7018

• UCREFRP Catalog Number: 7018
• Author: Muth, R. T. and T. P. Nesler
• Title: Marking Colorado Squawfish Embryos and Newly Hatched Larvae With Tetracycline
• USFW Year: 1989
• USFW - Doc Type: The Southwestern Naturalist
• Copyright Material: YES

September 1989 Notes 433
Tna1.E 1-Relative intensity of fluorescent mark in otoliths from Colorado squawfish larvae that
were immersed (whole body) for different lengths of time (4 to 36 h) in tetracycline hydrochloride
(TC) solutions (200 and 350 mg/1) as embryos or newly hatched larvae. Mark intensity was determined
for five larvae from each treatment and sampling time (days after hatching) and ranked 0 to 3 (absent
to bright). Values were averaged within each sample lot to produce a "batch" mark-intensity value.
Embryos Newly hatched larvae
200 mg TC/1 350 mg TC/I 200 mg TC/I 350 mg TC/l
Sampling
time 12h 24h 36h 12h 24h 36h 4h 12h 4h 12h
0 2.4 2.8 3.0 2.4 2.8 2.8 1.6 2.4 2.4 2.8
7 2.4 3.0 2.8 2.4 3.0 3.0 1.4 2.8 2.4 2.8
15 2.2 2.8 2.8 2.4 3.0 2.8 1.4 2.8 2.4 2.8
Fertilized Colorado squawfish eggs (1-day-old, postfertilization) were obtained from Dexter (New
Mexico) National Fish Hatchery in June 1987. Upon receipt, eggs were visually examined, "dead"
eggs were discazded, and "living" eggs were immersed in a 0.2% solution of formalin (saturated
formaldehyde solution) for about 20 min to treat against "fungal" infection (T. Winham, pers. comm.).
After Formalin treatment, eggs were rinsed with fresh well water, placed in glass trays covered with
fine-mesh screening, and incubated in a flow-through trough which received well water at 20°C
(Hammon, 1986). Water in the trough was aerated.
During the initial days of incubation, egg samples were taken twice daily and fixed and preserved
in 95% ethanol (Brothers, 1987; Muth et al., 1988). Soon after preservation, embryos were examined
under a dissecting microscope fitted with a polarizing filter (Brothers, 1987) for presence of otoliths.
Sagittae and lapilli (otoliths) were first visible in 3-day-old (postfertilization) embryos.
Embryo marking was started after otoliths were present in the embryos. About 50% of all living
eggs were divided into seven experimental groups (six treatments and one control), each with 185
specimens. The control group was placed in an incubation tray in the flow-through trough. Treatment
groups were placed in 1-1 glass beakers containing tris-buffered, near-neutral pH (Hetzler, 1984; Muth
et al., 1988) solutions of 200 or 350 mg TC/1 of distilled water for 12, 24, or 36 h. During TC
exposure, treatment groups were aerated and incubated in a covered water bath at 20°C.
Following TC exposure, treatment groups were placed in incubation trays in the flow-through
trough. After hatching was completed (4 to 5 days after fertilization; 12 to 24 h after treatment),
numbers of unhatched (dead) eggs were recorded for all experimental groups. Larvae were placed in
aerated well water in 3.8-1 rearing jars and incubated in a water bath at 20°C. Larvae were reared
for about 15 days after hatching. Rearing jars were cleaned and numbers of dead larvae were recorded
twice daily. After mouth and gut development were complete (about 6 days after hatching), larvae
were fed twice daily with live Artemis sp. nauplii and TetraMin Fry Diet.
Five living larvae from each experimental group were sampled immediately after hatching and at
weekly intervals thereafter. Sampled larvae were measured (millimeters total length, TL), examined
for any external abnormalities, and fixed and preserved in 95% ethanol. Using procedures described
by Muth et al. (1988) and Brothers (1987), Sagittae and lapilli were extracted from the larvae, mounted
in glycerin on glass slides, and examined with incident UV light under a compound microscope for
presence and intensity of the fluorescent mark. Mark intensity was determined for each larva examined
and ranked. Ranking categories used were absent, Faint, lucid, or bright with respective values of 0, 1,
2, or 3. Values were averaged within each sample lot of flue fish, and the mean was used to compare
mark intensity among sample lots.
For marking newly hatched larvae, eggs not used in the embryo-marking experiment were allowed
to hatch. Immediately after hatching was completed (5 days after fertilization), larvae were divided
into five experimental groups (four treatments and one control), each with 250 specimens. Control
larvae were placed directly in a rearing jar (previously described). Treatment groups were placed in
1-1 glass beakers rnntaining tris-buffered, near-neutral pH solutions of 200 or 350 mg TC/1 of distilled
water for 4 or 12 h. During TC exposure, treatment groups were aerated and incubated in a covered