Fill - Free fillable Valverde Et Al 2010 n12:77 PDF form

ENDANGERED SPECIES RESEARCH

Endang Species Res

Vol. 12: 77–86, 2010

doi: 10.3354/esr00296

Published online June 30

INTRODUCTION

The reproductive fitness of sea turtle females largely

depends on their selection of nesting beach and the

quality and timing of their nesting activities (Miller

1997). During incubation, the nests are affected by bi-

otic factors such as predation (Eckrich & Owens 1995)

and microorganism decomposers (Madden et al. 2008),

and abiotic factors such as temperature and moisture

(Ackerman 1997), density-dependent factors (Honar-

var et al. 2008) and respiratory gas fluxes (Wallace et

al. 2004). Of all these factors, temperature plays a criti-

cal role in embryo development (Miller 1985). When

incubated at a constant temperature, sea turtle embryo

development occurs within a thermal tolerance range

(TTR) of 25–27 to 33–35°C, above or below which em-

bryo development is impaired (Ackerman 1997). Like

crocodiles and many other reptiles, sea turtles exhibit

temperature-dependent sex determination (TSD; Yn-

tema & Mrosovsky 1980, Janzen & Paukstis 1991),

which is characterized by the exclusive production of a

single sex below (male) or above (female) a transitional

range of temperatures (TRT; Mrosovsky 1994, Wibbels

2003). Within the TRT, sex production is variable and

Field lethal incubation temperature of olive

ridley sea turtle Lepidochelys olivacea

embryos at a mass nesting rookery

Roldán A. Valverde

, Susanna Wingard

, Flor Gómez

Mark T. Tordoir

, Carlos M Orrego

Department of Biological Sciences, Southeastern Louisiana University, Hammond, Louisiana 70402, USA

Universidad Nacional San Antonio Abad del Cusco, Cusco, Perú

Department of Environmental Sciences, Van Hall Institute, Leeuwarden, Netherlands

Department of Bioscience and Biotechnology, Drexel University, Philadelphia, Pennsylvania 19104, USA

ABSTRACT: Hatching success in olive ridley mass nesting arribada beaches is typically low. We con-

ducted a short study to understand whether incubation temperature in dry months accounts for

exceedingly low hatching success at Ostional Beach, Costa Rica, a mass nesting rookery. We mea-

sured in situ incubation temperatures in nests from 4 arribadas recorded in October to December

2008, and January 2009. Mean incubation temperatures for all months exceeded the upper lethal

limit of 35°C, and no nests produced hatchlings when incubated at this or higher temperatures.

Embryo development was inversely related to mean incubation temperature. Hatching success was

low (2%) for the study period, and only 5 of 37 marked nests produced hatchlings. Mean incubation

temperature for successful nests was <35°C. Since incubation temperatures of 32°C and higher

recorded during the gonadal thermosensitive period were above the mean pivotal temperature of

30.5°C, the few hatchlings produced were presumably female. Incubation temperatures were signif-

icantly higher during the second and third trimesters of incubation during all months as a result of

metabolic heating. However, during January to March when embryos did not develop, higher incu-

bation temperatures of in situ nests relative to controls indicated that heating was a result of micro-

bial activity associated with egg decomposition. Our study demonstrates that after the onset of the

dry season, incubation temperatures at this beach become lethal.

KEY WORDS: Olive ridley · Lepidochelys olivacea · Ostional · Incubation temperature · Precipitation ·

Thermal tolerance range · Field lethal temperature · Arribada · Mass nesting

Resale or republication not permitted without written consent of the publisher

PEN

CCESS

Endang Species Res 12: 77–86, 2010

relative to incubation temperature and includes a

pivotal temperature (PT) where the sex ratio is 1:1

(Mrosovsky & Pieau 1991). PTs may vary inter- and

intra-specifically, which implies that temperature-

related studies need to be conducted at a population

level (Wibbels 2003). Turtle sex is determined during a

gonadal thermosensitive period (Mrosovsky & Pieau

1991), which occurs in the second trimester of embryo

development (Merchant-Larios et al. 1997). Studies

have shown that the mean PT for olive ridley arribada

populations nesting on Costa Rican Pacific northwest

coast is 30.5°C, the highest among sea turtles (McCoy

et al. 1983, Wibbels et al. 1998). These characteristics

make embryo development under high nest density

conditions vulnerable to changes in environmental

temperature, especially when incubation temperatures

fluctuate around the PT, where a change of 3°C or less

may result in the exclusive production of a single sex

(Ewert et al. 1994, Wibbels 2003). Thus, understanding

the relationship between incubation temperature, de-

velopmental stage and hatchling sex ratios is crucial

for effective sea turtle conservation and management.

Olive ridley sea turtles Lepidochelys olivacea, and to

a lesser extent the Kemp’s ridley L. kempii, are unique

in that in addition to nesting in solitary fashion like

other sea turtles, many females also aggregate and ar-

rive synchronously over a period of a few nights to nest

by the thousands to tens of thousands on a small section

of beach during events known as arribadas (Bernardo &

Plotkin 2007). Large arribadas surpassing 50 000 fe-

males per event are known to occur at Gahirmatha and

Rushikulya in India (Pandav et al. 1994, Shanker et al.

2004), at La Escobilla in Mexico (Márquez-M. et al.

1996), and at Ostional in Costa Rica (Richard & Hughes

1972), with smaller arribadas occurring elsewhere on

Eastern Pacific beaches (Bernardo & Plotkin 2007). The

vast majority, 98 to 99%, of the annual female nesting

population is estimated to lay eggs at these arribada

sites (Abreu-Grobois & Plotkin 2008). Although produc-

tion potential is extremely high at these beaches, the

aggregation of individuals on the beach and near shore

during these events makes populations vulnerable to

anthropogenic threats such as egg poaching (Cornelius

et al. 2007) and incidental capture of adults and juve-

niles by fisheries (Arauz et al. 1998, 1999, Frazier et al.

2007). In addition, developing turtle eggs at these

beaches are also subject to high natural mortality due

to density-dependent factors (Bernardo & Plotkin 2007).

This has both ecological and conservation implications

concerning the importance of arribada beaches for the

long term survival of this species, currently listed as

vulnerable by the IUCN Red List (Abreu-Grobois &

Plotkin 2008).

High nest densities alone at arribada beaches are cor-

related with low hatching success (Honarvar et al.

2008). Additionally, natural embryo mortality is high

during arribadas due to excavation of previously laid

nests by later nesting females (Cornelius et al. 1991). It

is this high loss of eggs that led Costa Rican authorities

to legitimize a large-scale egg extraction and market-

ing program in which Ostional villagers collect as many

eggs as possible during the first 36 h of each arribada to

sell in the domestic market (Campbell 1998, Hope 2002,

Campbell et al. 2007). Biologically, the large number of

broken eggs fosters microbial populations on the

beach, which use oxygen, in addition to that being con-

sumed by the many developing embryos (Madden et al.

2008). PO

is lower and incubation temperatures are

higher in high density nests compared to low density

nests as a result of microbial and embryo respiration

and metabolic heating (Clusella-Trullas & Paladino

2007, Honarvar et al. 2008). The extent of the influence

of metabolic (embryonic and microbial) activity on in-

cubation temperatures in areas with high nest density

is poorly understood. However, mass mortality of ridley

embryos during the dry, hot season has been suspected

for years at Ostional (Cornelius et al. 1991). Accord-

ingly, our main objective was to determine whether dry

season incubation temperatures are lethal to olive rid-

ley embryos at Ostional Beach, a mass nesting beach.

MATERIALS AND METHODS

Study site. Field work was conducted at Ostional

Beach (9° 59’ 46’’ N, 85° 42’ 13’’ W), Costa Rica (Fig. 1)

from late October 2008 to early March 2009. This dark

Fig. 1. Ostional Beach, Costa Rica. Sections (Rayo 1–4 and

main nesting beach [MNB]) in which the beach was divided

to facilitate the work. Typically, large arribadas occur at

MNB, although in large events as in 2008, the phenomenon

may be spread over the entire beach

Valverde et al.: Lethal incubation temperature of ridley sea turtle embryos

sand beach is located within the Ostional Wildlife

Refuge and measures 3.9 km in length. The region ex-

hibits fairly well marked climate conditions with a rainy

season that normally extends from May to November

and a dry season that extends from December to April.

The beach is divided into 5 sections and each section is

divided into subsections marked with posts every 50 m,

which are located on the vegetation line. These sections

are Rayo 1 (posts 1–17), Rayo 2 (posts 18–29), Rayo 3

(posts 30–40), Rayo 4 (posts 41–59) and main nesting

beach (MNB; posts 60–78). Arribadas tend to concen-

trate on the main nesting beach. Only during large ar-

ribadas do nesting females crowd the beach to such an

extent that the arribada encompasses most of the avail-

able beach sections. Arribadas peak in the rainy season

(August to December), with the September, October

and November arribadas being usually the largest, sig-

nificantly decreasing in abundance the rest of the year

(Cornelius 1986). Ostional Beach typically supports on

average 11 arribadas each year and ranks second be-

hind La Escobilla among the largest arribada assem-

blages, with between 50 000 and 200 000 nesting fe-

males per arribada (Plotkin 2007). Using the strip

transect in time method (Valverde & Gates 1999), arrib-

adas in 2008 were estimated to fluctuate between an

average of 117 996 ± 9395 females (mean ± SE; Septem-

ber to November; n = 2) and an average of 8101 ± 3573

(January to July; n = 9) (R. A. Valverde unpubl.).

Precipitation and nest and sand temperatures. Daily

precipitation (mm) was recorded every morning

(07:00 h) with a precipitation gauge throughout the study

period. In situ nests were selected during the arribadas

of October 28–30, November 21–23, and December

21–23 of 2008 and during January 22–23, 2009. Nest

selection was conducted by choosing any female laying

eggs in the open beach between the high tide mark and

the vegetation line, which is the area of the beach most

used by olive ridleys. Newly purchased HOBO pendant

temperature data loggers (Onset Computer, UA-001-08,

accuracy: ±0.5°C, resolution: 0.1°C, according to manu-

facturer) were inserted into the center of each nest with-

out disturbing the egg-laying process. Once nesting fe-

males began to fill the nest cavity, the turtles were

removed so the nest could be covered and protected

from subsequent nesters. Each nest was covered with a

40 × 40 × 25 cm wire mesh cage. From Day 40 of the incu-

bation period, each nest was monitored daily for signs of

neonate emergence and cages were either lifted or

removed upon sighting of a hatchling. Nests were ex-

humed 3 d after the last observed neonate emerged,

~50 d from oviposition. No live embryos were observed

in the nests during exhumation.

Control data loggers were buried in the sand (>1 m

distance from nests) and set to record sand tempera-

ture throughout the entire incubation period of the

respective arribada nests at a depth equal to that of the

center of an average nest (30 to 35 cm). All data loggers

were programmed to collect temperature data every

2 h. Recording consistency and accuracy of data log-

gers were ensured by placing all sensors in a bag

simultaneously and keeping them in a shaded room to

record air temperature for a period of 38 h (recordings

included 2 night periods).

Nest development and hatching success. Upon ex-

humation, data loggers were retrieved and the develop-

mental stage in each egg was recorded for all nests. Eggs

were classified into developmental stages according to

Chacón et al. (2007): Stage 0 (no apparent embryonic de-

velopment), Stage I (embryo fills 1 to 25% of amniotic

cavity), Stage II (26–50%), Stage III (51–75%), Stage IV

(76–100%), Stage V (empty egg shells from hatched tur-

tles). Eggs with indeterminable development were clas-

sified as ‘unknown stage of development’ (USD). Given

the relatively few marked nests with data loggers, we

decided to increase the sample size of nests in the study

by excavating additional 1 m

plots (10 m due north or

south of each study nest) concurrent with exhumations of

the study nests from October and November 2008. The

number of nests and each egg’s developmental stage

was recorded for each 1 m

plot.

Data analysis. Data from nest and control data log-

gers were downloaded and converted to spreadsheets

using HOBOware Pro 2.7.3. Temperature recordings of

data loggers were checked for consistency by testing

the hypothesis that the individual readings were no

different from the mean by running a χ

test for each of

the time points over a 38 h period (Fig. 2). No signifi-

cant differences were found.

Time (h)

Temperature (°C)

15:00

19:00

23:00

03:00

07:00

11:00

15:00

19:00

23:00

03:00

07:00

Fig. 2. Mean (±SE) air temperature and upper and lower con-

fidence intervals (95%) observed in all data loggers (n = 21)

kept in the shade for 38 h. Black bars on x-axis: scotophase.

Individual logger temperatures were not significantly differ-

ent from their respective mean (χ

= 45.31, df = 20, p < 0.001)

Endang Species Res 12: 77–86, 2010

Daily mean incubation temperatures were calcu-

lated for the incubation period for all nests and con-

trols. Monthly and trimester mean incubation tempera-

tures were also calculated. Hatching success was

calculated for each nest as: (empty egg shells)/(total

number of eggs) × 100%. Data analysis was conducted

with Systat 10.2 using a 2-sample t-test, 2-way

ANOVA, and regression analyses. For all tests,

α = 0.05. After completing a preliminary analysis, we

reviewed our data and found no outliers. We tested the

residuals for normality using a 1-sample KS test with

Lilliefor’s option and homogeneity of variance, and fit a

model by analyzing the plot of residuals against pre-

dicted values.

RESULTS

Precipitation

In general, monthly average precipitation was 3.69,

2.13, 0.08, 0.0, 0.0, 0.0, 0.10 mm d

–1

for the months of

October to April, respectively. Specifically, precipita-

tion throughout the incubation period (October 29 to

March 8) was low on average with 1.25 mm d

–1

. Of the

131 d in the study period, 118 d (90%) were without

precipitation and total precipitation for that period was

158.10 mm. Rainfall, when it occurred, only exceeded

3 mm on 3 occasions in the entire study period:

November 9 (52 mm) and 30 (57 mm), and December 1

(33 mm). Overall, the dry season started a month early

with November precipitation being lower than its

mean precipitation for the period 1995–2007 (Fig. 3).

This precipitation and associated cloudiness led to rel-

atively abrupt and brief decreases in mean incubation

temperature of 1.5, 2 and 1.3°C, respectively (Fig. 4).

No historical records for air temperature were avail-

able for the area.

Nest temperatures

A total of 42 data loggers were deployed in arribada

nests during the study. Loggers were placed in nests

distributed throughout the beach (Rayo 1 to MNB)

between the high tide and away from vegetation and

estuaries, which represents the bulk of the incubation

environment in Ostional. Due to logistic problems, we

were able to record incubation temperatures from only

37 nests. Excluded from analyses were data loggers

that could not be relocated (December, n = 2), prema-

turely stopped recording, did not record any data

(November, n = 2), or were excavated by raccoons

(January, n = 1). Daily mean (±SE) incubation temper-

atures are shown in Fig. 4. The mean incubation tem-

perature for October nests was 35.90°C (mean mini-

mum and maximum: 34.17 and 37.66°C, respectively;

n = 8), for November was 35.67°C (32.99–38.16°C; n =

19), for December was 35.39°C (34.23–37.06°C; n = 6),

and for January nests it was 37.58°C (36.85–38.26°C;

n = 4). Incubation period lasted about 47 d, which is

when the last hatchling was observed.

The total range in recorded temperatures for the

entire study period was 17.49°C (min. = 25.03°C,

Time (mo)

Jun DecAug Oct Feb Apr

Precipitation (mm)

200

400

600

800

1000

2008-09

1995-2007

Fig. 3. Mean (±SE) total monthly precipitation recorded at

Nosara Beach (adjacent to study site; gray bars) for 1995–

2007 and total precipitation at Ostional Beach (dark bars)

for 2008–2009. Data shows the dry season (Dec–Mar) and

rainy season (May–Nov). November 2008 (

) was drier than

average

Incubation day

010203040

Mean incubation temperature (°C)

Oct (n = 8)

Nov (n = 19)

Dec (n = 6)

Jan (n = 4)

52 mm

57 mm

FLT

Fig. 4. Mean (±SE) daily incubation temperatures for nests

laid during arribadas of Oct, Nov, and Dec 2008, and Jan

2009. Days with precipitation >3 mm are annotated. PT =

mean pivotal temperature; FLT = field lethal temperature

Valverde et al.: Lethal incubation temperature of ridley sea turtle embryos

max. = 42.52°C; both temperatures recorded in Octo-

ber nesters). Mean daily nest temperatures ranged

from 28.41°C in November to 40.14°C in January.

Incubation temperature was not related to the beach

sector where nests were laid, as there was no signifi-

cant relationship between mean incubation tempera-

ture and nest location by beach section. Study nests

in adjacent sections of beach exhibited mean temper-

atures that differed by 0.30 to 2.68°C, indicating that

the thermal microenvironment was heterogeneous.

Mean incubation temperatures in nests laid in Janu-

ary were significantly higher than in all other

months, 37.58°C compared to 35.68°C, respectively

(F = 6.02 p = 0.00083, Fig. 5B). Ranges in mean tem-

peratures between clutches was 5.17°C for nests laid

during the November arribada, the month with the

greatest inter-nest temperature variability as well as

lowest mean incubation temperature in a nest

(32.99°C). Mean incubation temperatures were most

uniform among nests laid during the January arrib-

ada (range of 1.42°C). One January nest had the

highest mean incubation temperature of 38.26°C.

Nest temperatures generally increased throughout

the incubation period. Mean incubation temperatures

for the second (36.11°C) and third (36.97°C) tri-

mesters were significantly higher than in the first

(34.57°C, F = 9.75, p = 0.00014; Fig. 5A). Incubation

temperatures <35°C, the upper limit of the TTR

(Ackerman 1997), only occurred in the first trimesters

of nests laid in November (33.74°C) and December

(34.70°C). Though no month had mean temperatures

below the upper lethal limit of 35°C, several nests

laid in October (n = 2), November (n = 5), and De-

cember (n = 3) had mean temperatures <35°C. Mean

daily temperatures exceeded 35°C for an average

69% of the incubation period of all nests (30.95 d,

95% confidence interval [CI

95%

] 26.78 to 35.12 d;

means and SE are reported in figures to increase fig-

ure clarity), and mean temperatures for all nests laid

in January exceeded 35°C throughout the entire

incubation period.

Control temperatures

Temperatures in controls (Fig. 6) increased signifi-

cantly toward the end of the study period. Mean tem-

peratures in January controls (36.14°C, CI

95%

34.50 to

37.27°C) were significantly higher than those in

November (34.09°C, CI

95%

32.63 to 35.55°C; 2-sample

t-test, t = 2.85, p = 0.017).

Nest versus control temperatures

Mean incubation temperatures from nests laid in

November (35.67°C, CI

95%

34.91 to 36.43°C) were

significantly higher than their respective controls

(34.10°C, CI

95%

32.63 to 35.55°C, t = 2.193, p = 0.039).

Temperatures within nests were on average 1.57°C

warmer than those in controls. The difference in-

creased with each trimester, with nests being 0.66,

1.76, and 2.23°C warmer than controls for the first to

third trimesters, respectively (Fig. 6A). There was a

significant difference between mean temperatures

in controls (36.14°C, CI

95%

35.00–37.27°C) and in nests

(37. 58°C, CI

95%

36.45–38.70°C) laid in January

(Fig. 6B; t = 2.320, p = 0.049). January nests were on

average 1.47°C warmer than controls, though they did

not become increasingly warmer throughout the incu-

bation period (1.32, 1.26, and 1.79°C, for the first to

third trimesters, respectively).

Incubation trimester

1st 2nd 3rd

Mean incubation temperature (°C)

Time of oviposition (month)

Oct Nov Dec Jan

FLT FLT

Fig. 5. Mean (±SE) incubation temperature per (A) trimester and (B) month of oviposition. Mean temperatures were significantly

higher during 2nd and 3rd trimesters and during the entire incubation period for nests laid in January (pairwise comparison,

Bonferroni adjusted means). Lowercase letters above bars indicate significant differences at 95% level of confidence. See Fig. 4

for definitions

Endang Species Res 12: 77–86, 2010

Embryo development and hatching success in study

nests

Of the 37 nests with usable data loggers, 16 nests ex-

hibited no development. Hatching rates for study nests

were 2.14% (19 of 889 eggs) for October, 2.54% (44 of

1646 eggs) for November, 1.72% (10 of 580 eggs) for

December, and 0% (0 of 439) for January, for an overall

hatching rate of 2% (73 hatchlings out of 3554 eggs).

In order to study the effect of incubation temperature

on embryo development, we staged the embryos from

all nests with data loggers. Given the large number of

eggs in each plot, we used a simple staging table in

which embryos were divided in the 5 stages described

above according to Chacón et al. (2007). These stages

corresponded roughly with those of Crastz (1982) as

follows: Stage I: Stages 1–10 (appearance of eye con-

sidered Stage II); Stage II: Stages 11–20 (appearance of

pigmentation considered Stage III); Stage III: Stages

21–28 (onset of vitelline sac reabsorption considered

Stage IV); Stage IV: up to hatching.

Mean incubation temperatures were significantly

lower in nests with development compared to nests

without development, i.e. 34.97 (CI

95%

34.47–35.46°C)

and 37.08°C (CI

95%

36.59–37.58°C), respectively (t =

6.27, p ≤ 0.0001). The highest developmental stage

reached within a nest was inversely related to mean in-

cubation temperature (regression analysis: r

= 0.706,

p ≤ 0.0001, Fig. 7). The highest developmental stage

reached was also inversely proportional to the total

number of days with incubation temperatures >35°C

(regression analysis: r

= 0.804, p ≤ 0.0001; Table 1).

Mean incubation temperatures were also signifi-

cantly lower within the 5 nests with hatchlings

(33.84°C, CI

95%

33.24–34.44°C; Stage 5) compared to

nests without (36.20°C, CI

95%

35.73–36.67°C; Stages

0–4; t = 3.98, p ≤ 0.0003). The rate of 14.81% hatching

success was low within nests with hatchlings (Table 1).

The nest with the highest percent emergence of

34.29% (n = 36 hatchlings from 105 eggs) had the

lowest mean incubation temperature.

Embryo development and hatching success in

density plots

A total of 8697 eggs were exhumed from 29 × 1 m

plots. Clutch density was too high to tell individual

nests apart, which is similar to what was found previ-

ously (Cornelius et al. 1991). For this reason we

counted all eggs within each plot. Thus, 4183 eggs

from 8 plots from the October arribada yielded a den-

sity of 522.88 eggs m

–2

, and 4514 eggs from 21 plots

Incubation day

0 10203040 10 203040

Mean incubation temperature (°C)

November nests (n = 19)

November controls (n = 6)

January nests (n = 4)

January controls (n = 6)

FLT

Fig. 6. Mean (±SE) incubation temperatures of nests and controls from (A) November and (B) January. See Fig. 4 for definitions

Mean incubation temperature (°C)

32 33 34 35 36 37 38 39

Highest developmental stage reached

III

Fig. 7. Last developmental stage reached per nest in relation

to mean incubation temperature. Solid line: linear trend.

Vertical dashed line: field lethal temperature of 35°C. Stage 0:

no embryonic development reached; Stage I: 1–25% egg vol-

ume occupied by developing embryo; Stage II: 26–50%;

Stage III: 51–75%, Stage IV: 76–100%; Stage V: hatchlings

Valverde et al.: Lethal incubation temperature of ridley sea turtle embryos

from the November arribada yielded a density of

214.95 eggs m

–2

. Taking the average clutch size as 100

eggs (CI

95%

94.55–105.45; n = 41 nests with data log-

gers), the nest densities for October and November

plots were 5.23 and 2.15 clutches m

–2

, respectively.

There was no embryo development in most nests

(92.83 and 88.99% of eggs lacked any visible develop-

ment in October and November, respectively). Within

nests that had embryonic development, a significantly

greater percentage of developing eggs from Novem-

ber nests resulted in hatchlings (97.79% or 486 hatch-

lings; t = 2.1161, p = 0.375) compared to those from

October nests (21.67% or 65 hatchlings). Likewise, the

hatching success rate for November nests was signifi-

cantly higher (10.77%, CI

95%

7.17–30.32; t = 2.8196,

p = 0.006) than for October nests (1.55%, CI

95%

0.94–4.53). Mean hatching rate for both months com-

bined was 6.34%.

DISCUSSION

Most studies of incubation temperature, sea turtle

embryo development and TSD are concerned with the

potential feminization of sea turtle populations in the

face of global warming temperatures. The reason for

this is that elevated temperatures tend to promote

skewed sex ratios toward a female bias (Wibbels 2003,

2007), which may curtail the reproductive potential of

a population by decreasing the number of male turtles

(Fuentes et al. 2010). However, our data indicate that

an issue of greater concern at Ostional Beach is that

incubation temperatures measured in the dry season

are not only on the high end of the TRT, above PT, but

are near the upper limit of the TTR and surpass lethal

temperatures, leading to developmental arrest and to

the death of olive ridley embryos. Indeed, our data are

consistent with a field lethal incubation temperature of

~35°C when most nests yielded embryos with little or

no appreciable development. This effect of tempera-

ture was most conspicuous in January nests when

development was negligible and no hatchlings were

produced. This field lethal incubation temperature is

consistent with the lethal temperature determined

under laboratory conditions for sea turtle embryos

when eggs are incubated at a constant temperature

(Ackerman 1997). Because our study focused only on a

part of the year when incubation temperatures were

significantly elevated, we cannot conclude that the his-

torically low hatching rates documented at Ostional

are due to lethal incubation temperatures. However,

our study supports the claims of the Ostional commu-

nity that during the dry season hatchling production is

nearly zero (Cornelius et al. 1991). Our study provides

empirical evidence that this lack of hatchling produc-

tion during the dry months is due to the lethal effect of

temperature.

Although both humidity and temperature constitute

powerful regulators of embryonic development (Miller

1985), we believe that temperature plays a more signif-

icant role in this study. The reason is that lethal tem-

peratures from laboratory studies are derived from

clutches incubated in hydric environments that pro-

mote optimal embryo development (e.g. McCoy et al.

1983, Wibbels et al. 1998). Under these conditions,

high embryo mortality is observed when constant incu-

bation temperatures extend beyond TTR (Yntema &

Mrosovsky 1980, Ackerman 1997).

Interestingly, incubation temperature seems to

exhibit a tonic effect on embryo mortality and develop-

ment on the high end of the TTR given that it is the

time spent at high temperatures and above 35°C that

determines embryo arrest and death and not the mere

fact of reaching this temperature. This indicates that

embryos have physiological mechanisms that allow

them to withstand exceedingly high temperatures, and

that these mechanisms will fail if temperatures do not

decrease below the lethal limit. Presumably, cooling

periods (e.g. rain) may rescue embryos at risk of over-

heating. This observation supports the idea of develop-

ing artificial ways to cool down incubating nests dur-

ing the dry months by providing them with irrigation

or shading (Cornelius et al. 1991) to increase hatchling

production during those months, if desirable.

Month Mean Nests with hatchlings Clutch Hatching

inc

(°C) D 1st D 2nd D 3rd D Hatchlings size success (%)

October 34.17 10 33.76 0 33.94 2 34.81 8 19 100 19.00

November 33.71 15 30.60 0 33.41 0 37.13 15 3 81 3.70

November 33.85 16 31.44 0 34.06 1 36.05 15 5 94 5.32

November 33.12 7 30.86 0 33.46 0 35.02 7 36 105 34.29

December 34.37 17 33.32 0 34.19 2 35.61 15 10 85 11.76

Average 33.84 13 32.00 0 33.81 1 35.72 12 14.6 93.00 14.81

Table 1. Mean incubation temperatures per month (T

inc

) and per trimester (1st, 2nd, 3rd), number of hatchlings, clutch size,

and percent hatching success of the 5 nests that yielded emerging hatchlings. D = number of days for which mean incubation

temperature >35°C for successful nests

Endang Species Res 12: 77–86, 2010

Olive ridleys, like other sea turtle species, are known

to nest year round in Costa Rica. Indeed, small arrib-

adas and solitary nesting have been documented for

the olive ridley on the Pacific coast every month of the

year, with the larger arribadas occurring between

August and November (Hughes & Richard 1974, Cor-

nelius 1986). Although far less abundant, other turtle

species such as greens and leatherbacks also exhibit

year round nesting, with a discrete peak reproductive

window including the months between October and

March (Cornelius 1986, Reina et al. 2002). These

months coincide with some of the hottest, driest

months of the year. This presumes a similar scenario

for these species as in our study with the consequent

detrimental impact on embryo development. However,

both leatherbacks and greens exhibit strategies that

allow them to nest during the most thermally challeng-

ing months of the year. For instance, although

leatherbacks prefer to nest in the open beach, they do

so at a greater depth than ridleys (Binckley et al. 1998),

whereas greens prefer to nest in the vegetation where

temperatures are lower (Cornelius 1976, 1986), thus

escaping the detrimental effect of the high tempera-

tures. In contrast, ridleys prefer to nest in the open

beach (Hinestroza & Páez 2001) and the top of their

nests is at an average 22 cm from the surface (Vega &

Robles 2005), which makes their nests more suscepti-

ble to the high temperatures of the dry season. As

such, the high embryo mortality of ridley embryos dur-

ing the dry months explains the selection pressure of

this species to time the bulk of their reproductive effort

to coincide with the rainy season when lower incuba-

tion temperatures are more favorable for development.

Rainfall was virtually absent during our study with

only 2 brief periods of significant rain, both recorded in

early and late November. We observed a decrease in in-

cubation temperature associated with these periods of

rain. This cooling effect of rain has been recognized in

many studies (Leslie et al. 1966, Godfrey et al. 1996,

Binckley et al. 1998, Matsuzawa et al. 2002, Houghton et

al. 2007). Thus, the lack of rainfall likely led to the ex-

ceedingly high incubation temperatures recorded dur-

ing the study period, which reached their zenith during

January and February when nests incubated at mean

temperatures >36°C and embryo development was cur-

tailed. This situation was expected to continue

throughout the rest of the dry season. Would hatching

rates recover to match those of solitary nesting rates of

59% and higher for olive ridley nests (Francia 2004,

Lopez-Castro et al. 2004) during the next rainy season

when incubation temperatures were expected to de-

crease? We believe that this is unlikely. Although our

data supports a preponderant effect of temperature dri-

ving embryo mortality at Ostional, hatching rates at

Costa Rican arribada beaches have been historically low

(Cornelius et al. 1991, Valverde et al. 1998, Fonseca et al.

2009), even during the rainy months. Factors such as

density-dependent effects and high microorganism load

due to decomposing organic matter and associated oxy-

gen depletion in the nest (Valverde et al. 1998, Clusella-

Trullas & Paladino 2007, Honarvar et al. 2008, Fonseca et

al. 2009) are thought to be involved in the embryonic

death in Costa Rican arribada beaches during the rainy

months. Thus, our study highlights the importance of

continuing research focusing on the mechanisms that

drive embryo mortality under various rainfall, tempera-

ture and nest density conditions and over seasons to

verify whether our results are typical of these beaches.

The large differences between cooler controls and

warmer study nests in the present study indicate that

metabolic heating occurred during the incubation peri-

ods for nests laid in November and January. November

arribada nests exhibited metabolic heating that was

primarily of embryological origin. Embryonic meta-

bolic heating occurs during the second half of embryo

development, especially during the last trimester

(Maxwell et al. 1988, Maloney et al. 1990, Godfrey et

al. 1997, Broderick et al. 2001). Thus, temperature dif-

ferences between controls and nests increase with the

progression of the incubation period, as observed in

December when November nests incubated. However,

in the case of January, nests embryo development was

minimal while incubation temperatures were higher in

nests than in controls. Adult and larval insects, fungi,

and microorganisms have been associated with dead

turtle eggs and dead hatchlings at Ostional (Madden et

al. 2008), and large quantities of these organisms may

contribute to metabolic heating. Thus, we feel that the

difference in temperatures between nests and controls

in January and February is most likely due to micro-

bial, or decomposer, metabolic heating.

Lastly, our exhumation plots yielded additional em-

pirical embryo development data and support our find-

ings of low development and hatching rates from

marked nests. The higher hatching rates in exhumation

plots from November could be attributable to a lower

nest density than October nests as it has been reported

that density-dependent effects may impair embryo de-

velopment (Honarvar et al. 2008). Thus, it is tempting to

suggest that, under in situ conditions at Ostional, den-

sity-dependent effects that limit embryo development

manifest at nest densities >2 nests m

–2

, though this can-

not be determined from our study due to the overriding

effect of the high incubation temperatures.

SUMMARY

Our results are consistent with a field lethal incuba-

tion temperature of 35°C for olive ridley embryos. Our

Valverde et al.: Lethal incubation temperature of ridley sea turtle embryos

data suggest that at Ostional, high incubation temper-

ature is the primary factor limiting embryo develop-

ment during the dry season. Our results indicate that

nests laid during arribadas are vulnerable to short term

weather fluctuations, especially droughts. Without the

rain’s cooling effect, metabolic heating within densely

spaced nests from arribadas may push the already high

incubation temperatures towards and beyond the

upper limit of the TTR, thus disrupting or terminating

embryo development or producing single sex clutches.

The findings of this study provide a glimpse of the

effects of the expected global warming temperatures

on sea turtle populations around the world. It has been

forecasted that global temperatures may increase by

up to 3.5°C or more by the year 2100 (IPCC 2007).

According to our data, not only is feminization of sea

turtle populations possible, but the increase in embryo

mortality due to lethal field temperatures may be a

reality for some populations. Ostional Beach, and per-

haps other arribada beaches in the Eastern Pacific,

seem particularly poised to suffer the detrimental

effects of global warming. Finally, it is important to

emphasize that this study was conducted during the

dry season, a time characterized by high temperatures,

low precipitation and lower nesting relative to the

peak nesting months. Extension of this work to the

rainy months is essential to obtain an improved under-

standing of the mechanisms leading to the high

embryo mortality at arribada beaches.

Acknowledgements. We thank L. S. Brenes (Administrator of

the Ostional National Wildlife Refuge), the Tempisque Con-

servation Area, and the Costa Rican Ministry of the Environ-

ment and Energy for their support and for providing the per-

mits to conduct this research. We also thank the people of

Ostional for their assistance and support during the field

work. L. G. Fonseca and 3 anonymous reviewers offered valu-

able criticism to this manuscript. D. Solís kindly generated

Fig. 1. Finally, we thank the many volunteers who assisted

with data collection. This research was funded by the Marine

Turtle Conservation Program of the USFWS (98210-8-G515).

This study was conducted under Southeastern’s IACUC per-

mit #004.

LITERATURE CITED

Abreu-Grobois A, Plotkin PT (2008) Lepidochelys olivacea.

In: IUCN 2009. IUCN Red List of Threatened Species. Ver-

sion 2009.1, available at www.iucnredlist.org

Ackerman RA (1997) The nest environment and embryonic

development of sea turtle. In: Lutz PL, Musick JA (eds) The

biology of sea turtles. CRC Press, Boca Raton, FL, p 83–106

Arauz RM, Vargas R, Naranjo I, Gamboa C (1998) Analysis of

the incidental capture and mortality of sea turtles in the

shrimp fleet of Pacific Costa Rica. In: Epperly SP, Braun J

(compilers) Proc 17th Annu Sea Turtle Symp. NOAA Tech

Memo NMFS-SEFSC-415, p 1–5

Arauz R, Rodriguez O, Vargas R, Segura A (1999) Incidental

capture of sea turtles by Costa Rica’s longline fleet. In:

Kalb HJ, Wibbels T (compilers) Proc 19th Annu Symp Sea

Turtle Biol Conserv. NOAA Tech Memo NMFS-SEFSC-

443, p 62–64

Bernardo J, Plotkin PT (2007) An evolutionary perspective on

the arribada phenomenon and reproductive behavioral

polymorphism of olive ridley sea turtles (Lepidochelys oli-

vacea). In: Plotkin PT (ed) Biology and conservation of rid-

leys sea turtles. Johns Hopkins University Press, Balti-

more, MD, p 59–87

Binckley CA, Spotila JR, Wilson KS, Paladino FV (1998) Sex

determination and sex ratios of Pacific leatherback turtles,

Dermochelysc oriacea. Copeia 1998:291–300

Broderick AC, Godley BJ, Hays GC (2001) Metabolic heating

and the prediction of sex ratios for green turtles (Chelonia

mydas). Physiol Biochem Zool 74:161–170

Campbell LM (1998) Use them or lose them? Conservation

and the consumptive use of marine turtle eggs at Ostional,

Costa Rica. Environ Conserv 25:305–319

Campbell LM, Haalboom BJ, Trow J (2007) Sustainability of

community-based conservation: sea turtle egg harvesting

in Ostional (Costa Rica) ten years later. Environ Conserv

34:1–10

Chacón D, Sánchez J, Calvo JJ, Ash J (2007) Manual para el

manejo y conservación de las tortugas marinas en Costa

Rica con énfasis en la opercaión de proyectos en playa y

viveros. Sistema Nacional de Areas de Conservación,

Ministerio de Ambiente y Energía, San José

Clusella-Trullas S, Paladino FV (2007) Micro-environment of

olive ridley turtle nests deposited during an aggregated

nesting event. J Zool 272:367–376

Cornelius SE (1976) Marine turtle nesting activity at Playa

Naranjo, Costa Rica. Brenesia 8:1–27

Cornelius SE (1986) The sea turtles of Santa Rosa National

Park. Fundación de Parques Nacionales, Madrid

Cornelius SE, Ulloa MA, Castro JC, Mata del Valle M, Robin-

son DC (1991) Management of olive ridley sea turtles

(Lepidochelys olivacea) nesting at Playas Nancite and

Ostional, Costa Rica. In: Robinson JG, Redford KH (eds)

Neotropical widlife use and conservation. University of

Chicago Press, Chicago, IL, p 111–135

Cornelius SE, Arauz R, Fretey J, Godfrey MH, Marquez-M. R,

Shanker K (2007) Effect of land based harvest of Lepi-

dochelys. In: Plotkin PT (ed) Biology and conservation of

ridley sea turtles. Johns Hopkins University Press, Balti-

more, MD, p 231–251

Crastz F (1982) Embryological stages of the marine turtle

Lepidochelys olivacea (Eschscholtz). Rev Biol Trop 30:

113–120

Eckrich CE, Owens DW (1995) Solitary versus arribadas nest-

ing in the olive ridley sea turtles (Lepidochelys olivacea): a

test of the predator-satiation hypothesis. Herpetologica 51:

349–354

Ewert MA, Jackson DR, Nelson CE (1994) Patterns of temper-

ature-dependent sex determination in turtles. J Exp Zool

270:3–15

Fonseca LG, Murillo GA, Guadamúz L, Spínola RM, Valverde

RA (2009) Downward but stable trend in the abundance

of arribada olive ridley (Lepidochelys olivacea) sea turtles

at Nancite Beach, Costa Rica for the period 1971–2007.

Chelonian Conserv Biol 8:19–27

Francia AG (2004) Predación y eclosión en nidos solitarios y

de arribadas de tortugas loras (Lepidochelys olivacea),

baulas (Dermochelys coriacea) y negras (Chelonia mydas

agassizi) en las playas Nancite y Naranjo e incidencia

humana sobre las anidaciones de tortugas marinas en

playa Junquillal, Costa Rica. Programa Regional en

Manejo de Vida Silvestre para Mesoamérica y el Caribe.

MSc thesis, Universidad Nacional, Heredia

➤

➤➤

➤

Endang Species Res 12: 77–86, 2010

Frazier J, Arauz R, Chevalier J, Formia A and others (2007)

Human-turtle interactions at sea. In: Plotkin PT (ed) Biol-

ogy and conservation of ridley sea turtles. Johns Hopkins

University Press, Baltimore, MD, p 253–296

Fuentes MMPB, Hamann M, Limpus CJ (2010) Past, current

and future thermal profiles of green turtle nesting

grounds: implications from climate change. J Exp Mar Biol

Ecol 383:56–64

Godfrey MH, Barreto R, Mrosovsky N (1996) Estimating past

and present sex ratios of sea turtles in Suriname. Can J

Zool 74:267–277

Godfrey MH, Barreto R, Mrosovsky N (1997) Metabolically

generated heat in sea turtles nests and its potential effect

on the sex ratio of hatchlings. J Herpetol 31:616–619

Hinestroza LM, Páez VP (2001) Anidación y manejo de la tor-

tuga golfina (Lepidochelys olivacea) en la Playa La Cue-

vita, Bahía Solano, Chocó, Colombia. Cuad Herpetol 14:

131–144

Honarvar S, O’Connor MP, Spotila JR (2008) Density-depen-

dent effects on hatching success of the olive ridley turtle,

Lepidochelys olivacea. Oecologia 157:221–230

Hope RA (2002) Wildlife harvesting, conservation and

poverty: the economics of olive ridley egg exploitation.

Environ Conserv 29:375–384

Houghton JDR, Myers AE, Lloyd C, King RS, Isaacs C, Hays

GC (2007) Protracted rainfall decreases temperature

within leatherback turtle (Dermochelys coriacea) clutches

in Grenada, West Indies: ecological implications for a spe-

cies displaying temperature dependent sex determination.

J Exp Mar Biol Ecol 345:71–77

Hughes DA, Richard JD (1974) The nesting of the Pacific rid-

ley turtle Lepidochelys olivacea on Playa Nancite, Costa

Rica. Mar Biol 24:97–107

IPCC (Intergovernmental Panel on Climate Change) (2007)

Climate change 2007: summary for policy makers. Synthe-

sis Report. IPPC, Valencia, p 22

Janzen FJ, Paukstis GL (1991) Environmental sex determina-

tion in reptiles: ecology, evolution, and experimental

design. Q Rev Biol 66:149–179

Leslie AJ, Penick DN, Spotila JR, Paladino FV (1966)

Leatherback turtle, Dermochelys coriacea, nesting and

nest success at Tortuguero, Costa Rica, in 1990–1991.

Chelonian Conserv Biol 2:159–168

Lopez-Castro MC, Carmona R, Nichols WJ (2004) Nesting

characteristics of the olive ridley turtle (Lepidochelys oli-

vacea) in Cabo Pulmo, southern Baja California. Mar Biol

145:811–820

Madden D, Ballestero J, Calvo C, Carlson R, Christians E,

Madden E (2008) Sea turtle nesting as a process influenc-

ing a sandy beach ecosystem. Biotropica 40:758–765

Maloney JE, Darian-Smith C, Takahashi Y, Limpus CJ (1990)

The environment for development of the embryonic log-

gerhead turtle (Caretta caretta) in Queensland. Copeia

1990:378–387

Márquez-M. R, Peñaflores C, Vasconcelos J (1996) Olive rid-

ley turtles (Lepidochelys olivacea) show signs of recovery

at La Escobilla, Oaxaca. Mar Turtle Newsl 73:5–7

Matsuzawa Y, Sato K, Sakamoto W, Bjorndal KA (2002) Sea-

sonal fluctuations in sand temperature: effects on the incu-

bation period and mortality of loggerhead sea turtle

(Caretta caretta) pre-emergent hatchlings in Minabe,

Japan. Mar Biol 140:639–646

Maxwell JA, Motara MA, Frank GH (1988) A micro-environ-

mental study of the effect of temperature on the sex-ratios

of the loggerhead turtle, Caretta caretta, from Tongaland,

Natal. S Afr J Zool 23:342–350

McCoy CJ, Vogt RC, Censky EJ (1983) Temperature-con-

trolled sex determination in the sea turtle Lepidochelys

olivacea. J Herpetol 17:404–406

Merchant-Larios H, Ruiz-Ramirez S, Moreno-Mendoza N,

Marmolejo-Valencia A (1997) Correlation among ther-

mosensitive period, estradiol response, and gonad differ-

entiation in the sea turtle, Lepidochelys olivacea. Gen

Comp Endocrinol 107:373–385

Miller JD (1985) Embryology of marine turtles. In: Gans C,

Billett F, Maderson FPA (eds) Biology of the reptilia,

development A. Wiley, New York, NY, p 269–328

Miller JD (ed) (1997) Reproduction in sea turtles. The biology

of sea turtles. CRC Press, Boca Raton, FL, p 51–81

Mrosovsky N (1994) Sex ratios of sea turtles. J Exp Zool 270:

16–27

Mrosovsky N, Pieau C (1991) Transitional range of tempera-

ture, pivotal temperature and thermosensitive stage for

sex determination in reptiles. Amphib-reptil 12:169–179

Pandav B, Choudhury BC, Kar CS (1994) Discovery of a new

sea turtle rookery in Orissa, India. Mar Turtle Newsl 67:

15–16

Plotkin PT (2007) Olive ridley sea turtle (Lepidochelys oli-

vacea). Five-year review: summary and evaluation.

USFWS, Silver Spring, MD and NMFS, Jacksonville, FL

Reina RD, Mayor PA, Spotila JR, Piedra R, Paladino FV (2002)

Nesting ecology of the leatherback turtle, Dermochelys

coriacea, at Parque Nacional Marino Las Baulas, Costa

Rica: 1988–1989 to 1999–2000. Copeia 2002:653–664

Richard JD, Hughes DA (1972) Some observations of sea tur-

tle nesting activity in Costa Rica. Mar Biol 16:297–309

Shanker K, Pandav B, Choudhury BC (2004) An assessment of

the olive ridley turtle (Lepidochelys olivacea) nesting pop-

ulation in Orissa, India. Biol Conserv 115:149–160

Valverde RA, Gates CE (1999) Population surveys on mass

nesting beaches. In: Eckert KL, Bjorndal KA, Abreu-

Grobois FA, Donnelly M (eds) Research and management

techniques for the conservation of sea turtles. IUCN/SSC

Marine Turtle Specialist Group Publication No. 4, Wash-

ington, DC, p 56–60

Valverde RA, Cornelius SE, Mo CL (1998) Decline of the olive

ridley sea turtle (Lepidochelys olivacea) nesting assem-

blage at Nancite beach, Santa Rosa National Park, Costa

Rica. Chelonian Conserv Biol 3:58–63

Vega AJ, Robles Y (2005) Descripción del proceso de

anidación y biometría de hembras, huevos y nidos en tor-

tuga golfina Lepidochelys olivacea (Eschscholtz, 1829) en

Isla de Cañas, Pacífico panameño. Tecnociencia 7:43–55

Wallace BP, Sotherland PR, Spotila JR, Reina RD, Franks BF,

Paladino FV (2004) Biotic and abiotic factors affect the

nest environment of embryonic leatherback turtles, Der-

mochelys coriacea. Physiol Biochem Zool 77:423–432

Wibbels T (2003) Critical approaches to sex determination in

sea turtles. In: Lutz PL, Musick JA, Wyneken J (eds) The

biology of sea turtles II. CRC Press, Boca Raton, FL,

p 103–134

Wibbels T (2007) Sex determination and sex ratios in ridleys

turtles. In: Plotkin PT (ed) Biology and conservation of rid-

ley sea turtles. Johns Hopkins University Press, Baltimore,

MD, p 167–189

Wibbels T, Rostal DC, Byles R (1998) High pivotal tempera-

ture in the sex determination of the olive ridley sea tur-

tle from Playa Nancite, Costa Rica. Copeia 1998:

1086–1088

Yntema CL, Mrosovsky N (1980) Sexual differentiation in

hatchling loggerheads (Caretta caretta) incubated at dif-

ferent controlled temperatures. Herpetologica 36:33–36

Editorial responsibility: Matthew Godfrey,

Beaufort, North Carolina, USA

Submitted: January 18, 2010; Accepted: June 4, 2010

Proofs received from author(s): June 28, 2010

➤

➤➤

➤

➤➤

➤

➤➤

➤

➤➤

➤

Download
Save PDF to desktop

Email
Email completed PDF

Send for Signing
Email for others to sign

Print
Print document

NAME

Valverde Et Al 2010 n12:77

View Audit Log
Print With Audit Log
Duplicate Document

Fill Online, Printable, Fillable, Blank Valverde Et Al 2010 n12:77 Form

Related forms

Fillable Valverde Et Al 2010 n12:77

Fill Online, Printable, Fillable, Blank Valverde Et Al 2010 n12:77 Form

Related forms

First 20 documents completely FREE!