor holding tanks and fasted for 24 h to ensure that fish were in a post-absorptive state prior to surgery. Following surgery, the fish were held for 10 days and fed a maintenance ration of fish pellets mixed with blood worms twice daily. The tanks were monitored regularly for the duration of the 10 day holding period. When a fish was observed to exhibit loss of equilibrium and reactivity during the holding period the fish was removed from the tank. After 10 days, a blood sample was obtained (see below) prior to euthanizing the fish via cerebral percussion to enable a post-mortem health assessment (see below). Two different experiments were performed, using new fish for each.
Experiment 1: Entry of water into incision
This experiment included five test groups (N = 15 per group); a control group that was anesthetised but not surgically implanted, a “water-free” group that received an implant but no water injection, as well as lake water (collected from Lake Opinicon), distilled, and saline (physiological saline) treatment groups that received an implant and were injected with 1 mL of water through the incision. The water-free test group surgery used a surgical drape to aid in keeping water out of the incision site.
Experiment 2: Variation in surgical sterility
The general materials and methods for the second experiment were similar to the first experiment except that water was not introduced into the incision. However, the degree of sterility was manipulated. All surgeries were performed on 27 June 2009 and involved four different test groups (N = 15 per group). The first test group had the highest grade of sterility (herein called “sterile”). The foam surgical board was soaked with gluteraldehyde for a 24 h prior surgery and rinsed with distilled water. In between surgeries the foam board was cleaned with a betadine solution. A fresh pair of sterile gloves were used for each successive surgery and placed on the hands of the surgeon by an assistant so that they were not contaminated. The dummy tags were sterilized 24 h prior to surgery using gluteraldehyde, and the surgical tools were sterilized via autoclave for at least 1 h. A fresh (sterile) suture package was used for each fish and no contact between the fish and the surgeon was allowed at all. All surgical tools remained in autoclave packets until used for surgery; the assistant would peel back the packet, and the surgeon would grasp the tool to maintain sterility. A surgical drape (new drape for each fish) with a small window over the incision site was present during surgery to aid in preventing water from entering the incision site. The second test group received aseptic techniques reminiscent to those used in the field (herein called “field”). Non-sterile latex gloves were used and rinsed with betadine between surgeries. The dummy tags, surgical tools, and sutures were cleaned in betadine and then rinsed with distilled water before surgical use. In this treatment, contact between the surgeon and the fish was allowed as well as contact between the suture and the fish. The same sutures were reused on multiple fish after cleaning with betadine. The third test group was subjected to the unsterile conditions (herein called “non-sterile”). No gloves were used and the dummy tags were not sterilized. The surgical tools were left unsterilized from previous treatments surgeries and reused for each consecutive surgery. Fleshy pieces and mucus left from the previous subjects was removed from the surgical tools with a lake water rinse. Leftover sutures from previous fish were not sterilized and reused multiple times. The final test group acted as a control and was anaesthetized but underwent no surgery. The duration of the surgery time was measured to the nearest second. Unlike experiment 1, fish were held in three separate smaller tanks (150 L each). Five fish from each treatment were introduced into each tank. Fish were held for 10 days and sampled as above.