Comparison of Metabolic Rates of Guppy, Poecilia reticulata (Poecilidae), and Black Molly, Poecilia latipinna (Poecilidae), at Different Temperatures.

Julius O. Ikenga [1]

This research was designed to compare the metabolic rates (mRs) of guppy, Poecilia reticulata (Poecilidae) and black molly, Poecilia latipinna (Poecilidae) at 23-25[degrees]C and at 15[degrees]C, using the indirect respirometry. Most aquaria kept at homes are maintained in the 23-25[degrees]C temperature range. The 15[degrees]C was chosen to test the fishes' survivability at a lower temperature and to compare the mRs at different temperatures. Guppies and mollies are popular, small aquarium pets. The male guppies are about 3 centimeters long and are naturally smaller than the male black mollies. Both fishes, along with other similar aquarium pets, are viviparous; which enabled them to occupy an important niche in fish evolution. Using end-point titration, we measured the metabolic carbon dioxide ([CO.sub.2]) produced by the guppy and the black molly that displaced 1 and 2 milliliters of water, respectively. Endpoint data collected were adjusted and used to calculate the mRs in [micro]M [CO.sub.2]ml/hr. The a verage mRs of the guppy was found to be 7.11, 10.45, 11.58, and 9.02 [micro]M [CO.sub.2]/ml/hr at 15, 23, 24, and 25[degrees]C, respectively. The black molly at the above temperatures had average mRs of 4.78, 6.6, 7.40, and 5.78 [micro]M [CO.sub.2]/ml/hr, respectively. Both fishes survived testing at 15[degrees]C. The black molly however, regulated its mR much more tightly (5.4 to 4.5 [micro]M [CO.sub.2]/ml/hr) at 15[degrees]C than the guppy did (8.0 to 6.3 [micro]M [CO.sub.2]/ml/hr) in the same temperature range. The two species responded metabolically to changes in water temperature. But, it is unclear how much of the observed differences in mR were due to species or weight differences. Similar research using the female guppies which can measure up to 6 cm long and the female black mollies of the same body weight is recommended to resolve the question on weight differences.

Guppy and black molly are small, tropical aquarium pets. The male guppy is about 3 cm long and by nature is smaller than the male black molly. Both the guppies and the black mollies are closely related to the swordtails, platys and mosquito fish. Their uniqueness as live bearers has enabled them to occupy an important niche in the evolution of fishes. Guppy is a native to the fresh waters of Venezuela and the Caribbean Islands, where water temperature is usually at least 20[degrees]C. The demand for guppy in the last decade has increased immensely that it now rivals goldfish as the world's most popular aquarium pet. Molly on the other hand, is generally found in the fresh-waters between Mexico and Venezuela. It is particularly vulnerable to cold temperatures. Ready availability from commercial sources and low maintenance make both fishes very attractive for use in research. The guppies in particular have already been dubbed the laboratory mice in the aquaria. However, majority of natural habitats for the gup py and the molly and other live bearers is under threat through improper landscaping, logging, or recreational use.

Guppy and black molly, like many other gill-breathing animals, continually exchange carbon dioxide for oxygen with their ambient environment. Most fishes further enhance exchange of gases by using ram ventilation (Muir and Kendall, 1968; Eckert and Randall, 1983), or the buccal-opercular pump (Steffensen and Lambolt, 1983; Steffensen, 1985). Additionally, many fishes are able to switch selectively between the two modes of gas exchange. Steffensen (1985) suggested possible involvement of both chemoreceptors and mechanoreceptors in the mode-switching reflex. At the cellular level, oxygen is used as the final electron acceptor during aerobic respiration. As a result, energy, metabolic carbon dioxide and water are produced (Pelster and Driedzic, 1994; Pelster, 1995a, 1995b; Pelster and Port, 1996). The rate of production of metabolic carbon dioxide depends on the animal's complexity, activity level, and temperature. These same factors also affect the amount of oxygen the animal consumes (Hughes, 1965). In this research we quantified the metabolic rates (mRs) of guppy (Poecilia reticulata) and the black molly (Poecilia latipinna) using the indirect respirometry at 23-25[degrees]C and at 15[degrees]C. The latter technique enabled measurements of metabolic rates (mRs) using the produced carbon dioxide as a target molecule.

MATERIALS AND METHODS

This research was conducted at Mississippi Valley State University, Itta Bena, MS from October 1997 to April 2000. Three 10 gallon aquarium tanks filled with de-chlorinated tap water were stocked with guppies and black mollies from a local supplier. Each guppy was found to displace 1 ml of water in a graduated cylinder while each molly displaced 2 ml of water in a graduated cylinder. The water in the stock tanks were continuously filtered and aerated with Aqua-Tech filtration units. All fishes were allowed to acclimate for about two weeks before use in the experiment (Hochachaka and Somero, 1971). Fishes were fed once a day with Tropical flakes. The research methods we used were modified from Skavaril et al. (1993). Fifty ml of aquarium water at 23[degrees]C was measured with a graduated cylinder and poured into three, clean 100 ml beakers labeled 1, 2, and 3. A guppy and a black molly were scooped each from its tank with a dip net and gently transferred into beakers 2 and 3, respectively. Beaker 1 served as the control setup. Each beaker was tightly sealed with a clinging-plastic wrap and secured with a rubber band. The setups were then allowed to sit undisturbed for 30 minutes, during which the fishes carried on their routine activities. Next, the plastic seal was quickly removed from the three test-beakers and the fishes in beakers 2 and 3 were transferred with a perforated plastic spoon into 50 ml graduated cylinders. The latter cylinders contained 20 ml of aquarium water that was taken from the respective stock tanks. Contamination or loss of water from the above beakers was avoided during the fishes' transfer by using dedicated, perforated plastic spoon for each fish and completely draining all the water from the spoon back into the beaker.

Eight drops of phenolphthalein solution were then added to the test-waters in each of the three beakers as a color indicator. The test-waters were each titrated to endpoint with a burette that contained 2.5 [micro]M of sodium hydroxide (NaOH) solution. The amount of NaOH solution used to neutralize the carbonic acid in the control was deducted from the amounts used in beakers 2 and 3. The resulting data represent the adjusted volumes of NaOH used for the test-waters in beakers 2 and 3 that previously contained the test fishes and hence, were used to calculate the metabolic rates (mRs) of the fishes in [micro]M [CO.sub.2]/ml/hr. The mass of each fish was determined as the difference between the final and the initial water volumes in the 50 ml graduated cylinders. The above procedures were repeated at 24, 25, and 15[degrees]C and the fishes were rested 24 h between tests. The mR was calculated using the formula: mR [ml NaOH (adjusted)] x [2.5 [micro]M NaOH/ ml] /[Fish volume (ml)] x [0.5 hr].

RESULTS

The mRs of black molly at 23 to 25[degrees]C ranged from 5.0 to 14.3 [micro]M [CO.sub.2]/ml/hr (Table 1). For guppy in the above temperature ranges, the mRs ranged from 9.0 to 13.5 [micro]M [CO.sub.2]/ml/hr (Table 1). The average mRs for guppy were 10.45, 11.58, and 9.02 [micro]M [CO.sub.2]/ml/hr at 23, 24, and 25[degrees]C (Table 2). In comparison, the average mRs for black molly at the above temperatures were 6.6, 7.40, and 5.78 [micro]M [CO.sub.2]/ml/hr (Table 2). At 15[degrees]C the mRs of the black molly ranged from 4.5 to 5.4 [micro]M [CO.sub.2]/ml/hr (Table 3). Also at 15[degrees]C, the mRs of guppy ranged from 6.3 to 8.0 [micro]M [CO.sub.2]/ml/hr (Table 3). Figure 1 shows the average mRs of black molly and guppy in [micro]M [CO.sub.2]/ml/hr at 15[degrees]C and at 23 to 25[degrees]C. Metabolism for both fishes was lowest at 15[degrees]C and highest at 24[degrees]C (Fig. 1). No mortality was observed at any of the temperatures tested.

DISCUSSION

In the absence of high technological equipment, the indirect respirometry is a low cost and effective means of measuring the metabolic rates of small aquatic animals. Our data show that both the guppy and the black molly actively carried on metabolic processes during the test periods. The metabolic rates (mRs) for both fishes appear to increase as the ambient temperature increased (Fig. 1). Such increases in mR relative to rising temperatures do not go on indefinitely, as is the case with the two fish species we tested.

The increases in the mR that both species exhibited were gradual from 15 to 24[degrees]C before dropping at 25[degrees]C. The decline in the mR observed for the guppy and the black molly after 24[degrees]C suggests that both fishes may have definite upper temperature tolerance for routine activities. The latter are activities supported by routine metabolism. Generally, when routine metabolism occurs, it permits fish to swim continuously, undisturbed, and without tiring (Hughes, 1965; Fry, 1971; Milligan, 1996). Such swimming has also been described as steady-state swimming and to involve a broad range of swimming speeds (Milligan, 1996), as are frequently observed in most home aquarium tanks. Routine metabolism is important for fish normal cruises in its territory and for long distance travels. Both activities are also essential for fish survival and are largely supported by aerobic respiration, which occurs in the red muscles. There, the pyruvate molecules are metabolized to [CO.sub.2] and [H.sub.2]O in the mitochondria using oxygen molecules as the final electron acceptors.

The black molly we tested has twice the mass of the guppy. The guppy consistently showed a higher mR at each temperature tested. This observation supports the general knowledge that smaller animals tend to have higher mRs when compared to larger animals on equal weight basis (Hughes, 1965). Black mollies are particularly vulnerable to cold temperatures. This may explain why they regulated their mRs much more tightly, between 4.5 and 5.4 [micro]M [CO.sub.2]/ml/hr at 1 15[degrees]C, as compared to the guppy (6.3 to 8.0 [micro]M [CO.sub.2]/ml/hr) at the same temperature. It is unclear how much of the differences in mR that we observed is purely attributable to differences in species, and if any, to weight differences. Additional study is therefore warranted to resolve the point of differences.

(1.) Author for correspondence.

LITERATURE CITED

Eckert, Roger, and David Randall, 1983. Animal Physiology: Mechanisms and adaptations. W H. Freeman and Co., NY, pp. 476-512.

Fry, F.E.J. 1971. The effect of environmental factors on the physiology of fish. Pages 11-98 in W.S. Hoar, and D.J. Randall, eds. Fish Physiology, vol. VII. Academic Press, NY.

Hochachka, P.W., and G.N. Somero. 1971. Biochemical adaptation to the environment. Pages 100-148 in W.S. Hoar and D.J. Randall, eds. Fish physiology, vol. VI. Academic Press; NY.

Hughes, G.M. 1965. Comparative physiology of vertebrate respiration. Harvard University Press, Cambridge, MA, pp. 1-40.

Milligan, C.L. 1996. Metabolic recovery from exhaustive exercise in rainbow trout. Rev. Comp. Biochem Physiol. 113A:51-60.

Muir, B.S., and I.J. Kendall. 1968. Structural modifications in the gills of tunas and some other oceanic fishes. Copeia 2:388-398.

Pelster, B., J. Hichs and W. R. Driedzic. 1994. Contribution of the phosphate shunt to the formation of [CO.sub.2] in swimbladder tissue of the eel. J. Exp. Biol. 197:119-128.

Pelster, B. 1995a. Metabolism of the swimbladder tissue. Biochem. Molec. Biol. Fishes 4:101-118.

Pelster, B. 1995b. Mechanisms of acid release in isolated gas gland cells of the European eel, Anguilla anguilla. Am. J. Physiol. 270: R793-R799.

Pelster, B., and L. Pott. 1996. Control of acid secretion in cultured gas gland cells of the European eel, Anguilla anguilla. Am. J. Physiol. 270: R578-R58.

Skavaril, R., M. Finnen, and S. Lawton. 1993. General biology lab manual: Investigations into life's phenomena. Sanders College Publishing, N.Y., pp. 73-75.

Steffensen, J.F., and J.P. Lamholt. 1983. Energetic cost of branchial ventilation in sharksucker, Echeneis naucrates. L. Exp. Biol. 103:185-192.

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Figure 1. Average metabolic rates of
guppy (Poecilia reticulata) and black
molly (Poecilia latipinna) based on
metabolic carbon dioxide produced in
[micro]M/ml/hr.
    Guppy  Black Molly
15   7.11     4.78
23  10.45      6.6
24  11.58      7.4
25   9.02     5.78
Note: Table made from bar graph
Table 1. Metabolic rates of guppy
(Poecilia reticulata) and black molly
(Poecilia latipinna) based on metabolic
carbon dioxide produced at
23-25[degrees]C. Gu, guppy, and Bm,
black molly.
Experiment                    2.5 [micro]M NaOH solution used (ml)
Test No.    Temp. [degrees]C              Experimental
                                             Setup
                                               Gu
1                  23                         2.40
2                  23                         2.20
3                  23                         2.50
4                  23                         2.60
5                  23                         2.40
6                  24                         3.00
7                  24                         3.00
8                  24                         2.50
9                  24                         2.70
10                 24                         2.65
11                 25                         2.85
12                 25                         2.70
13                 25                         2.50
14                 25                         2.80
15                 25                         2.60
Experiment
Test No.          Control  Adjusted        [CO.sub.2] Produced
                   Setup    Volume          ([micro]M/ml/hr)
             Bm               Gu      Bm           Gu             Bm
1           3.40   0.20      2.20    3.20         11.00          8.00
2           2.40   0.40      1.80    2.00          9.00          5.00
3           2.70   0.40      2.10    2.30         11.50          5.57
4           2.50   0.45      2.15    2.05         10.75          5.13
5           3.40   0.40      2.00    3.00         10.00          7.50
6           6.00   0.30      2.70    5.70         13.50         14.30
7           2.80   0.55      2.45    2.25         12.30          5.63
8           3.75   0.45      2.05    3.30         10.30          8.25
9           2.80   0.40      2.30    2.40         11.50          6.00
10          3.75   0.50      2.15    3.25         10.80          8.13
11          3.15   0.45      2.40    2.70         12.00          6.75
12          4.00   0.45      2.25    3.55         11.30          8.88
13          3.00   0.45      2.05    2.55         10.30          6.38
14          3.20   0.45      2.35    2.75         11.75          6.88
15          3.60   0.45      2.25    3.15         10.80          7.90
Table 2. Average metabolic rates for
guppy (Poecilia reticulata) and black
molly (Poecilia latipinna) based on
metabolic carbon dioxide produced in
[micro]M/ml/hr.
Fish         15[degrees]C  23[degrees]C  24[degrees]C  25[degrees]C
Guppy        7.11          10.45         11.58         9.02
Black Molly  4.78          6.60          7.40          5.78
Table 3. Metabolic rates of guppy
(Poecilia reticulata) and black molly
(Poecilia latipinna) based on metabolic
carbon dioxide produced at 15[degrees]C.
Gu, guppy, and Bm, black molly.
          2.5 [micro]M NaOH solution Used (ml)
Test No.  Experimental Control                        Control
                 Setup                                 Setup
                   Gu                            Bm
1                 1.90                          2.45   0.30
2                 1.60                          2.10   0.30
3                 1.50                          2.10   0.25
4                 1.65                          2.15   0.35
5                 1.75                          2.25   0.35
6                 2.05                          2.30   0.45
7                 1.75                          2.25   0.45
8                 2.00                          2.45   0.40
9                 1.85                          2.40   0.40
10                1.75                          2.25   0.35
Test No.  Adjusted        [CO.sub.2] Produced
           Volume          ([micro]M/ml/hr)
             Gu      Bm           Gu            Bm
1           1.60    2.15         8.00          5.40
2           1.30    1.80         6.50          4.50
3           1.25    1.85         6.30          4.60
4           1.30    1.80         6.50          4.50
5           1.40    1.90         7.00          4.80
6           1.60    1.85         8.00          4.60
7           1.30    1.80         6.50          4.50
8           1.60    2.05         8.00          5.10
9           1.45    2.00         7.30          5.00
10          1.40    1.90         7.00          4.80