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Effect of pH on oxygen consumption of the cestode Schistocephalus solidus
EXPERIMENTAL
PARASITOLOGY
22,
201-206
( 1968)
Effect of pH on Oxygen
Cestode
Schistocephalus
M. Walkey Department
Consumption
and
P. Spencer
of the
solicfus Davies
of Zoology, Queen Mary College, London, E.I. and Depurtment of Zoology, The University, Glasgow, W.2. (Submitted
for publication,
8 September
1967)
1968. Effect of pH on oxygen consumption WALKEY, M., AND DAVIES, I?. SPENCER. of the cestode Schistocephalus solidus. Experimental Parasitology 22, 201-206. The endogenous aerobic respiration rates of Schistocephalus solidus (Miiller) were determined for plerocercoid and adult stages of the tapeworm. At both 10°C and 4O”C, plerocercoids exhibited a constant rate of respiration over the entire pH range tested. Adult worms, by comparison, displayed irregular changes in respiratory rate which are difficult to interpret. The absence of respiratory response in plerocercoids can be largely explained by observations that pH in the internal medium of the parasite does not alter with change in the pH of the external medium. It is concluded that the ability to maintain an unchanged pH, and thus metabolic rate, is of adaptive value for the plerocercoid during its passage through the highly acid conditions in the stomach of the avian host prior to attaining sexual maturity in the intestine.
Endoparasitic helminths are generally adapted, by a variety of ecological, physiological, and morphological mechanisms, to tolerate considerable variation in environmental conditions during the course of their life cycles. In many cases resistance to environmental stress is effected by survival as small nonmotile, nonfeeding, or encysted stages, which are metabolically rather inactive compared with their preceding and succeeding stages. The pseudophyllidean cestode, Schistocephalus solidus ( Miiller) is of interest in that some of the most extreme environmental changes experienced occur not during a resistant stage, but when the organism has attained full somatic development as the plerocercoid larva in the body cavity of the three-spined stickleback. After ingestion of the stickleback by the definitive host, a fish-eating bird, the plerocercoid ceases somatic growth and completes genital development to become an adult. During this change of host, the 201
parasite experiences sudden and major changes in temperature, pH, and oxygen tension of its environment. The metabolic responses of the worm to temperature changes have been reported previously (Davies and Walkey, 1966). In the body cavity of the fish host, the plerocercoid is bathed in coelomic fluid of pH 7.2-7.6 ( Walkey, unpublished results). Upon transfer to the avian host however, the worm is subjected to a pH of 2.3 in the gizzard and then to a gradual return to a pH of approximately 7.5 in the caecum (Crompton, 1966). The experiments described below were designed to investigate the effects of this variation in environmental pH upon oxygen, consumption of the tapeworm. MATERIALS
AND METHODS
I Plerocercoids, obtained,..by dissection of infected sticklebacks, were collected &ring the months of May,: June, and Ju&- and .ps
202
WALKIZY A.\;D
used directly in the respiration experiments. Adult worms were obtained by feeding plerocercoids to 2day-old ducklings which were killed 48 hours later for recovery of the parasites. Aerobic endogenous respiratory rates of both adults and plcrocercoids were measured by standard Warburg manometry. Worms were placed singly in 5-ml Warburg flasks together with 2 ml of 25 mM Trismaleate buffered Krebs-Ringer saline (Read et aZ., 1963) adjusted to the appropriate PH. Experimental p1-I values of 4.0, 5.0, 6.0, 6.6, 7.4, and 8.0 were employed at temperatures of 10 and 40°C for plerocercoids and 40°C only for adult worms. Final pH values of incubation media varied by no more than 0.2 from the original levels. Approximately 25 worms were used in each
DAVIES
experiment. The gas phase employed was atmospheric air and the flasks were shaken at a rate of 100 5-cm strokes/minute. At the completion of each experiment, each worm was dried to constant weight and respiratory rates calculated in terms of ,ul OJmg dry weight/hour. RESULTS
To overcome the effects of body size upon metabolism, the statistical treatment followed in a previous work (Davies and Walkey, 1966) was employed for the analysis of respiratory, rates. Thus, for each experiment the rates.were plotted Iogarithmically against dry body weight and regression lines were derived for this relationship by the method of least squares ( Fig. 1) . . Regression coefficients ranged ‘. _
Plerocfrco~ds 40OC:pti
Adults 6.0
,,,I 5
I I I1111 10
5
50
I
I
I
I Illll 5
I
IO
l
1
I
10
‘1
I
50
I
I
I,, SO
III
100
FIG. 1. Examples of the relationship between respiration rate and dry body R/eight in S. solidn. Abscissae indicate dry body weight in milligrams. Ordinates indicate oxygen consuniption in microlitel per milligram dry weight per hour. The b’ values are given for the slopes of the last-squares regressio line.%
203
RESPIRATION IN %histocephah.&s solidus
from b’ = -0.1471 to b’ = -0.6053 (Table I). Covariance analysis of the results indicated no significant variation in these b’ values at the 5%level of significance. Hence, by pooling the sums of squares and products of deviations (Snedecor, 1961) a common regression coefficient for all worms, having the value b’ = -0.4919 was derived. Regression Equations
Temperature
The most interesting feature of the results is the marked lack of variation shown by plerocercoids over the pH range investigated. At both 10 and 40°C there was no significant variation in oxygen consumption at pH values between 4.0 and 8.0. Results for adult worms however were extremely variable and exhibited no discernible trend. These results could arise if either the
TABLE I ,, and Confidence Intervals of Respiration Rate and Dry Weight of Schistocephalus at Dijerent pH Values
and pH
Regression
equation
95ye Confidence limits of b’
95% Confidence limits of P
Plerocercoids 10°C pH 4.0 5.0 6.0 6.6 7.4 8.0
Y Y Y Y Y Y
= = = = = =
-.35Sl - ,569s - .5609 -.5067 - .5649 --.4974X
x X X X X
-.3070 - .1253 - .3216 -.3076 - .0949 -.3025
kO.1771 kO.1361 f0.1359 ,0.1183 stO.0827 *0.3503
iO.0819 +0.1131 kO.0504 f0.0588 io.0443 kO.0742
Plerocercoids 40°C pH 4.0 5.0 6.0 6.6 7.4 8.0
Y Y Y Y Y Y
= = = = = =
- .4377 - .4642 -.4362 -.5264 - .4604 - .4749
x X X X X X
f.7899 + .9392 +.8521 +.9829 + .9110 + .9485
+o. 1133 kO.2693 kO.0597 f0.0779 +o.ow3 rtO.0802
kO.0360 kO.0714 rto.0304 *0.0311 kO.0235 kO.0384
Adults 40°C pH 4.0 5.0 6.0 6.6 7.4 8.0
Y Y Y Y Y Y
= = = = = =
- .3828 - .3174 - .2013 --.1471 -.6053 - .3783
X x X X X x
+ .7421 + .7813 + .6604 +.6751 +1.2344 + .9990
kO.3751 10.1877 rto.0979 +0.1931 kO.4159 +o. 1078
10.0755 Iko.0371 kO.0257 *o.o4oF3 f0.0814 kO.0314
Replotting the regression lines of oxygen consumption upon dry weight to this common regression coefficient, resulted in a series of parallel lines (Fig. 2). From these graphs, the respiration rate of a worm of I8 mg dry weight was calculated for each experiment. Direct comparison of the effects of pH variations upon metabolic rate was now possible using these computed vaIues for a worm of standard weight (Fig. 3).
intracellular respiratory enzymes of the plerocercoid possessed a wide plateau of tolerance to pH change or if the tegument of the worm presented a barrier to the %ow of hydrogen ions so that the internal pH remained unchanged in the different media. In order to test whether pH equilibration took place between the external medium in the flask and the internal medium of the worms, both plerocercoids and adult worms were incubated for periods of up to 20
204
WALKEY
AND
DAVIES
Plerocercoids 4o" c
Adults
20
30
40
50
100
FIG. 2. Regression lines replotted to the common regression coefficient b’ = -0.4919. milligrams dry weight; ordinates = ~1 0, per milligram dry weight per hour.
hours in media of different pH. They were then removed, blotted dry on filter paper, and either homogenized with 0.25 ml of distilled water or pierced with a hypodermic needle containing the thin polythene catheter of a “Radiometer” micro-pH electrode. Both methods of measuring internal pH gave similar results. The results (Table II) show that except at the extremes of pH, under which conditions the worms became moribund within 2 hours, the pH of the internal medium of both plerocercoids and adults remained constant and independent of the pH of the external medium. DISCUSSION
It has been shown that for mammalian cells, both the growth rate (Paul, 1959) and respiration rate (Danes and Paul,
Abscissae =
1961) in tissue cultures are markedly influenced by the.pH of the external medium, having an optimum at about pH 7.4. The lack of response by plerocercoids of Schistocephalus to changes in pH of the medium is clearly at variance with these observations. However, the lack of pH equilibration between the external and internal media (Table II) would suggest that the actively respiring cells of the tapeworm do not experience any change in their environmental pH and this is probably adequate explanation for the absence of response by plerocercoids. A similar lack of respiratory response to pH change was recorded by von Brand (1943) in the larvae of the nematode Eustrongylides ignotus over the pH range 3.48.3. The mechanism involved in maintaining a constant internal pH has not been in-
L-a-
-*--“.--
,c@
Plerocercoids 4o’I c
0.5
0.25 -
l -.
/“\ePlerocercoids
,-‘-.
I 5.0
a.0
I
I
I
6.0
7.0
8.0
loo
c
PH
FIG. 3. Effect of pH on the oxygen Adult worms = 0; plerocercoids = 0.
consumption
pH of worm after incubation for pH of meI dium hours
2 hours
10 hours
20 hours
10°C Plerocercoids
2.0 4.0 6.0 8.0 10.0
8.7 8.5 8.3 8.5 9.3
8.7 8.5 8.5 8.5 9.2
(2.8)a 8.6 8.5 8.5 9.3
(2.8) 8.6 8.5 8.5 lC.0
40°C Plerocercoids
2.0 4.0 6.0 8.0
8.0 8.0 8.3
8.3 8.0 8.3 8.2
6.5 8.3 8.3
6.5 8.C 8.1
40°C Adults
2.0 4.0 6.0 8.0
8.2 8.3 8.3 8.5
(8.0) 8.0 8.3 8.5
(2.2) (7.6) 8.2 8.5
(2.0) (7.0) 8.0 8.5
worms
at 10°C and 40°C.
vestigated in the present work but the lack of respiratory response would suggest that an energy consuming regulatory process is not involved. Von Brand (1933) suggested that the intracellular calcareous corpuscles of the tapeworm Moniezia expansa serve to buffer acids absorbed through the tegument and those produced during anaerobic glycolysis. Calcareous corpuscles are also found within Schistocephalus and may quite possibly fulfill a similar function. The response of the adult worms presents something of an anomaly since it was found that the internal pH of these forms also remains constant during variations in external pH. It is possible that after 2 days in the gut of a duck, at environmental oxygen tensions down to 0.5 mm Hg, there is an alteration in the buffering capacity of adult worms which experiments at atmospheric oxygen tensions (Table II) failed to define. Von Brand et al. (1960) in fact recorded that the calcareous corpuscles of T. taeniae-
TABLE II Internal pH of Schistocephalus Following Incubation in Media oj Dijerent pH BalueP
Temperature and stage of development.
of 18 mg dry weight
Q Figures in parenthesis were derived from worms which were either moribund or exhibiting visual characteristics of stress. 205
206
WALKEY
formis tended to decrease in numbers when the tapeworm was maintained in a medium at low partial pressures of oxygen. For plerocercoids however, there is considerable advantage in possessing a mechanism for the maintenance of a constant internal pH, and hence metabolic rate. At the time when the parasite is experiencing its greatest variations in external pH, immediately after ingestion by the definitive host, it is still a plerocercoid. Transition to the adult stage is not completed until the worm has reached the intestine (McCaig and Hopkins, 1963), by which time the pH of the medium is again relatively stable at or near neutrality. ACKNOWLEDGMENTS We wish to thank Mr. Robert A. R. Young for technical assistance, and Dr. C. A. Hopkins for his assistance in the provision of infected sticklebacks. One of us (M.W.) would also like to acknowledge the receipt of a grant from the Wellcome Trust to finance technical assistance and from the Royal Society for the purchase of the Radiometer micropH assembly. REFERENCES CROMPTON, D. W. T. 1966. Measurements of glucose and amino acid concentrations, temperature and pH in the habitat of Polymorphus minutus ( Acanthocephala) in the intestine of domestic ducks. Journal of Experimental Biology 45, 279-284. DANES, B. S., AND PAUL, J. 1961. Respiration of
AND
DAVIES
normal and tumour cells in culture. Pathologic et biologie. Paris 9, 571-573. DAVIES, P., SPENCER, AND WALKEY, M. 1966. The effect of body size and temperature upon oxygen consumption of the cestode Schistocephalus solidus ( Miiller ). Comparative Biochemistry and Physiology 18, 415425. MCCAIC, M. L. O., AND HOPKINS, C. A. 1963. Studies on Schistocephalus solidus. 11. Establishment and longevity in the definitive host. Experimental Parasitology 13, 273-283. PAUL, J. 1959. Environmental influences on the metabolism and composition of cultured cells. Journal of Experimental Zoology 142, 475505. READ, C. P., ROTHMAN, A. H., AND SIMMONS, J. E. 1963. Studies on membrane transport, with special reference to parasite-host integration. Annals of the New York Academy of Sciences 113, 154-205. SNEDECOR, G. W. 1961. “Statistical Methods Applied to Experiments in Agriculture and Biology,” p. 398. 5th ed. Iowa State Univ. Press, Ames, Iowa. VON BRAND, T. 1933. Untersuchungen iiber den Stoffbestand einiger Cestoden und den Stoffweschsel von Moniezia expansa. Zeitschrift fiir vergleichende Physiologie 18, 56s-596. VON BRAND, T. 1943. Physiological observations upon a larval Eustrongylides. IV. Influence of temperature, pH and inorganic ions upon the Biological Bulletin. oxygen consumption. Marine Biological Laboratory, Woods Hole 84, 148-156. VON BRAND, T., MERCADO, T. I., NYLEN, M. U., AND SCOTT, D. B. 1960. Observations on the function, composition and structure of cestode calcareous corpuscles. Experimental Parasitology 9, 205-214.
PARASITOLOGY
22,
201-206
( 1968)
Effect of pH on Oxygen
Cestode
Schistocephalus
M. Walkey Department
Consumption
and
P. Spencer
of the
solicfus Davies
of Zoology, Queen Mary College, London, E.I. and Depurtment of Zoology, The University, Glasgow, W.2. (Submitted
for publication,
8 September
1967)
1968. Effect of pH on oxygen consumption WALKEY, M., AND DAVIES, I?. SPENCER. of the cestode Schistocephalus solidus. Experimental Parasitology 22, 201-206. The endogenous aerobic respiration rates of Schistocephalus solidus (Miiller) were determined for plerocercoid and adult stages of the tapeworm. At both 10°C and 4O”C, plerocercoids exhibited a constant rate of respiration over the entire pH range tested. Adult worms, by comparison, displayed irregular changes in respiratory rate which are difficult to interpret. The absence of respiratory response in plerocercoids can be largely explained by observations that pH in the internal medium of the parasite does not alter with change in the pH of the external medium. It is concluded that the ability to maintain an unchanged pH, and thus metabolic rate, is of adaptive value for the plerocercoid during its passage through the highly acid conditions in the stomach of the avian host prior to attaining sexual maturity in the intestine.
Endoparasitic helminths are generally adapted, by a variety of ecological, physiological, and morphological mechanisms, to tolerate considerable variation in environmental conditions during the course of their life cycles. In many cases resistance to environmental stress is effected by survival as small nonmotile, nonfeeding, or encysted stages, which are metabolically rather inactive compared with their preceding and succeeding stages. The pseudophyllidean cestode, Schistocephalus solidus ( Miiller) is of interest in that some of the most extreme environmental changes experienced occur not during a resistant stage, but when the organism has attained full somatic development as the plerocercoid larva in the body cavity of the three-spined stickleback. After ingestion of the stickleback by the definitive host, a fish-eating bird, the plerocercoid ceases somatic growth and completes genital development to become an adult. During this change of host, the 201
parasite experiences sudden and major changes in temperature, pH, and oxygen tension of its environment. The metabolic responses of the worm to temperature changes have been reported previously (Davies and Walkey, 1966). In the body cavity of the fish host, the plerocercoid is bathed in coelomic fluid of pH 7.2-7.6 ( Walkey, unpublished results). Upon transfer to the avian host however, the worm is subjected to a pH of 2.3 in the gizzard and then to a gradual return to a pH of approximately 7.5 in the caecum (Crompton, 1966). The experiments described below were designed to investigate the effects of this variation in environmental pH upon oxygen, consumption of the tapeworm. MATERIALS
AND METHODS
I Plerocercoids, obtained,..by dissection of infected sticklebacks, were collected &ring the months of May,: June, and Ju&- and .ps
202
WALKIZY A.\;D
used directly in the respiration experiments. Adult worms were obtained by feeding plerocercoids to 2day-old ducklings which were killed 48 hours later for recovery of the parasites. Aerobic endogenous respiratory rates of both adults and plcrocercoids were measured by standard Warburg manometry. Worms were placed singly in 5-ml Warburg flasks together with 2 ml of 25 mM Trismaleate buffered Krebs-Ringer saline (Read et aZ., 1963) adjusted to the appropriate PH. Experimental p1-I values of 4.0, 5.0, 6.0, 6.6, 7.4, and 8.0 were employed at temperatures of 10 and 40°C for plerocercoids and 40°C only for adult worms. Final pH values of incubation media varied by no more than 0.2 from the original levels. Approximately 25 worms were used in each
DAVIES
experiment. The gas phase employed was atmospheric air and the flasks were shaken at a rate of 100 5-cm strokes/minute. At the completion of each experiment, each worm was dried to constant weight and respiratory rates calculated in terms of ,ul OJmg dry weight/hour. RESULTS
To overcome the effects of body size upon metabolism, the statistical treatment followed in a previous work (Davies and Walkey, 1966) was employed for the analysis of respiratory, rates. Thus, for each experiment the rates.were plotted Iogarithmically against dry body weight and regression lines were derived for this relationship by the method of least squares ( Fig. 1) . . Regression coefficients ranged ‘. _
Plerocfrco~ds 40OC:pti
Adults 6.0
,,,I 5
I I I1111 10
5
50
I
I
I
I Illll 5
I
IO
l
1
I
10
‘1
I
50
I
I
I,, SO
III
100
FIG. 1. Examples of the relationship between respiration rate and dry body R/eight in S. solidn. Abscissae indicate dry body weight in milligrams. Ordinates indicate oxygen consuniption in microlitel per milligram dry weight per hour. The b’ values are given for the slopes of the last-squares regressio line.%
203
RESPIRATION IN %histocephah.&s solidus
from b’ = -0.1471 to b’ = -0.6053 (Table I). Covariance analysis of the results indicated no significant variation in these b’ values at the 5%level of significance. Hence, by pooling the sums of squares and products of deviations (Snedecor, 1961) a common regression coefficient for all worms, having the value b’ = -0.4919 was derived. Regression Equations
Temperature
The most interesting feature of the results is the marked lack of variation shown by plerocercoids over the pH range investigated. At both 10 and 40°C there was no significant variation in oxygen consumption at pH values between 4.0 and 8.0. Results for adult worms however were extremely variable and exhibited no discernible trend. These results could arise if either the
TABLE I ,, and Confidence Intervals of Respiration Rate and Dry Weight of Schistocephalus at Dijerent pH Values
and pH
Regression
equation
95ye Confidence limits of b’
95% Confidence limits of P
Plerocercoids 10°C pH 4.0 5.0 6.0 6.6 7.4 8.0
Y Y Y Y Y Y
= = = = = =
-.35Sl - ,569s - .5609 -.5067 - .5649 --.4974X
x X X X X
-.3070 - .1253 - .3216 -.3076 - .0949 -.3025
kO.1771 kO.1361 f0.1359 ,0.1183 stO.0827 *0.3503
iO.0819 +0.1131 kO.0504 f0.0588 io.0443 kO.0742
Plerocercoids 40°C pH 4.0 5.0 6.0 6.6 7.4 8.0
Y Y Y Y Y Y
= = = = = =
- .4377 - .4642 -.4362 -.5264 - .4604 - .4749
x X X X X X
f.7899 + .9392 +.8521 +.9829 + .9110 + .9485
+o. 1133 kO.2693 kO.0597 f0.0779 +o.ow3 rtO.0802
kO.0360 kO.0714 rto.0304 *0.0311 kO.0235 kO.0384
Adults 40°C pH 4.0 5.0 6.0 6.6 7.4 8.0
Y Y Y Y Y Y
= = = = = =
- .3828 - .3174 - .2013 --.1471 -.6053 - .3783
X x X X X x
+ .7421 + .7813 + .6604 +.6751 +1.2344 + .9990
kO.3751 10.1877 rto.0979 +0.1931 kO.4159 +o. 1078
10.0755 Iko.0371 kO.0257 *o.o4oF3 f0.0814 kO.0314
Replotting the regression lines of oxygen consumption upon dry weight to this common regression coefficient, resulted in a series of parallel lines (Fig. 2). From these graphs, the respiration rate of a worm of I8 mg dry weight was calculated for each experiment. Direct comparison of the effects of pH variations upon metabolic rate was now possible using these computed vaIues for a worm of standard weight (Fig. 3).
intracellular respiratory enzymes of the plerocercoid possessed a wide plateau of tolerance to pH change or if the tegument of the worm presented a barrier to the %ow of hydrogen ions so that the internal pH remained unchanged in the different media. In order to test whether pH equilibration took place between the external medium in the flask and the internal medium of the worms, both plerocercoids and adult worms were incubated for periods of up to 20
204
WALKEY
AND
DAVIES
Plerocercoids 4o" c
Adults
20
30
40
50
100
FIG. 2. Regression lines replotted to the common regression coefficient b’ = -0.4919. milligrams dry weight; ordinates = ~1 0, per milligram dry weight per hour.
hours in media of different pH. They were then removed, blotted dry on filter paper, and either homogenized with 0.25 ml of distilled water or pierced with a hypodermic needle containing the thin polythene catheter of a “Radiometer” micro-pH electrode. Both methods of measuring internal pH gave similar results. The results (Table II) show that except at the extremes of pH, under which conditions the worms became moribund within 2 hours, the pH of the internal medium of both plerocercoids and adults remained constant and independent of the pH of the external medium. DISCUSSION
It has been shown that for mammalian cells, both the growth rate (Paul, 1959) and respiration rate (Danes and Paul,
Abscissae =
1961) in tissue cultures are markedly influenced by the.pH of the external medium, having an optimum at about pH 7.4. The lack of response by plerocercoids of Schistocephalus to changes in pH of the medium is clearly at variance with these observations. However, the lack of pH equilibration between the external and internal media (Table II) would suggest that the actively respiring cells of the tapeworm do not experience any change in their environmental pH and this is probably adequate explanation for the absence of response by plerocercoids. A similar lack of respiratory response to pH change was recorded by von Brand (1943) in the larvae of the nematode Eustrongylides ignotus over the pH range 3.48.3. The mechanism involved in maintaining a constant internal pH has not been in-
L-a-
-*--“.--
,c@
Plerocercoids 4o’I c
0.5
0.25 -
l -.
/“\ePlerocercoids
,-‘-.
I 5.0
a.0
I
I
I
6.0
7.0
8.0
loo
c
PH
FIG. 3. Effect of pH on the oxygen Adult worms = 0; plerocercoids = 0.
consumption
pH of worm after incubation for pH of meI dium hours
2 hours
10 hours
20 hours
10°C Plerocercoids
2.0 4.0 6.0 8.0 10.0
8.7 8.5 8.3 8.5 9.3
8.7 8.5 8.5 8.5 9.2
(2.8)a 8.6 8.5 8.5 9.3
(2.8) 8.6 8.5 8.5 lC.0
40°C Plerocercoids
2.0 4.0 6.0 8.0
8.0 8.0 8.3
8.3 8.0 8.3 8.2
6.5 8.3 8.3
6.5 8.C 8.1
40°C Adults
2.0 4.0 6.0 8.0
8.2 8.3 8.3 8.5
(8.0) 8.0 8.3 8.5
(2.2) (7.6) 8.2 8.5
(2.0) (7.0) 8.0 8.5
worms
at 10°C and 40°C.
vestigated in the present work but the lack of respiratory response would suggest that an energy consuming regulatory process is not involved. Von Brand (1933) suggested that the intracellular calcareous corpuscles of the tapeworm Moniezia expansa serve to buffer acids absorbed through the tegument and those produced during anaerobic glycolysis. Calcareous corpuscles are also found within Schistocephalus and may quite possibly fulfill a similar function. The response of the adult worms presents something of an anomaly since it was found that the internal pH of these forms also remains constant during variations in external pH. It is possible that after 2 days in the gut of a duck, at environmental oxygen tensions down to 0.5 mm Hg, there is an alteration in the buffering capacity of adult worms which experiments at atmospheric oxygen tensions (Table II) failed to define. Von Brand et al. (1960) in fact recorded that the calcareous corpuscles of T. taeniae-
TABLE II Internal pH of Schistocephalus Following Incubation in Media oj Dijerent pH BalueP
Temperature and stage of development.
of 18 mg dry weight
Q Figures in parenthesis were derived from worms which were either moribund or exhibiting visual characteristics of stress. 205
206
WALKEY
formis tended to decrease in numbers when the tapeworm was maintained in a medium at low partial pressures of oxygen. For plerocercoids however, there is considerable advantage in possessing a mechanism for the maintenance of a constant internal pH, and hence metabolic rate. At the time when the parasite is experiencing its greatest variations in external pH, immediately after ingestion by the definitive host, it is still a plerocercoid. Transition to the adult stage is not completed until the worm has reached the intestine (McCaig and Hopkins, 1963), by which time the pH of the medium is again relatively stable at or near neutrality. ACKNOWLEDGMENTS We wish to thank Mr. Robert A. R. Young for technical assistance, and Dr. C. A. Hopkins for his assistance in the provision of infected sticklebacks. One of us (M.W.) would also like to acknowledge the receipt of a grant from the Wellcome Trust to finance technical assistance and from the Royal Society for the purchase of the Radiometer micropH assembly. REFERENCES CROMPTON, D. W. T. 1966. Measurements of glucose and amino acid concentrations, temperature and pH in the habitat of Polymorphus minutus ( Acanthocephala) in the intestine of domestic ducks. Journal of Experimental Biology 45, 279-284. DANES, B. S., AND PAUL, J. 1961. Respiration of
AND
DAVIES
normal and tumour cells in culture. Pathologic et biologie. Paris 9, 571-573. DAVIES, P., SPENCER, AND WALKEY, M. 1966. The effect of body size and temperature upon oxygen consumption of the cestode Schistocephalus solidus ( Miiller ). Comparative Biochemistry and Physiology 18, 415425. MCCAIC, M. L. O., AND HOPKINS, C. A. 1963. Studies on Schistocephalus solidus. 11. Establishment and longevity in the definitive host. Experimental Parasitology 13, 273-283. PAUL, J. 1959. Environmental influences on the metabolism and composition of cultured cells. Journal of Experimental Zoology 142, 475505. READ, C. P., ROTHMAN, A. H., AND SIMMONS, J. E. 1963. Studies on membrane transport, with special reference to parasite-host integration. Annals of the New York Academy of Sciences 113, 154-205. SNEDECOR, G. W. 1961. “Statistical Methods Applied to Experiments in Agriculture and Biology,” p. 398. 5th ed. Iowa State Univ. Press, Ames, Iowa. VON BRAND, T. 1933. Untersuchungen iiber den Stoffbestand einiger Cestoden und den Stoffweschsel von Moniezia expansa. Zeitschrift fiir vergleichende Physiologie 18, 56s-596. VON BRAND, T. 1943. Physiological observations upon a larval Eustrongylides. IV. Influence of temperature, pH and inorganic ions upon the Biological Bulletin. oxygen consumption. Marine Biological Laboratory, Woods Hole 84, 148-156. VON BRAND, T., MERCADO, T. I., NYLEN, M. U., AND SCOTT, D. B. 1960. Observations on the function, composition and structure of cestode calcareous corpuscles. Experimental Parasitology 9, 205-214.