1999

Vol. 8, pp. 53-59

Asiatic Herpetological Research

Utilization of Energy and Material in Eggs and Post-hatching Yolk in an

Oviparous Snake, Elaphe taeniura
XlANG Jl', PlNG-YUE SUN

1,

SHUI-YU FU

2,

AND HUA-SONG ZHANG 2

Department of Biology, Hangzhou Normal College, Hangzhou 310036, Zhejiang, China; Email (XJ):
xji@puhlic.hz-zj.cn; ''Department of Chemistry, Hangzhou Normal College, Hangzhou 310036, Zhejiang, China

Abstract. -The duration of incubation of Elaphe taeniura eggs at 30±0.3°C averaged 54.9 days. During
incubation, pliable-shelled eggs of E. taeniura increased in wet mass. Dried shells of the freshly laid egg
caveraged 17.6 /< of the entire egg dry mass. Freshly laid eggs had significantly heavier shells than did hatchedwiththesame wet mass at oviposition. Dry mass conversion from egg contents of the freshly laid egg to
eggs

hatchling averaged 84.5%. During incubation, approximately 74.6% of non-polar lipids and 80.8% of energy inegg contents of the freshly laid egg were transferred to the hatchling, with 25.4% of non-polar lipids and 19.2%

of energy used for embryogenesis. Shells from freshly laid eggs had higher levels of calcium and magnesium
than did shells from hatched eggs. Fully developed embryos could obtain almost all magnesium from the yolkA

but withdraw approximately 35.6% of their total calcium requirements from the eggshell.
few days afterain
decrease
mass
was
an
increaseincarcassmass, indicating that
hatching,
post-hatching yolk
accompanied bypost-hatching yolk could be used to support early growth of hatchlings.

Key Hwd.s.-Reptilia: Squamata: Elaphe taeniura: Incubation; Egg; Post-hatching yolk; Hatchling

Introduction1997). In this paper,

As for reproductive investments in eggs and embryonic development, there are two patterns that seem tobe common in oviparous reptiles. One pattern is thatenergy and material stored in eggs generally
exceed the needs for producing a complete hatchling.
total

Hence, a portion of yolk, namely post-hatching yolkor residual yolk, may remain unutilized at the time ofhatching. Post-hatching yolk represents a supply ofenergy and material for early activities of hatchlings

we present data on a colubrid

snake. Elaphe taeniura. We address the following topics: ( 1 ) conversion of energy and material

from egg tohatchling during incubation, (2) sources of calcium

and magnesium during embryogenesis, and (3) posthatching yolk and its contribution to early growth of

newly emerged hatchlings.

Material and Methods
Elaphe taeniura is one of the most common snakes in(Kraemer and Bennett, 1981; Troyer, 1983, 1987;
Wilhoft, 1986; Congdon and Gibbons, 1989); how-our study areas in the Zhoushan Islands (29° 32'-31°of this portion of resources

The distributional range of E. taeniura covers mostprovinces of China (including Taiwan and Hainan),ever, the exact function

allocated by themothernot very clear.
There is a growing evidence showing that resources in
in

eggs

is

the post-hatching yolk can be transferred to the car-04' N, 121° 30'-123° 25' E), Zhejiang, eastern China.

India (Darjeeling and Assam), Indochina, and thenorthern half of Malay Peninsula (Zhao and Adler,

cass (=total hatchling-yolk sac-fat bodies). The conse-

1993). For this species, many aspects of biology spe-quence of this transference is that carcass increases inmass during the first post-hatching days of hatchlings

cies have(Ji et al.,1997a). The other pattern

is

that

embryosmust mobilize minerals (e.g., calcium) from the eggcomplete development (e.g.. Bustard et al.,
1969: Jenkins, 1975: Packard and Packard, 1984,
1989; Packard et al„ 1984a, b; Shadrix et al„ 1994; Ji

shell toet al„

1996, 1997a,b).

As the consequence of thismobilization, eggshell decreases in mass and ash contents during incubation, particularly at the late stageof incubation (Ji et al., 1996, 1997a, b; Zhao et al„

been previously examined, but little infor-

mation on incubation and reproductionisavailable

other than incidental notes (see Huang and Jin, 1990).

Four gravid E. taeniura [snout-vent length: 1 10.0155.0 cm; body mass (excluding the clutch): 233.4778.7 g] were obtained from a private collector

inBaiquan, Dinghai, the Zhoushan Islands, in mid-June
1994. The snakes were individually maintained in ourlaboratory in 80 x 80 x 80 cm wire cages until oviposition(mean=16.3 days). We removed eggs from the1999

Asiatic Herpetological ResearchVol. 8, p. 54

h of ovicages, measured and weighed them within 6
fromtwo
selected
then
and
eggs
randomly
position,each of the first three clutches and one egg from thelast clutch to determine egg composition. Egg con-stored in a refrigerator until analysis for calcium andmagnesium. Concentrations of the two elements in thedigestates were determined using a WFX-1B model
atomicabsorption

spectrophotometer

(The

2"

tents (embryo plus yolk) of the dissected freshly laid

Beijing Optical Instruments). To check if there were

eggs were removed, placed in pre-weighed small
0.1 mg.
glass dishes, and weighed to the nearest
Shells from the freshly laid eggs were rinsed briefly,

any differences in calcium and magnesium contents
between shells from freshly laid eggs and hatchedfrom each
eggs, we took equal amount of sample

weighed to the nearest o.l mg, and then were saved

shell,

for later analysis. All dissected freshly laid eggs con-laid eggs andtained a small embryo, which was too small and frag-different samples.

ile

to

be

sampledseparately,

and therefore wasincluded with yolk.

pooled separately the samples from the freshly
hatched eggs, and treated them as two

All variables were tested for normality using Kolmog-

orov-Smirnov

test

and for homogeneity of variance

Eighteen eggs, 1/3-buried in moistened substratum, were incubated in a constant temperature chamber at 30±0.3°C. The incubation medium consisted ofand arc-sine transformation was performed for percentage data. We used analyses of variance (ANOVA),sand to water in a ratio of 4: 1 and water was added
content. We meaperiodically to keep the initial water

tics,

,

sured and weighed the incubating eggs at weeklyintervals before day 42. and daily intervals thereafter.

Four eggs

failedtohatchfollowing

incubation.

using Bartlett's test prior to further statistical analysis,

analysis of covariance (ANCOVA), regression
and partial correlation analysis to analyze ourstatis-data. Significance level

was set at a=0.05. Prior tothe homogetesting for differences in adjusted means,neity of slopes

was checked. Throughout this paper,

Hatchlings were measured and weighed immediately
after they left the eggs. Shells from hatched eggs werevalues are presented as mean±l standard error.

rinsed briefly, weighed to the nearest 0.1 mg, and then

Results

were frozen for later analysis. Nine hatchlings (2-3from each clutch; hereafter 0-day hatchling) were froafter hatching. The remaining 5
hatchlings (1-2 from each clutch; hereafter 7-day
hatchling) were fasted at room temperatures (26-

zen immediately

38°C)

for 7 days,

and then frozen. The preserved

hatchlings were later thawed, dissected, and separated
into the carcass, yolk sac. and fat bodies.
All samples for determinations of non-polar lipids,
ash, calories, calcium, andmagnesium were ovendried to constant mass at 65 °C, weighed, and thenground in a mortar and pestle. Non-polar lipids wereextracted from all samples of egg contents, carcass,of
post-hatching yolk, and fat bodies for a minimum
5.5 h using absolute ether in a Soxhlet apparatus. The

mass of non-polar lipids in each sample was calculated as the difference in sample dry mass before andafter extraction.

Ash and calories of samples of egg contents, car-

Elaphe taeniura laid pliable-shelled eggs. Clutch sizein our sample averaged 8.8+0.9 (range=8-ll, N=4).
Freshly laid eggs averaged 26.8±0.6 g (range=20.8-

N=35) wet mass, 54.1±0.9 mm (range=45.7N=35) length, and 28.8±0.4 mm (range=25.033.1, N=35) width. During incubation, eggs increased

32.3,
63.2,

wet mass and, one day prior to hatching, weighed
110.0±3.5% (range=98.6-135.9, N=14) of egg wet
mass at oviposition. The incubation time averaged
54.9±0.2 days (range=54. 1-55.7, N=14). Newlyinemerged young averaged 17.1±0.7 g (range=13.221.8, N=14) wet mass, 381.4±3.5 mm (range=357.0405.0, N=14) SVL, and 88.5±1.9 mm (range=77.0103.0, N=14) tail length.

The data on components of the freshly laid eggs and0-day hatchlings are given in Table 1. Egg contents
averaged 74.0% water by mass; egg contents averaged

92.4% organic material, 7.6% ash, 31.5% non-polar.36% calcium, and 0.39% magnesium by dry
lipid,
mass (Table 1 ). Shells from the freshly laid eggs averaged 17.6% of total egg dry mass, and 81.7% organic
material and 18.3% ash by shell dry mass (Table 1).
1

cass, post-hatching yolk, and fat bodies were determined using a GR-2800 adiabatic bomb calorimeter(Changsha Instruments). Titrations were performed of
the residue after calorimetry to correct for nitrogenous

wastes. Samples of eggshells were burned in a muffle

furnace at 550 °C for 24 h to determine ash mass.

Samples for calcium and magnesium determinations were weighed out into glass tubes and digested

completely in hot concentrated nitric acid. Digestates
were brought to volume in volumetric glassware andShells from freshly laid eggs had higher levels of cal-cium (8.21%) and magnesium (0.75%) than did shells
from hatched eggs (calcium: 6.31%; magnesium:
0.61%).
mass.
0-day hatchlings averaged 70.6% water bymaterial.
89.1%
These hatchlings averaged
organic

Asiatic Herpetological Research

1999

Vol. 8, p. 55

Components and F values of the ANCOVA for 7 Elaphe taeniura freshly laid eggs and nine 0-day
hatchlings. Data are expressed as adjusted meam+1SE with total egg wet mass at oviposition as the covariate.
*"***P<0.001.
P<0.01, andSymbols immediately after F values represent significant levels: NS P>0.05, P<0.05,

Table 1.

Vol. 8, p. 56

Asiatic Herpetological Research

1999

Table 2. A comparison between nine 0-day and five 7-day hatchlings of Elaphe taeniura. Data are expressed as

mean±1 SE; all mass units are in grams.

Hatchling wet mass at hatching

0-day hatchling

7-day hatchling

18.8±0.915.7±0.3

Hatchling wet mass 7 days after hatchling

15.0±0.2

Decrease in wet mass

0.70±0.22

Hatchling dry mass

5.28±0.31

4.00±0.06

Carcass

3.51 ±0.20

3.18±0.05

Yolk sac0.66±0.05

0.09±0.01

Fat bodies

1.11±0.09

0.7310.02

70.6+0.573.2±0.3% water of hatchlingP<0.001 of the 0-day hatchlings were both correlated)

from the substrate on which the eggs were incubated.

with total hatchling dry mass. In the 7-day hatchlings,

However, water uptake seemed not to be obligate for

we only found a positive correlation between carcass

E. taeniura eggs, because some eggs whose final massdry

mass andF, 3=22.34,

total

hatchlingdrymass (r~=0.88,

P<0.05). 7-day hatchlings had signifi-

cantly heavier carcasses than did 0-day hatchlings
with the same wet mass at hatching (ANCOVA:F |=16.38, P<0.01) (Table 2). There was a negativeucorrelation between post-hatching yolk dry mass andcarcass dry mass

when holding total hatchling dry

mass and fatbody dry mass constant (r=-0.90, t=6.53,
df=10, P<0.001 ). There was no significant correlation
between post-hatching yolk dry mass and fatbody dry

mass when holding total hatchling dry mass and car-was less than initial mass also hatched successfully.
Small E. taeniura embryos were present

in allfreshly laid eggs, but they, relative to the large

eggsize, were too small to be considered as an importantpart of the egg at oviposition. Therefore, the transfer-ence of energy and material from egg to hatchling
during incubation was approximately equal to thetransference overall. This makes it possible to compare our data with those for other oviparous reptiles
whose freshly laid eggs also contain small embryos

and embryonic stage is near the oviparous end in thecassoviparity-viviparity continuum (Shine, 1983). Elaphecarcass dry mass and fatbody dry mass when holding

energy and material from egg to hatchling. The values
in Table 3 show that the conversion efficiencies ofdry mass constant (r=0.43, t=1.50, df=10,
P>0.05). There was no significant correlation between

dry mass and post-hatching yolk dry
mass constant (r=-0.20, t=0.65, df=10, P>0.05). 7-day
hatchlings (23.2±0.4%, range=22.1-24.1%, N=5) had
total hatchlingsignificant lower levels of non-polar lipids than did 0-

taeniura exhibitedhigh conversion efficiencies ofenergy and material recorded in E. taeniura werehigher than those reported for any other studied reptiles.

The values in the Table also show that the con-day hatchlings (27.4±0.6%, range=25.3-29.5%, N=9)

version efficiencies vary considerably among species;(F U2 =32.1,P<0.001).

however, the explanations to these differences are
unknown at this time. It has been known that costs of

Discussion

embryonic development vary considerably among
reptiles (Dmf el, 1970; Black et al., 1984), parental

Similar to that reported for pliable-shelled eggs ofother reptiles (e.g.. Fitch, 1954; Fitch and Fitch, 1967;

investment in each offspring should be related to its

Andrews and Sexton, 1981; Vitt and Cooper, 1986;

survivorship (Congdon ad Gibbons, 1989; Fischer etal., 1991), and incubation environments may influ-

Vleck, 1991: Ji et al., 1996, 1997a, b), eggs of E. tae-ence embryonic development (Gutzke and Pachard,niura overall increased in wet mass and swelled dur-

1987). So, further studies in a wider field covering

ing incubation due to a net gain of water absorbed

parental reproductive investment, embryonic metabo-

1999

Asiatic Herpetological ResearchVol. 8, p. 57

Table 3. Comparison of conversion efficiencies of dry mass, non-polar lipids, and energy between Elaphe taeniura

and other oviparous reptiles.

embryos deplete the yolk of almost of its calcium
before hatching and none of the calcium withdrawn
from the eggshell is deposited in the yolk. This patternof mobilization and deposition of calcium is similar toobserved in other non-crocodilian reptiles (e.g.,
Packard et al., 1984b, 1985, 1987; Packard and Pack-

that

ard. 1986: Packard and Packard, 1988: Shadrix et al.,1994).

The estimated amount of magnesium in egg contents of the freshly laidegg (95.9% of total magne-

sium in the hatchling) was slightly less than that in thehatchling.

1999

Asiatic Herpetological ResearchVol. 8, p. 58

We are presently not very certain that E.

taeniura embryos use the eggshell as an additionalsource of magnesium, because any slightly biased
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post-hatching yolk can be used to support early
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The work was supported by a grant from the Natural
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