1. Introduction
It is important that fish be individually identified because performance varies Ž
substantially among individuals of different or the same size e.g., Jobling et al. 1989; .
Alanara and Brannas 1993; McCarthy et al. 1994 . This should ideally be done from the
¨ ¨ ¨ ¨
youngest possible age and extend over long periods of time. Passive integrated transpon- Ž
. ders PITs; Prentice et al. 1984, 1990a are among the best candidates for tagging small
Ž .
fish. These are low weight F 100 mg tags, with unlimited functional life and billions of individual codes, and can thus be applied to large samples of fish. The use of data
entry stations allows the automatic detection of PIT tagged fish passage or presence at Ž
. Ž
close range within 20 cm of detecting antennas Prentice et al. 1990b,c; Brannas and
¨ ¨
. Alanara 1993; Brannas et al. 1994; Armstrong et al. 1996 .
¨ ¨ ¨ ¨
Ž PIT tags have been applied mainly to large fish in husbandry management e.g.,
. Jenkins and Smith, 1990 . The tag is inserted into the dorsal musculature of broodfish,
represents a tiny proportion of their body weight, and has no marked effect on their physiology. Few studies have documented the feasibility of implanting PIT tags into the
Ž body cavity of small juveniles, and most of these concerned salmonids Prentice et al.,
. 1990a; Brannas and Alanara, 1993; Peterson et al., 1994 . Behavioural and physiological
¨ ¨ ¨ ¨
reactions to identical tagging procedures may differ substantially between species Ž
. Winter, 1983; Summerfelt and Smith, 1990; Baras et al., in press , and feasibility
studies on tag acceptance are strongly encouraged when no detailed data are available on the species of interest, both for ethical considerations and validation of results. This
Ž .
becomes crucial when using high tag ratios tag weightrfish weight , close to or above Ž
the mean adjustment capacity of the swimbladder in teleost fish ca. 1.75, Alexander, .
Ž 1966 , as most tagging biases were reported in these circumstances Greenstreet and
. Morgan, 1989; Summerfelt and Smith, 1990; Baras et al., in press . More recent studies
with carefully tailored procedures suggest that higher tag ratios can be used without Ž
causing substantial biases in the long run Claireaux and Lefranc
¸
ois, 1998; Martinelli et .
al., 1998; Baras et al., 1999 . This paper is a component of research programmes on the biology of cultured
juveniles of the Eurasian perch Perca fluÕiatilis, which rely on monitoring the activity of PIT tagged individuals through the use of automatic data entry stations. To evaluate
the adequacy of the tagging technique and to determine the minimum size at which perch can be tagged successfully, the intensity and duration of the perturbation caused
by the tagging procedure and tag presence were measured based on tag retention, survival, growth, healing progress and physiology of juvenile perch of different body
Ž .
weights 1.67–10.62 g, 55–96 mm fork length . As the detection of PIT tags is
governed by the laws of inductive coupling, the orientations of implanted tags and their stability over time were also examined to test whether all tagged fish would have the
same chances of being detected by automatic stations.
2. Material and methods
2.1. Fish origin and experimental infrastructure The fish used in the experiment were hatchery reared age-0 perch obtained from
natural spawning by captive breeders in the rearing facilities of the Tihange Aquaculture
Ž .
Station of the University of Liege Belgium . Larvae and young juveniles were reared
`
Ž .
2
Ž
3
. over 6 weeks at constant temperature 22 18C in 10 m
4 m outdoor flow through
Ž tanks, then were transferred into an indoor recirculating circuit
250-l aquarium, .
12L:12D . Water temperature was maintained at 22 18C and monitored by a data Ž
. logger TidBit, ONSET Comp. . Oxygen concentration in the aquarium was measured
once a day and was never - 5.5 mg l
y1
. Water was supplied from well water, and the infrastructure was deemed to be pathogen-free, except when fish from outdoors facilities
were transferred for establishing experimental groups. The first experiment was con- ducted in a non-pathogen-free environment to test for the effect of pathogens on the
survival of tagged fish, and whether survival and retention were affected by the tagging protocol.
2.2. Comparison between tagging protocols Experiment 1 Ž
Three protocols were assayed on groups of 30 fish each 2.78–8.54 g, 65–90 mm .
Ž fork length , using alcohol sterilised TROVAN transponders model ID100, 11 = 2.2
. mm in diameter, 100 mg in the air, 58 mg in the water . In all three protocols, juvenile
Ž
y1
. perch were anaesthetised with tricaine methanesulfonate 90 mg l
and placed ventral side up in a support made of wet paper. In the first group, the tag was injected into the
body cavity, using a standard 12-gauge needle injector passed through the midventral body wall. In the second and third groups, the tag was inserted through a 3-mm long
incision made with a scalpel on the linea alba, ca. 2–3 mm anterior to the papilla. In the second group, the incision was left open, as it normally is, following the injection of the
tag with a syringe. In the third group, it was closed with a single stitch of polyamide
Ž .
monofilament suture material 0.7 Dec fixed to a 12-mm curved cutting needle. On average, the surgical implantation was completed within 60 s, and suturing required
further 60–80 s. The syringe injection procedure never took more than 30 s. The 90 tagged fish were reared over 4 weeks in the same aquarium, together with 30
Ž .
control handled but not tagged fish of similar body weight, so control and tagged fish Ž
were exposed to the same pathogens. Formulated feeds 50 protein, 11 lipid
. contents were distributed during daytime by an automatic feeder, starting ca. 1 h after
Ž .
illumination. The daily food ration DFR, BW was close to the optimum ration at
m y0 .24
Ž . Ž
22–238C, and was 3.30 W , where W
is the mean body weight g of fish Melard
´
m m
. et al., 1996 . The aquarium was searched twice a day for dead fish and lost tags. Fish
were examined once a week to measure individual growth rates; specific growth rates Ž
.
y1
w xŽ
.
y1
SGR , d s 100 = ln W y ln W
t y t , where W and W
were the body
2 1
2 2
2 1
weights at times t and t , respectively. Upon the first examination and weighing, the
2 1
stitch was removed to avoid any detrimental long term effect inherent to the presence of Ž
a transcutaneous foreign body Knights and Lasee, 1996; Thoreau and Baras, 1997; .
Baras et al., in press . Ž
. Prentice et al. 1990a provided evidence for a good correspondence between visual
observation and histological examination when evaluating the healing progress in PIT tagged salmonids. The incision was considered as completely healed when the external
layers of the body wall had closed up, and the perch was regarded as having recovered
its physical integrity when the incision was covered by regenerated scales. The mean healing time for each treatment was deduced from the distribution frequency of healed
fish at each inspection.
2.3. Determination of minimum fish size for pit tagging Experiment 2 Approximately 2000 juvenile perch were acclimatized for 10 days to the experimental
infrastructure and no pathology was observed prior to tagging. Two hundred and twelve Ž
. fish 1.67–10.62 g, 55–96 mm fork length were tagged with transponders, using
Ž .
surgical implantation and suturing see justification in Table 1 . No fish smaller than 55 Ž
. mm was tagged because the length of its body cavity excluding the pericardial cavity
was similar to the length of the tag. Analyses were made on eight weight classes, Ž
corresponding to mean tag ratios in the water from 0.75 to 2.62 corresponding .
ratios in the air from 1.29 to 4.52; Table 2 .
Table 1 Comparison of SGR, survival, tag expulsion, and healing progress of PIT tagged age-0 perch P. fluÕiatilis,
Ž
y1
. depending on the tagging protocol 30 fish in each group; 250-l aquarium; G 5.5 mg l
O ; 22–238C
2
Ž .
For each line of the table, the categories sharing at least one common superscript a, b, c are not significantly Ž
different, whereas the other comparisons differ at P - 0.05 Fischer PLSD comparison of means for growth .
Ž .
and body weight, contingency tables for tag expulsion, mortality and healing progress . ‘‘ 3D 5’’ in the expelled tags lines means that tags were expelled by five fish, of which three eventually died.
Syringe Surgery,
Surgery, Control
injector no suture
suture Body weight BW, mean SD, g
a a
a a
d 0 4.780.97
5.061.37 5.011.08
4.881.47
a a
a a
d 42 7.301.78
7.582.63 7.051.98
7.882.77
y 1
SGR mean SD, BW d
ab ab
a b
dd 1–7 1.600.85
1.581.09 1.301.34
2.001.39
a a
a a
dd 8–14 1.380.72
1.520.87 1.660.79
1.670.72
a a
a a
dd 15–21 1.660.60
1.900.76 1.680.56
1.920.49
a a
a a
dd 22–28 1.200.66
1.130.61 1.270.46
1.390.66 Dead fish running sum
a ab
a b
d 7 8
3 7
a ab
ab b
d 14 12
6 7
3
a ab
ab b
d 21 12
7 7
3
a ab
ab b
d 28 12
7 7
3
a ab
b a
Initial body weight of dead fish 4.430.80
4.950.79 5.621.00
3.561.73 Ž
. meanSD, g
Expelled tags running sum
a a
b
Ž .
d 7 3D 5
4 –
a a
b
Ž .
d 14 6D 7
6 –
a a
b
d 21 7
6 –
a a
b
d 28 7
6 –
a a
b
Ž .
Healing time mean, d 20.1
18.0 11.8
–
E. Baras
et al.
r Aquaculture
185 2000
159 –
173
163 Table 2
Comparisons between SGR, survival and healing progress of juvenile P. fluÕiatilis tagged with surgically implanted PIT tags, depending on the initial body weight Ž
y1
. 250-l aquarium; G 5.5 mg l
O ; 22–238C except from days 22 to 32: 20–218C
2
Tag retention was 100. Tag ratio stands for tag weightrfish weight. Growth rates were independent of tag ratio on any of the four controls between days 43 and 126, and are not illustrated separately.
Ž .
For each line of the table, the categories sharing at least one common superscript a, b, c are not significantly different, whereas the other comparisons differ at Ž
. P - 0.05 Fischer PLSD comparison of means for growth and body weight, contingency tables for mortality and healing progress .
Ž . Weight class g
- 2.5
2.5–3.0 3.0–3.5
3.5–4.0 4.0–4.5
4.5–5.5 5.5–6.5
G6.5 Mean tag ratio BW
In air 4.52
3.65 3.09
2.69 2.35
1.97 1.66
1.29 In water
2.62 2.12
1.80 1.56
1.37 1.15
0.97 0.75
Initial numbers 34
29 34
31 27
21 17
19 Dead fish running sum
a a
a a
a a
a a
d 42 1
1 1
1 1
1
b a
a a
a a
a a
d 126 11
2 3
4 3
1 1
1 Sex ratio F:M
d 0 22:12
12:17 17:17
12:19 14:13
12:9 10:7
12:7 d 126
18:5 11:16
15:16 10:17
13:11 12:8
10:6 11:7
Body weight BW mean SD, g
d 0 2.210.24
2.740.16 3.240.12
3.720.17 4.260.14
5.080.26 6.030.23
7.731.03 d 42
3.950.65 5.071.13
5.761.22 6.220.99
7.080.77 8.471.35
9.741.60 11.832.83
d 126 13.534.07
17.226.42 19.435.65
21.597.25 25.898.08
27.607.26 34.3911.41
44.4014.20
y 1
SGR BW d mean SD
a a
a a
a b
b b
dd 1–11 1.001.07
1.170.66 1.100.76
1.120.86 1.000.67
1.620.87 1.720.64
1.580.46
a a
a ab
b bc
c c
dd 11–21 1.950.80
1.820.68 1.810.51
1.580.67 1.420.53
1.310.44 0.980.62
1.060.62
a a
a a
a a
a a
dd 22–32 0.910.60
1.040.45 1.000.54
0.970.44 1.060.31
0.910.35 1.050.38
1.150.35
a a
a a
a a
a a
dd 33–42 2.361.95
1.991.58 1.880.77
1.690.75 1.880.37
1.700.48 1.640.51
1.930.39
a a
a a
a a
a a
dd 43–126 1.480.21
1.500.29 1.500.32
1.510.37 1.600.32
1.490.55 1.550.38
1.660.14
b a
a a
a a
a a
Healing time mean, d 15.8
13.1 13.2
13.6 12.4
11.6 11.9
11.6
Ž Perch were reared over 126 days in groups initially three groups of 344 fish each,
rearranged on day 43 into nine groups of 114–115 each, containing fish of all weight classes, at a ratio of ca. one tagged to three untagged fish. Food distribution, search for
dead fish or expelled tags, assessment of healing progress and growth were as above, Ž
except for the periodicity of the assessments twice every 3 weeks until day 42, then .
once every 2 weeks . The remote detection of transponders relies on inductive coupling, and is dependent
on the orientations of the detecting loop and tag coil. Maximum coupling is obtained when the tag crosses perpendicular to the plane of the loop, and other angles cause a
reduction in detection range, which is inversely proportional to the cosine of this angle with the ideal trajectory. Tags that are misplaced at the time of tagging or become
disorientated during fish growth, may compromise coupling, reduce the detection range
Ž w
x. and the probability that the fish be detected i.e., reduction s 1 y cos angular deviation .
X-ray photographs of all tagged fish were made under anaesthesia on days 73, 94 and Ž
. 115 using an X-ray TR 80r20 Todd Research, UK and films Kodak X-OMAT MA,
developed with Fuji Film FPM 100 A. Tilt angles to the sagittal axis of the body were measured to the nearest degree, assuming that this axis was the ideal trajectory for a fish
crossing perpendicular to the antenna. Associated risks of detection failure were calculated as above. Photographs were used to determine the positions and migrations of
the tags in the body cavity, using five classes of horizontal and vertical coordinates.
At the end of the experiment, all fish were dissected, and examined visually for gross Ž
. morphological effects from surgery e.g., muscle necrosis, erythema and host tissue
Ž .
response to the tag e.g., embedded in adipose tissue, encapsulation . In all tagged fish and in a sample of 50 control fish, the abdominal adipose tissue was extracted with
Ž .
forceps and weighed. The gonado-somatic index GSI was measured and the sex of all Ž
fish was determined from visual examination of gonads single ovary in female perch .
and two testes in male perch; Mallison et al., 1986; Craig, 1987 . The aceto-carmin Ž
. squash technique Guerrero and Shelton, 1974 was also used on randomly sampled fish
to validate the visual assessment of their sex.
2.4. Statistical analyses Contingency analyses were used to compare fish survival, tag retention rates and
healing progress in both experiments, and to test for the effect of initial body weight at tagging on host tissue reaction and tag positioning in Experiment 2. Analyses of
Ž .
variance one-way ANOVA and Fischer PLSD comparisons of means were used to compare the orientations of tags, the body weights and growth rates of fish. The growth
rates of controls were calculated from the ordinated distributions of their body weights at the beginning and end of each rearing period. This calculation relied on the
assumption of the primacy of early size differences, which is not an invalid assumption
Ž .
for short term experiments see Baras, 1999 . Factorial ANOVA was used to test for differences between the GSI and proportion of perivisceral fat, depending on fish sex
and weight class. Student’s t-test and simple regressions tests were also used when appropriate. Null hypotheses were rejected at P - 0.05.
3. Results
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