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Spawning and Captive Propagation of Blackside Dace,
Chrosomus cumberlandensis
Patrick L. Rakes1,*, Melissa A. Petty1, J.R. Shute1, Crystal L. Ruble1,
and Hayden T. Mattingly2
Abstract - Chrosomus cumberlandensis (Blackside Dace) is a federally protected stream
fish endemic to the upper Cumberland River drainage of Kentucky and Tennessee. Captive
propagation of the species has been conducted in only one previous study. Here
we report new findings from spawning and rearing the species for the past three years,
2011–2013, at the Conservation Fisheries, Inc. (CFI) facility in Knoxville, TN. Brood
stock (n = 80 adults) were collected in 2011–2012 from Big Lick Branch in Pulaski
County, KY. Spawning at the CFI facility occurred in April and May of each year when
CFI water temperatures were between 16 and 21 °C. We compare CFI spawning dates
and temperatures to those measured in a 2006 field study. We also provide video footage
of spawning behavior, which conformed to that previously described in the literature.
Per capita production of fry (i.e., number of juveniles reared per breeding adult) in 2012
was more than twice that achieved in the previous study. We progressively reduced the
presence of (and cues from) other fish species in 2011, 2012, and 2013, with the eventual
achievement of having Blackside Dace spawn completely independent from any heterospecific
cues. Our results collectively increase the capacity to propagate Blackside Dace
in captivity to support ongoing recovery ef forts for this threatened species.
Introduction
Captive breeding and reintroduction programs have a number of recognized
limitations, yet they often play vital roles in the conservation of endangered
species. Among freshwater fishes, there are several examples of successful restoration
of species to their native waters (e.g., Mueller and Wydoski 2004, Shute et
al. 2005). However, protocols for captive propagation and/or translocation have
not been developed for many freshwater fishes, thereby restricting conservation
options for those species.
Successful captive propagation of any given fish species is facilitated by
advance knowledge of its spawning habits in a natural setting. However, in the
diverse minnow family, Cyprinidae, spawning modes are known for only 13 of
46 imperiled species (Johnston 1999). Johnston and Page (1992) reviewed the
reproductive strategies of cyprinids and identified eight different categories. Six
of the eight strategies involve preparation or use of substrate to form a nest in
which eggs are deposited and fertilized. Parental care in the form of nest guarding
is provided by males in some instances.
One important aspect of cyprinid reproduction is the nest association behavior
displayed by at least 33 species (Johnston and Page 1992). Nest association
1Conservation Fisheries, Inc., 3424 Division Street, Knoxville, TN 37919. 2Department
of Biology, Box 5063, Tennessee Technological University, Cookeville, TN 38505. *Corresponding
author - xenisma@gmail.com.
Ecology and Conservation of the Threatened Blackside Dace, Chrosomus cumberlandensis
2013 Southeastern Naturalist 12(Special Issue 4):162–170
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occurs when one species, the host, prepares a nest that another species, the
associate, uses for spawning (Johnston and Page 1992). The host species is typically
another cyprinid, but some associates use centrarchid or other nests (e.g.,
Fletcher 1993, Hunter and Hasler 1965, Johnston and Page 1992). Certain cyprinids
show flexibility by spawning either (1) as an associate in the nests of hosts
or (2) independent of the host by broadcasting or by building their own nests
(Johnston and Page 1992). Understanding the degree of host dependence and
host specificity can inform captive breeding protocols and establish criteria for
selecting reintroduction sites to ensure that appropriate hosts are present.
Chrosomus cumberlandensis (Starnes and Starnes 1978) (Blackside Dace) is
a federally protected cyprinid whose known native range is restricted to small
tributaries in the upper Cumberland River system in southeastern Kentucky and
northeastern Tennessee (Black et al. 2013a [this issue]; Eisenhour and Strange
1998; O’Bara 1988, 1990; Starnes and Etnier 1986; Starnes and Starnes 1978,
1981; USFWS 1988). The species inhabits cool (less than 20 °C) headwater streams with
stable substrates, low water conductivity (less than 240 μS), and sufficient in-stream
cover (Baxter 1997, Black et al. 2013b [this issue], O’Bara 199 0).
Blackside Dace reproduction in a natural stream was first reported by Starnes
and Starnes (1981), who observed the species using a broadcasting spawning
mode over substrate in a nest of Campostoma anomalum (Rafinesque) (Central
Stoneroller). These authors observed one spawning event on 17 May 1981 at a
water temperature of 17.5 °C. Cicerello and Laudermilk (1996) later observed
a school of nuptial Blackside Dace over the occupied nest of Semotilus atromaculatus
(Mitchill) (Creek Chub) on 12 May 1993, although actual spawning was not
observed. Mattingly and Black (2013 [this issue]) observed 25 Blackside Dace
spawning events from 12 May to 12 June 2006 at water temperatures of 11.9 to
18.2 °C, all of which occurred over Creek Chub nests. These field observations
confirm that Blackside Dace is a nest-associating species that uses at least two
cyprinid host species. No field reports of Blackside Dace spawning independent
of cyprinid hosts are known, although most authors have presumed that independent
spawning may occur.
Rakes et al. (1999) conducted the first study of Blackside Dace propagation
in captivity. Twenty-four adult Blackside Dace were collected from Buck
Creek (Whitley County, KY) on 7 May 1993. The dace were maintained at 16
°C in Knoxville, TN, at the Conservation Fisheries, Inc. hatchery (hereafter CFI
facility) in two 75-L aquaria lined with gravel and pebble substrate. Mounds of
substrate were arranged in the aquarium to create artificial “chub nests” (20 cm
wide, 30 cm long, 5 cm high), thereby simulating a cyprinid host nest. Flowing
water was directed over the mounds to mimic natural conditions. The fish
spawned upon first arrival at the CFI facility but subsequently ceased reproductive
activity. Rakes et al. (1999) then used milt (see Hunter and Hasler 1965)
from a reproductively mature male Central Stoneroller and Nocomis micropogon
(Cope) (River Chub) to induce additional Blackside Dace spawning activity.
Three hundred and thirty fertile eggs (230 before addition of milt and 100 after
P.L. Rakes, M.A. Petty, J.R. Shute, C.L. Ruble, and H.T. Mattingly
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milt) were siphoned from the mounds and moved to incubation trays. Selected
early life-history information was recorded, including egg diameter (1.0 mm),
egg characteristics (demersal, non-adhesive), egg deposition (among gravel and
pebbles of artificial minnow nests), hatchling size (5 mm total length), characteristics
of embryos and larvae (benthic approximately 48 h), foods used by larvae
(live copepods, brine shrimp nauplii, commercially prepared rotifer-sized powdered
food), and survival of fertile eggs to the juvenile stage (87%).
The goal of the present study is to build on the knowledge provided by Rakes
et al. (1999) to further advance our understanding of Blackside Dace captive
propagation and early life history. We propagated Blackside Dace in captivity
for three consecutive years, 2011–2013, to accomplish this goal. Our specific
objectives were to (1) refine captive propagation techniques to allow greater
production of offspring, (2) determine the duration and survival of early life
stages, (3) compare dates and temperatures of spawning in captivity to field
observations, (4) determine whether Blackside Dace are capable of spawning
independently in captivity without the presence of host fishes, and (5) describe
spawning behavior, including the presentation of video footage of spawning.
Methods
Collection of wild brood stock
We collected a total of 80 adult Blackside Dace (n = 50 on 17 March 2011
and 3 May 2011, plus an additional n = 30 on 7 February 2012) from Big Lick
Branch in Pulaski County, KY. Stream water temperatures were 7 °C, 13 °C,
and 5.5 °C on the three collection dates, respectively. Big Lick Branch harbors
one of the more robust populations known for Blackside Dace (Black
et al. 2013a [this issue]). All fish were collected using a fine mesh seine and
transported to the CFI facility in bags within insulated containers, then slowly
acclimated to laboratory aquaria.
Presence of other fish species in aquaria
Multi-aquaria recirculating systems were utilized to house the brood stock
and rear young in 2011, 2012, and 2013 (see Fig. 1 in Rakes et al. 1999). Each
year, we progressively reduced the presence of cyprinid host fishes and cues from
host fishes as summarized in Table 1. Briefly, in 2011 all 50 Blackside Dace and
two reproductively mature male stonerollers (Campostoma sp.) were housed in
a 380-L spawning aquarium that was part of a larger recirculating system. The
stonerollers and then the Blackside Dace suffered a disease outbreak during early
May 2011. The infection was successfully eliminated with medications, but only
after losing over 50% of the fish. In 2012, brood stock were housed in two 170-L
spawning aquaria that were part of a larger recirculating system; one housed the
remaining older 20 dace collected in 2011 and the other housed the 30 additional
Blackside Dace collected in 2012. Two male Creek Chub were present in another
tank in the aquarium system, sharing the same water. In 2013, all 40 surviving
Blackside Dace were housed in a single, 170-L aquarium with no contact with
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other cyprinids (or even water from other cyprinids) for approximately one year.
Blackside Dace sex ratios were roughly equivalent in all three years, but precise
determination was not made.
Introduction of milt to aquaria
Milt from reproductively mature male stonerollers (in 2011) as well as Creek
Chubs and Moxostoma sp. “Sicklefin Redhorse” (in 2012) was used to induce
spawning in 2011 and 2012. Dates of milt addition are provided in Table 1. Reproductive
behavior and intensity of nuptial coloration instantly increased upon
exposure to all the introductions of milt, and breeding continued for weeks following
this stimulus. However, in 2013 we purposely did not introduce milt or
any other chemical or visual cues.
Underwater physical conditions in aquaria
In a field setting, Blackside Dace have only been observed spawning over
clean gravel nests constructed by Central Stonerollers and Creek Chubs (Cicerello
and Laudermilk 1996, Starnes and Starnes 1981). Further, Mattingly and
Black (2013 [this issue]) observed that Blackside Dace only spawned or exhibited
spawning behaviors in areas where Creek Chubs had constructed a pit and
ridge nest. Therefore, field surveys support the assumption that Blackside Dace
may have a strong relationship with nest-building cyprinids. To simulate the
nests over which Blackside Dace spawn, we provided our aquaria with flowing
water and artificial “chub nests,” with dimensions and details remaining the same
as described above for the Rakes et al. (1999) study .
Photoperiod and temperature
Blackside Dace were maintained and conditioned in preparation for captive
spawning. Photoperiod was controlled with an astronomic timer to mimic
natural winter lighting conditions by slowly changing day length on a schedule
comparable to the actual season. Food quantities were provided dependent upon
water temperature and the accompanying activity levels of the fish. As spring
approached, both water temperatures and day length were increased, closely following
natural conditions (in Knoxville, TN). Feeding was increased as appetites
tracked rising water temperature. High feeding activity and provision of food
(2–3 times/day) began in early March as the fish came into breeding condition
and feeding activity remained at high levels through mid-May ea ch year.
In 2006, we measured water temperatures at 5 sites in 4 streams where Blackside
Dace spawning events were observed by Mattingly and Black (2013 [this issue]).
A single temperature datalogger per site was deployed in April 2006 and retrieved
in December 2006. In 2011–2013, water temperatures at the CFI facility were measured
and recorded by temperature dataloggers (T & D Corporation, Model RTR
500NW and Model RTR 502) and archived in an online storage database. We calculated
daily mean temperature values from the hourly field and CFI-facility data
from March or April through June each year, and used these mean values in qualitative
comparisons between field and hatchery temperature conditions.
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Collection and incubation of embryos
Fertilized and unfertilized eggs, as well as detritus, were removed every 2
to 10 days by vacuuming through the artificial pebble nests with an aquariumcleaning
siphon. The few eggs collected in 2011 appeared to be from one
spawning event. A total of eight egg collections were performed throughout
the spawning period in 2012, with each certainly encompassing multiple
spawning events as evidenced by varied egg developmental stages. Likewise,
we collected eggs from at least four different spawning events in 2013. Eggs
were transferred with a pipette to the incubation trays; water temperature during
incubation was 15–20 °C. Unfertilized eggs were discarded and all counts
reported below are of fertilized eggs (i.e., embryos).
Our production goal in 2013 was different than in 2011 and 2012. In 2013
we simply wanted to produce ≈700 embryos for use in toxicity studies with collaborating
researchers. After reaching our production goal, we stopped collecting
additional eggs and ceased monitoring spawning activities of brood stock. We
also did not track survival of embryos in 2013.
Results and Discussion
Production and survival of offspring
Only the largest, oldest Blackside Dace collected in 2011 successfully
spawned that year, and only the fish collected in 2011spawned in 2012. Though
apparently sexually mature, no eggs or larvae were recovered from the younger
fish collected in 2012. Post-hatch yolk-sac larval production increased from
a low of 71 fry produced from 117 eggs collected in 2011 (60% survivorship)
to a peak of 1910 fry produced from the 2855 eggs collected in 2012 (67%
survivorship). Production in 2013 was 840 eggs, but additional eggs were not
collected and survival was not reported that year.
Our 2011–2012 survival rates (60-67%) were slightly lower than reported
previously (87%) by Rakes et al. (1999). However, our per capita production
(number of fry reared per breeding adult) in 2012 was 38.2, more than twice that
achieved in the previous study (13.75; Rakes et al. 1999). We attribute the increased
production capacity in 2012 to a variety of factors, including the 44-day
duration of spawning activities that year, the health and condition of the brood
stock, an increased variety of foods offered, and the absence of any serious disease
outbreaks.
Eggs hatched relatively quickly (≈3 days), producing unpigmented immature
yolk-sac larvae that remained benthic for ≈5 days of further development before
swimming up to feed. The amount of time spent in benthic development in 2011–
2012 was longer than the ≈2 days reported by Rakes et al. (1999). After swim-up,
larvae were sufficiently large enough to feed on smaller brine shrimp nauplii,
but rotifers (Brachionus sp.) and early water flea (Ceriodaphnia dubia Richard)
instars were also provided, thereby expanding the early-life-stage dietary options
reported by Rakes et al. (1999).
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Dates and temperatures of spawning
Spawning at the CFI facility began in April when water temperatures warmed
to approximately 16 °C and ended in mid-May when water temperatures reached
20 or 21 °C (Fig. 1). Spawning lasted 14 days in 2011, 44 days in 2012, and at
least 18 days in 2013 (Table 1). We did not track spawning activities beyond 15
May 2013. Mattingly and Black (2103 [this issue]) observed a 32-day spawning
season in their 2006 field study, with spawning events first observed in mid-May
and last observed in mid-June, representing a shift of a few weeks later than captive
spawning. In both field and captive settings, however, spawning ceased when
water temperatures approached 20 or 21 °C (Fig. 1), suggesting that Blackside
Dace may generally cease reproduction above that temperature. Water temperature
may be a stronger cue than day length for initiation of Blackside Dace
spawning, given the notable shift in CFI versus field spawning dates illustrated
in each panel of Figure 1.
Independent spawning
Blackside Dace spawned completely independent of any cues from other host
fish species in 2013, representing the first observation of independent spawning
by the species in captivity. Independent spawning has never been reported in field
studies, but we now know the species is capable of independent spawning in a
captive setting. It remains unknown whether spawning and captive propagation is
enhanced by independent spawning, because we did not measure survival in 2013
as we did in 2011 and 2012. Nevertheless, we do know that captive propagation
is not entirely reliant on obtaining and introducing milt from other fish species.
Table 1. Hatchery conditions and results obtained during 2011, 2012, and 2013 spawning and rearing
of Blackside Dace, Chrosomus cumberlandensis, at the CFI facility in Knoxville, Tennessee.
NM = not recorded or not measured, C = Campostoma sp., Sa = Semotilus atromaculatus, M =
Moxostoma sp.
Year
Characteristic 2011 2012 2013
Number of adult Blackside Dace brood stock 50 50 40
Number and size of spawning aquaria 1, 380-L 2, 170-L 1, 170-L
Other fish species in the same spawning aquaria Yes (C) No No
Other fish species in shared recirculating system Yes (NM) Yes (Sa) No
Addition of milt from other fish species Yes (C) Yes (M, Sa) No
Dates of milt addition 26 April 2 April, 10 April None
Date of first spawning observation 28 April 2 April 28 April
Date of last spawning observation 11 May 15 May NM
Water temperatures during spawning (°C) 17—20 16—21 17—21
Number of fertile eggs collected 117 2,855 840
Time to hatching 3 d 3 d NM
Post-hatching time spent as benthic 5 d 5 d NM
Number of fry produced 71 1,910 NM
Survival from embryo to fry stage 60% 67% NM
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Figure 1. Diamonds represent mean daily water temperatures from the Conservation
Fisheries, Inc. (CFI) facility in Knoxville, TN measured during a) 2011, b) 2012, and c)
2013. Circles represent mean daily water temperatures averaged from five sites in four
Chrosomus cumberlandensis (Blackside Dace) streams in 2006 to generate a single fieldwater-
temperature profile to visually compare to CFI temperatures. Black diamonds and
circles indicate when spawning was observed in each setting, and gray diamonds and
circles indicate when spawning was not observed. Arrows indicate when heterospecific
milt was added in 2011 and 2012 to stimulate Blackside Dace spawning activities. Milt
was not added in 2013.
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Spawning behavior
Spawning behavior generally conformed to that described by Starnes and
Starnes (1981). Spawning activity and nuptial coloration peaked in the morning
hours (in the absence of milt additions), and usually “pods” of 2–3 males were
observed following or courting a gravid female. We observed spawning events
taking place on the mounded substrate “chub nests” provided in the spawning
aquaria. Although presumed spawning acts were observed via closed circuit
video (see Supplemental Video File 1, available online at http://www.eaglehill.
us/SENAonline/suppl-files/s12-Sp4-1040i-Rakes-s1, and, for BioOne subscribers,
at http://dx.doi.org/10.1656/S1040i.s1), actual egg releases were difficult to
observe and searches of the substrate immediately afterwards only occasionally
yielded eggs. At the peak of breeding when water temperature was approximately
18 °C, brilliant coloration was displayed by males and females during spawning
and was most pronounced in late April. Males had a golden dorsum and bright
yellow paired fins and anal fin, while females exhibited more subdued coloration
and lacked yellow or gold.
Conclusions
We successfully spawned and reared Blackside Dace in captivity for the past
three years, with desirable rates of juvenile production in 2012 and 2013. Water
temperature and heterospecific milt appear to be strong abiotic and biotic cues
for induction of spawning, with egg deposition in captivity beginning in April
when water temperatures exceeded 16 or 17 °C. Spawning in a 2006 field study
by Mattingly and Black (2013 [this issue]) started later in May when water temperatures
were generally cooler than at CFI. Spawning in both settings ended as
water temperatures approached or reached 20–21 °C in May (CFI) or June (field).
Captive spawning of reproductively mature and conditioned Blackside Dace can
be triggered within 0–2 days by addition of milt from other fishes, as also noted
by Rakes et al. (1999). However, as we observed in 2013, captive spawning also
can be initiated independently without the presence of other fishes or their milt.
Our observation of independent spawning by Blackside Dace in captivity is the
first reported for this species, yet it remains unknown whether the species practices
independent spawning in a wild setting.
Acknowledgments
This work was completed with funds provided by the Kentucky Aquatic Resources
Fund, Frankfort, KY. Collection and handling of adult Blackside Dace and any offspring
used in toxicity testing were addressed in Kentucky and USFWS permits for obtaining
this federally listed species. Jessica Hendricks and Rebecca Xiques performed important
work at the CFI facility, Tyler Black collected field temperature data, and Matthew
Padgett constructed Figure 1. The manuscript was improved by comments from the guest
editor and two anonymous reviewers.
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Supplemental Video File 1 is available online at https://www.eaglehill.us/SENAonline/
suppl-files/s12-Sp4-1040i-Rakes-s1, and, for BioOne subscribers, at http://dx.doi.
org/10.1656/S1040i.s1.