Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

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Journal of Experimental Marine Biology and Ecology

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Intake passage time, digesta composition and digestibility in East Pacific green turtles(Chelonia mydas agassizii) at Gorgona National Park, Colombian Pacific

Diego F. Amorocho, Richard D. Reina ⁎School of Biological Sciences. Monash University, Clayton, Victoria 3800, Australia

⁎ Corresponding author. Tel.: + 61 3 99055600; fax: +E-mail address: richard.reina@sci.monash.edu.au (R.

0022-0981/$ – see front matter. Crown Copyright © 20doi:10.1016/j.jembe.2008.04.009

A B S T R A C T

A R T I C L E I N F O

Article history:

We investigated the food d Received 6 February 2008Received in revised form 25 February 2008Accepted 9 April 2008

Keywords:DigestibilityIntake passage timeGreen turtleNutrition

igestibility of East Pacific green turtles, Chelonia mydas agassizii, at tropical coralreefs of Gorgona National Park in the Colombian Pacific and calculated the intake passage time (IPT) of 3different diets. We collected 150 faecal samples from turtles (mean straight carapace length 61.3±4.12 cmand mean mass 32.3±6.67 kg) to determine digesta composition and for measurement of neutral detergentfibre (NDF), acid detergent fibre (ADF), sulphuric acid lignin and protein. The mean (± S.D) IPT to recover atleast 73% of external markers (plastic beads) in the faeces was 23.3±6.6 days. The true NDF digestibility anddry matter digestibility were determined for high protein (fish), plant (fresh leaves of Araceae, Moraceae andBombaceae) and mixed (combination of both high protein and plant) diets. NDF values obtained fordigestibility of the protein, plant and mixed diets were 1%, 63% and 49% respectively. There was a largeamount of undigested plant material in the faeces, dominated by fruits of red mangrove (Rhizophora mangle).We considered the relationships between the type of food, IPT and apparent digestibility in the context of thenutritional contribution of an omnivorous diet. Our results suggest that for juvenile green sea turtles in theEastern Pacific combined diets are an opportunistic strategy responding to habitat features and supply tomaximise energy acquisition in transitional habitats such as Isla Gorgona.

Crown Copyright © 2008 Published by Elsevier B.V. All rights reserved.

1. Introduction

The acquisition of nutrients for all animals depends not only onfeeding but also on mechanisms of digestion and absorption toprocess the foods obtained in an interrelated set of needs andprocesses (Will et al., 2004). The green sea turtle (Chelonia mydas) isconsidered to be primarily a herbivore that feeds throughout most ofits range on seagrass pastures, but that will also consume some animalmatter (Seminoff et al., 2006), which it readily accepts in captivity(Mortimer,1982). Green turtles can feed on algaewhere seagrasses arenot available (Bjorndal, 1980; Bjorndal 1985; Forbes, 1994; Read, 1991)or consume both when they are present in the same area (Brand-Gardner et al., 1999; Ferrerira et al., 2006). Other plant material suchasmangrove leaves and fruits have been described as a substantial andnutritionally important part of green turtle diets in Australia (Limpusand Limpus, 2000) and juveniles of the East Pacific green turtle(Chelonia mydas agassizii) in Colombia (Amorocho and Reina, 2007).Consumption and effective digestion of a plant diet by sea turtlespresents a significant challenge. Turtles rely on microbial fermenta-tion in the large intestine to digest plant cell walls, but they can notchew their food to reduce particle size and facilitate this process(Bjorndal et al., 1990) because they don't have teeth. Thus, only by

61 3 99055613.D. Reina).

08 Published by Elsevier B.V. All rig

retaining material for longer in the gut with a slow passage rate canthey equal ruminants in the digestion of plant material by fermenta-tion. The primary nutritional end products of this fermentation are thevolatile fatty acids (VFA), an important source of energy in herbivoroussea turtles (Bjorndal, 1997). The quality of forage is a major deter-minant of the efficiency of digestion and can be assessed by mea-surement of acid detergent fibre, (ADF, cellulose and lignin) andneutral detergent fibre (NDF, cellulose, hemicellulose and lignin) usingthe detergent system (Van Soest, 1963; Van Soest and Wine, 1967).Lignin is the prime factor influencing the digestibility of plant cell wallmaterial and as lignin increases the digestibility, intake and animalperformance usually decrease because the percent ADF and NDFincrease (Van Soest, 1994).

Green sea turtle juveniles over about 40 cm straight carapacelength (SCL) shift their feeding behaviour from carnivorous toherbivorous when they move from pelagic to neritic developmentalhabitats (Bjorndal and Bolten, 1988; Hirth, 1997; Musick and Limpus,1997). In turtles that feed on seagrass, Thalassia testudinum, in theBahamas, nutritional assimilation occurs at a very low intake rate withhigh efficiency digestibility of cellulose (89%), hemicellulose (75%) andorganic matter (67%) by cellulolytic microflora present in the gut(Bjorndal, 1980). The same author suggests that the digestibilitycoefficients of NDF and ADF of green turtles foraging selectively onyoung blades of the seagrass reflect the specificity of their intestinalmicroflora. Bjorndal (1997), proposed that the optimal forage forgreen turtles may be that to which its gut microflora are adapted

hts reserved.

118 D.F. Amorocho, R.D. Reina / Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

(either seagrass or algae). Thus, it is expected that a less selective dietor one which is mixed (omnivorous) may change the composition andfunction of fermenting bacteria to affect the efficiency of digestion.However, nutrient gain in the form of VFA may be maximised by theability to ingest a greater quantity of food more rapidly (Bjorndal,1997).

Large juvenile and adult East Pacific green sea turtles are describedas primarily herbivorous feeding on seagrasses and/or marine algae inGalapagos and Mexico (Fritts, 1981; Green, 1983; Seminoff et al.,2002). Nonetheless, along the Coast of Baja California, they occasion-ally consume red crabs (López-Mendilaharsu et al., 2005) and otherchordates (Casas-Andreu and Gómez-Aguirre, 1980). A significantamount of animal matter (molluscs, amphipods, sardines andanchovies) was found in the stomachs of subadult and adult greenturtles captured in the Pacific coastal waters of Peru (Hays and Brown,1982) and fish eggs, molluscs, polychaetes and jellyfish have beenrecovered from the stomachs of turtles caught near the Pacific coast ofEcuador (Fritts, 1981). These findings suggest a carnivorous trend inthe feeding habits of East Pacific green turtles in the region. Thisbehaviour has also been observed in individuals consuming a mixeddiet of tunicates (Salpas spp.), terrestrial leaves (Ficus spp.) and algae(Gelidium spp.) at Gorgona National Park, in the Colombian Pacific(Amorocho and Reina, 2007). The high frequency of tunicates (73.8%)recovered through oesophageal lavages conducted in 84 juveniles(N40 cm SCL), showed that Colombian green turtles are omnivoreswith a diet biased towards animal matter (Amorocho and Reina,2007). Sea turtles consistently ingesting a mixed diet would almostcertainly develop a different microbial community than exclusivelyherbivorous turtles, in order to degrade the various complexcarbohydrates required to digest each food item efficiently (Bjorndal1985). So, long intake passage time (IPT) might occur to enable highlyefficient fibre digestibility because the microbial populations in thegreen turtle gut may have to adapt continually to changes in the typeand proportions of selected diet components.

Other than stomach content analyses and anatomical descriptions(Bjorndal, 1997; Green, 1994; Green and Ortiz, 1982; Hays and Brown,1982), studies on the digestive system of green turtles in the southeastPacific are lacking. Information about the digestibility of a mixed diet

Fig. 1. Map of Gorgona National Park in the Colombian Pacifi

will indicate the real benefit to turtles of eating animal matter toincrease energy gain in a transitional habitat such as Gorgona.Nutrient availability is limited by the amount of food and bycompetition between the rates of digestion and passage. This can bemaximised by ingesting a mixed diet as a way to obtain the essentialaminoacids required for protein synthesis and we may expect thatgreen turtles at Gorgona will try to obtain the maximum nutritionalbenefit by consuming an omnivorous diet. This might suppose adigestibility limitation because of the diverse range of food items theyneed to eat and the time required for microbial fermentation toproduce VFA. To elucidate these nutritional aspects of the East Pacificgreen turtle population foraging at Gorgona National Park in theColombian Pacific, we investigated the apparent digestibility of threetypes of diet, measured IPT and considered the effect of diet quality onjuveniles' growth. We addressed the following questions: 1. Do turtlesingesting an omnivorous diet in Gorgona have low digestiveefficiencies? 2. If so, can nutrient gain be maximised by the abilityto ingest a greater quantity of different types of food more rapidly? 3.If the East Pacific green turtle has a slow fermenting strategywith highdigestibility of fibre and organic matter, is ingesta retained within thefermentation chamber for long periods? By revealing these aspects ofdiet selection and digestive process, we can begin to understand hownutrition acts as a regulating mechanism for growth and maturity injuveniles and the importance of Gorgona as a foraging and develop-mental habitat for the East Pacific green sea turtle.

2. Materials and methods

2.1. Study site

Fieldwork was carried out at Isla Gorgona National Park (2° 50' - 3°00'N, 78°10' - 78°15' W) in Colombia. The island is 9 km long and2.5 km wide with a total protected area of 617 km2 and a maximumheight of 338 m, located 56 km offshore from the town of Guapi in thesouthern Colombian Pacific coast (Fig. 1). The shores of Gorgona arepredominantly steep plunging cliffs, with small sandy and pebblybeaches supplied on its eastern side by coral reef detritus. The island issurrounded by near-shore coral reefs where East Pacific green sea

c, showing location of the sea turtle in-water enclosure.

Table 1Total mass (g) of protein and plant feeding rations supplied every 3 days to 9 turtlesconfined in a near shore captive enclosure at Gorgona National Park, Colombia

Turtle Type of diet Plant ration (g) Protein ration (g) Total intake (g)

T1 Mixed 175 320 495T2 Mixed 225 100 325T3 Mixed 125 85 210T4 Mixed 223 210 433T5 Mixed 500 500 1000T6 Plant 292 - 292T7 Plant 300 - 300T8 Protein - 534 534T9 Protein - 500 500

Fig. 2. In-water enclosure to keep turtles confined near shore El Poblado beach. The enclosure was sunk to the water level after being placed in position.

119D.F. Amorocho, R.D. Reina / Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

turtles can be found resting or foraging. The IPT, digesta compositionand digestibility experiments were conducted within the constraintsof limited logistic and infrastructure conditions in the field. Gorgona isan isolated place without permanent electricity to adequately storecollected samples for some specialised analyses.

2.2. Turtles sampled, confinement and faecal collection

We conducted IPT and digestibility experiments between July andOctober 2005. Nine green turtles were caught by hand using snorkelat a depth of up to 7 m in coral reefs of La Azufrada and Playa Blanca.Straight carapace length (SCL) of all animals was measured asdescribed by Bolten (1999) and turtles were weighed and doubletagged in the front flippers with Inconel 1005-681S tags (NationalBand & Tag Co.), following standard techniques (Balazs, 1999). Aplastic Ziploc bag cut at the bottom was wrapped inside anotherplastic bag and covered with nylon cloth to be attached to theanimal's cloaca using a surgical thread, leaving the zip side at the endto enable collection of faeces in a modification of Bjorndal'stechnique (1980). Turtles were confined in an enclosure made up ofbamboo, PVC and plastic mesh, in water of approximately 3 m depth(Fig. 2). Three turtles were held at a time, each in a 2 m×2 m×2 mseparated compartment of the in-water enclosure for 3 consecutivesampling periods. Turtles were acclimated to the enclosure for 7 daysand fed with fresh leaves and small fish supplied ad libitum. Wecovered the enclosure with 6 mm square plastic mesh to reduce thepossibility of turtles eating food other than that provided in thelaboratory and we checked the enclosure daily to keep it free ofleaves, small fish and tunicates.

2.3. IPT and dietary treatments

After acclimation, we administered external markers to each turtlebefore being feeding it with the experimental diet and placing it backinto the enclosure. External markers are synthetic indigestible organicsubstances such as plastic beads, that are used in gut passage studiesbecause they can be counted, are not chewed into smaller pieces andcan easily be recovered from the faeces (Van Soest, 1994). This isprobably the best single measure of food passage through the entiregut if faeces can be collected (Warner, 1981). We used cylindricalyellow beads (2.0 - 3.0 mm×1 mm packaged into gelatin capsulescontaining 20 beads each) and introduced between three and fivecapsules into the lower oesophagus by pushing them through a plastichose. IPT was calculated from the integrated average of markersrecovered in the faeces following administration.

Animals were moved every three days into the laboratory to be fedwith 1 – 2% body mass of wet food as a maintenance diet (Higgins,

2003). The three experimental diets given to the turtles were: highprotein (fish); plant (a blend of Araceae, Moraceae and Bombacaeaeleaves) andmixed (a combination of animal and plant material). Mealswere prepared in the form of approximately 8 cm3

fish cubes and smallpackages of fresh leaves. The mass of food given each time varied withthe willingness of the turtle to ingest it. The diet rations were intro-duced using 2 plastic hoses to push the food deep into the oesophagus.The high protein group was composed of turtles with mass rangingfrom 27 to 31 kg; the plant group mass was from 31 to 36 kg and themixed diet group was from 33 to 48 kg. The three dietary treatmentsand mass of food administered to animals are presented in Table 1.

Turtles were taken out of the water every day at 3 p.m. and faecescollected from each individual's attached bag before returning them tothe enclosure. Mean value for water temperature recorded throughoutthe three experimental seasons at 3 m below the surface, was 28.3±0.3 °C; ranging from a minimum of 27.7 °C to a maximum of 29.0 °C.We measured body mass every 7 days and replaced faecal collectionbags as required.

2.4. Digesta composition and digestibility analyses

Collected faecal material was initially sorted into categories (fruits,leaves,mixture, coral and plastic).Wetmasswasmeasured for thefirst 3categories and then samples were preserved in 70% ethanol in plasticbags and stored at 4 °C in a portable fridge for organicmatter (OM), aciddetergent fibre (ADF), neutral detergent fibre (NDF) and lignin analyses.At the conclusion of the experiment, attached cloacal bags werecarefully removed from turtles and iodine (10%) applied around thecloaca before releasing the animals at the approximate site of capture.

Faecal samples were dried to constant mass at 60 °C for 48 to 60 hand stored in desiccators (Wood andWood, 1981). Following return tothe mainland, samples were dried again at 105 °C for 12 hours,

Table 2Recovery of administered markers from 6 turtles. Mean Ingesta Passage Time (IPT)=23.3±6.6 days

Turtle SCL(cm)

Number ofadministeredmarkers

Percentageof markersrecovered

First recoveryday

Last recoveryday

T2 59.0 100 96% 16 17T3 60.5 95 86% 14 19T4 60.8 60 100% 19 21T5 54.8 60 82% 23 27T6 62.2 60 100% 30 32T9 52.0 64 73% 30 32Mean±SD 58.2±3.6 73.2±17.3 89.4±10.0% 22.0±6.3 24.7±6.0

T1 was excluded because no markers were recovered by day 38 and animal wasreleased. T7 died after 12 days of treatment due to a hook ingested prior to experiment.T8 was released after day 31 without releasing markers.

Fig. 4. Percentage of different categories of ingested items collected from faecal samplesof 9 green sea turtles at Gorgona National Park.

Table 3Percentages of organic matter (OM), neutral detergent fibre (NDF), acid detergent fibre(ADF), lignin and protein of diet components and 3main faecal categories (leaves, fruits,mixture) collected from 9 green turtles at Gorgona National Park, Colombia

120 D.F. Amorocho, R.D. Reina / Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

weighed and ground in a Wiley Mill. Coral and plastic debris wereexcluded because they were not digested and were probablyconsumed incidentally by turtles while feeding on other food items.Faecal samples were aggregated for the whole collection period ofeach animal, then all faecal and food samples were analysed fororganic matter by heating in a muffle furnace at 550 °C for 12 h. Totalnitrogen was measured by the micro-Kjeldhal method and crudeprotein including both true protein and non-protein nitrogen wasestimated by multiplying total nitrogen content by 6.25, while ADF,NDF and sulphuric acid lignin analyses were performed followingstandard procedures (Van Soest, 1963; Van Soest and Wine, 1967).Food samples were also analysed for organic matter, crude protein,ADF, NDF and lignin. Hemicellulose content was calculated bysubtracting ash-free ADF from ash-free NDF, and cellulose wascalculated by subtracting lignin from ash-free ADF following proce-dures applied for green turtles previously (Bjorndal, 1980). Duplicateanalyses were performed when the quantity of material permitted.

3. Results

All results are reported as the mean±S.D.

3.1. Animal size and gut length

The mean SCL of the 9 studied turtles was 58.2±3.6 cm. (rangingfrom 52.2 to 62.2 cm) and mean mass was 32.3±6.7 kg (ranging from26 to 48 kg). One of the turtles (T7) died 12 days after capture and anecropsy revealed a 5 cm long-line “J” shaped hook # 7 (Mustard Co.)embedded in the oesophagus, that was presumably accidentallyingested by the turtle from some fisheries activity prior to capture.Large pieces of plastic debris were distributed all along the gut andmay have also contributed to the cause of death. The entire digestivetract of this turtle (63.7 cm SCL, 36.0 kg mass) from the beak to the

Fig. 3. Percentage of markers recovered and time elapsed for over 73% of markers to berecovered from faecal samples of 6 green turtles under 3 dietary treatments. P=plant,M=mixed, F=fish (high protein).

cloaca was 6.53 m in length. The necropsy showed that plant matterconsumed by the turtle had travelled along the gut in the form ofingesta packages (bolus), with fresh leaves observed between theoesophagus and the stomach (0 - 70 cm), boluses of partially digestedplant material located in the midgut (70 - 318 cm) and large amountsof mangrove fruits were distributed from mid to hindgut (318 -650 cm). All of the external markers given to the animal on day one ofthe experiment were contained within boluses distributed along themidgut (371 - 411 cm) at the time of death 12 days later.

3.2. IPT and dietary treatments

No markers were recovered from Turtle T1 after 38 days and onlyone marker was recovered from turtle T2 in 32 days, so these twoturtles were released to avoid health problems that could result fromextended captivity. Excluding T1, T7 and T8 due to zero or low markrecovery, the mean IPT for the 6 remaining turtles was 23.3±6.6 days(559 hr) ranging from 22.0±6.9 days (528 hr) for the first recovery dayand 24.7±6.6 days (593 hr) for the last recovery day of markers(Table 2). We ended the experimentwhenwe recovered at least 73% ofbeads from faeces (Fig. 3).

3.3. Digesta composition and digestibility analyses

Faecal sample percentage wet mass of mangrove fruits, leaves,remains composed of fragments of leaves and metabolic excretionstermed ‘mixture’, coral and plastic debris are shown in Fig. 4. Entire or

Diet Sample OM(%) NDF(%) ADF(%) Lignin(%) Protein(%)

Fish 87 1 0.1 0 98Leaves 73 63 53 38 8

Faecal Sample OM(%) NDF(%) ADF(%) Lignin(%) Protein(%)

MIXTUREMean±SD 68±3 74±19 58±6 28±5 16±2Min 63 0 47 20 12Max 74 87 68 36 19

LEAVESMean±SD 52±33 45±42 45±27 22±13 12±8Min 3 0 6 5 2Max 74 87 68 36 19

FRUITSMean±SD 58±24 61±30 49±20 25±11 14±6Min 3 0 6 5 2Max 74 87 68 36 19

Table 4Number of days that turtles were confined, total food intake, mean Intake Passage Time(IPT), total mass of faeces, percent neutral detergent fibre (NDF) and percent apparentdrymatter digestibility (ADM) of 9 green sea turtles at Gorgona National Park, Colombia

Turtle Daysconfined

Numberof feeds

Totalintake (g)

IPT(days)⁎

Wet massof collectedfaeces (g)

NDF(%)

ADM(%)

T1 38 13 3519 - 725 77 89T2 25 8 5850 16 866 82 67T3 19 6 1853 16 321 77 75T4 25 9 4138 25 841 75 78T5 31 10 10000 - 3377 60 66T6 19 7 2051 20 687 62 67T7 13 4 1200 - 1649 -61 -37T8 31 11 5674 31 535 19 91T9 31 10 5000 31 746 22 85Mean±SD 26±8 9±3 4365±2700 23.3±7.2 1083±933 59±25 77±10

⁎ Mean time elapsed between first and last recovery day of markers . NDF and ADM

121D.F. Amorocho, R.D. Reina / Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

chopped pieces of mangrove fruit were present in faeces throughoutthe experiment in all 9 assessed turtles. Percentages of OM, NDF, ADF,lignin and protein from fish and plant dietary treatments, as well asthe 3main faecal components are presented in Table 3. Through directobservation of the faecal material it appears that animal food (i.e. fish)was entirely digested, but some plant material (particularly mangrovefruits) showed little visible sign of digestion (Fig. 5). True NDF andapparent dry matter digestibility (ADM) of the assessed turtles arepresented in Table 4.

4. Discussion

4.1. Turtle size class

East Pacific green turtles using coral reefs and surrounding watersof Gorgona National Park were small and large juveniles (52.0 – 62.2

Fig. 5. Pieces of a) the mangrove fruit Rhizophora mangle and b) leaves of Ficus spp.collected in the faeces of 9 assessed turtles.

values of T 7 are negative because intake was less than the excreta produced after12 days of captivity prior to death from unrelated hook ingestion. These values were notincluded for mean and SD estimation.

SCL), confirming recruitment of young turtles at the size of 50.0 -70 cm SCL to neritic habitats of this protected area. The mean size of58.2±3.6 cm at Gorgona is similar to that measured from juveniles atBahía Magdalena and Estero Banderitas in Baja California Sur, Mexico(López-Mendilaharsu et al., 2005) and is typical of juveniles on thePacific coast of the Baja California Peninsula (Koch et al., 2007) inprotected reefs or mangrove habitats where they can find food andshelter. However, juveniles in Gorgona, consume a mixed diet biasedtowards animal matter (Amorocho and Reina, 2007) while those inBahía Magdalena feed mostly on algae and seagrasses (López-Mendilaharsu et al., 2003; López-Mendilaharsu et al., 2005; Seminoffet al., 2002). At Gorgona, it is advantageous for juvenile green turtlesto consume a high protein diet with good nutrient assimilation tocontribute energy for growth while juveniles migrate betweenforaging and developmental habitats. In comparison, stable isotopesin scutes of N36 cm young green turtle recruited to neritic habitats offGreat Inagua (Bahamas), reveal rapid ontogenetic shift from carnivor-ous to herbivorous lifestyle (Reich et al., 2007). Green sea turtlejuveniles at a size class of 50 - 70 cm continue feeding mainly onanimal matter or at least consuming an omnivorous diet in neriticnear shore habitats of Gorgona (Amorocho and Reina, 2007) in anontogenetic pattern different to that observed in green turtles of theCaribbean. These different dietary patterns are probably indicative ofthe transitional nature of the Gorgona habitat as a stop-over point inturtle migration.

4.2. IPT and dietary treatments

There is very little information about the use of external markers(beads) to estimate time elapsed between ingestion and egestion insea turtles. The percentage of bead recovery in faeces of 6 turtles wasnot always over the 95% typically applied in ruminant passage studies(Van Soest, 1994). However, our mean of 89% (from the 73% to 100%range) of markers recovered was similar to the 88% - 100% collected in3 juveniles of similar size class (50.3 – 55.2 cm SCL) in Australia (Brandet al., 1999). It is possible that some markers were lost through smallholes between the collection bags and the skin around the cloaca. It isunclear why no markers were recovered after at least 32 days from 2turtles. We suspect that they may have been regurgitated followingadministration and escaped through the mesh of the enclosure butcannot exclude the possibility that their movement through the gutwas retarded for some unknown reason.

Mean IPT for captive turtles was long with a mean time of first andlast marker recovery 528 to 593 h compared to digesta retention timesof 156 to 325 h estimated in wild immature green turtles at Moreton

122 D.F. Amorocho, R.D. Reina / Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

Bay, Australia (Brand et al., 1999). This difference is a consequence ofthe experimental design and different techniques used in both studies.In the Moreton Bay study, turtles were freed into the wild to resumefeeding after the markers were administered, then recaptured andsacrificed after 5 – 9 days. This was before they excreted any beads, soIPT was estimated from beads counted at some point along the gutfollowing necropsy. We think that our measurements of IPT are moreprecise because the passage of beads was measured all the waythrough from ingestion to egestion, with excreted beads collecteddaily in attached cloacal bags. Additionally, the necropsy of turtle T7that died from the unrelated hook ingestion, showed that markerswere only half to two thirds the way along the gut after nearly 300 h(12 days). The point of the gut that the markers reached was similar tothat of Brand et al. (1999) in a comparable time, but our resultsindicate that it took about another 11 days for markers to reach theend of the gut and be excreted.

4.3. Digesta composition

The presence of large amounts of undigested mangrove fruits infaeces of all sampled turtles reflects the abundance of this food itemin Gorgona waters. Mangrove fruits are found drifting in surfacecurrents carrying organic matter from the mainland rivers of la Tola,Sanquianga and Patia in South Western Colombian Pacific. Theabundance and availability of Rhizophora mangle fruits in superficialwaters of Gorgona varies between seasons and may influence die-tary changes in foraging green turtles from plant to animal matter(tunicates). The presence of undigested mangrove fruits in the faecesof all turtles can be associated with seasonal abundance prior to theexperiment. The low percentage of NDF (63%) in freshmangrove fruitsmight be responsible for the high intake of this food item before ourstudy was conducted. The consequence of consuming Rhizophoramangle needs to be considered in further studies, since mangrovesplay a key role in the nutritional ecology of immature green turtlepopulations in the Pacific (Limpus and Limpus, 2000).

4.4. Digestibility

Fromdirect observation of digesta composition in the entire gut, it isevident that green turtles at Gorgona are opportunistic consumers.Distinct packages of food separated in boluses containing mangrovestalks, small fish and leaves reflects the omnivorous behaviour ofjuveniles in thewild. This also indicates how often they switch betweenanimal and plant food items. A similar pattern of shifting diet wasobserved in Australia, with separate boluses representing differentfeeding bouts divided along the digestive tract (Brand et al., 1999).

The NDF digestibility of six turtles from which we recoveredmarkers varied, with apparent dry matter digestibility (ADM) highestin the protein (fish) diet group, lowest in the plant diet group andintermediate in the mixed diet group, although sample size was smallin the first two groups. The protein diet (NDF 21±2%) seems to be welldigested, with our measured ADM of 85 – 91% very similar to that forgreen turtles (82 – 90%) fed commercially prepared high protein diets(Hadjichristophorou and Grove, 1983; Wood and Wood, 1981). As aresult, juveniles in Gorgona might grow more rapidly than thosefeeding on sea grass or algae in the Caribbean (Bjorndal, 1985) andAustralia (Brand et al., 1999), considering protein as the limitingnutrient responsible for growth and sexual maturity. Thus, we concurwith Bjorndal (1985) that growth rates in wild green turtles are undernutritional rather than genetic control. We propose that the need forrapid growth is indicated by the diet biased toward animal matter ofjuvenile turtles at Gorgona (Amorocho and Reina, 2007), in contrast tothe assumption that juveniles must exclusively consume plant matteras they approach adulthood.

For animals fed with plant diet, 63% of organic matter consisted ofNDF and ADM was 67%, in agreement with the ADM of 66% measured

from green juveniles consuming Thalassia testudinum blades fromgrazed plots in Union Creek, Bahamas (Bjorndal, 1980). NDF digest-ibility for turtles varies depending on its composition, becausehemicellulose can be partially digested by sea turtles but lignin cannot. In addition, lignin and cutin appear to reduce digestibility of otherNDF components. NDF digestibility also depends on the species and itsdigestive anatomy, size and strategy and so ruminants digest moreNDF than hind-gut fermenters such as sea turtles.

4.5. Comparisons with ruminants

In general there are two options available for a herbivore: to eithermaximise digestive efficiency or to maximise intake (White et al.,2007). These are competing process because digestion is a time-dependent process, so that maximum digestion requires a long time. Ifeach unit of forage must be held in the digestive tract a long timeduring microbial fermentation, then a species maximising digestiveefficiency cannot eat a large amount of food because the capacity ofthe gut is limiting. Conversely, a species maximising intake willsacrifice digestive efficiency because rate of passagemust be increasedto make room for the additional forage and therefore potentiallydigestible material will pass through the digestive tract (Mertens andEly, 1979). Ruminants are more efficient at digesting the low andmedium quality forage normally consumed by these animals, usuallyin 24 - 48 h, but ruminants are constrained by rumen fill and rate ofpassage on how much they can eat in a day (Leng, 1990). Hind-gutfermenters like green sea turtles are not similarly constrained andthus can compensate for lower digestive efficiency by simply eatingmore. When food is not limiting, hind-gut fermenters can attain anequal or higher rate of energy (or digestible dry matter) intake.Ruminants may be at an advantage when food quantity is limited, buthind-gut fermenters such as green sea turtles usually have the optionof consuming more of a higher quality food, which is generallyabundant in Gorgona. Analysis of plant and animal characteristics thatinfluence digestion and intake suggests that maximum intake ofdigestible drymatter is influencedmore by the proportion of fiber thatis indigestible and the rate of passage than by the rate of fiberdigestion. Thus, with a slow passage rate and increased digestibility,the green turtles at Gorgona maximise nutrient gain in the form ofVFA through an omnivorous diet.

4.6. Temperature implications

Temperature can be a major variable limiting digestive efficienciesin poikilothermic animals. For this reason, the intake rate anddigestive efficiency of turtles will be affected by geographical andseasonal variations in water temperature. Body temperature of greenturtles in Gorgona are not more than one or two degrees C differentto the water temperature at 3 m depth, which is not sufficient tosignificantly affect IPT or decrease digestibility. Thus, the microbialincubation temperature of these turtles was considerably cooler thanthat of endothermic ruminants, further contributing to relativelylengthy gut passage times. Reptiles can attain digestive efficienciesroughly comparable to those of ruminants by subjecting feed tomicrobial fermentation processes for longer periods of time (Mackie,2002). This strategy is not feasible for endothermic herbivores with arapid metabolic rate and limited time available to extract energy andnutrients (Farlow, 1987). Thus the main advantage of mastication inruminants is to reduce the time needed to attain a digestibility thatreptiles can achieve simply through a longer IPT. Consumption of amixed diet and a regulated temperature are the key aspects to main-tain the efficiency of themicrobial fermentation process driving intakerate, passage time and digestibility of green turtles in developmentaland feeding grounds of Gorgona. Water temperatures of 21.3 - 25.5 °Crecorded in feeding grounds of Moreton Bay, Australia (Brand et al.,1999) and of 19 – 28 °C in Baja California, Mexico (Lluch-Belda et al.,

123D.F. Amorocho, R.D. Reina / Journal of Experimental Marine Biology and Ecology 360 (2008) 117-124

2000), are similar to those measured in Gorgona and are typical oftropical waters. Fluctuation in temperature over the course of the yearwill have some effect on the fermentation process, digestive efficiencyand IPT for a poikilotherm animal. This might be the case for turtlesfeeding on seagrasses in the Bahamas during atypical El Niño years,when normal temperature ranging from 24 – 29 °C rises to above 34 °C(Bjorndal, 1980), increasing the rate of digestion and food intake.

4.7. Conservation and management issues

The death of turtle T7 due to a hook embedded in its oesophagusraises the issue of high sea turtle mortality in artisanal and industrialfisheries. Incidental capture in long-line fishing gear has beendocumented as the major threat for green sea turtle populations inthe Pacific Ocean (Seminoff, 2004). Technological changes in currentfishing practices and the use of de-hookers need to be rapidlyintroduced in order to reduce green sea turtle by-catch in Colombia. Inaddition, the amount of plastic (1.6%) that we recovered from thefaeces of turtles indicates the magnitude of marine pollution affectingthe feeding behaviour and health of green sea turtles. It is important tocontinue documenting the occurrence and impact of plastic debris indiets consumed by sea turtles for the design and implementation of aregional management strategy in the Pacific. Results from our studyshow how the transitional feeding grounds of Gorgona National Parkprovide nutritional opportunities that benefit turtles' digestiveefficiency that can be translated into growth and fitness. For managersto better protect them, it would be important to address theabundance of debris and quality of food items ingested by omnivorousjuveniles in the protected habitats and to ensure that Gorgonaremains available as a transitional habitat.

4.8. Conclusions

It seems that for juvenile green sea turtles in the Eastern Pacificcombined diets are an opportunistic strategy responding more tohabitat features and supply rather than to the species' herbivorouspreference. Perhaps foraging patterns in juveniles are driven moreby the rookery geographical distribution, type of habitat and foodavailability than by ontogenetic changes experienced by animalswhen they enter neritic shallow waters. This relationship may be thereason for IPT differences observed in assessed turtles, suggesting thatthe cellulolytic gut microflora is capable of degrading plant cell wallsand proteinwith acceptable digestibility efficiency under amixed diet.Thus, juveniles recruited to Gorgona's habitats seem to be consuminga better quality and more degradable mixed forage that increasesgrowth rates for more rapid achievement of sexual maturity. In regardto our experimental questions posed earlier, we can say the following:1) turtles in Gorgona consuming an omnivorous diet exhibited highfibre digestibilities (82 – 88%), similar to the 72 – 91% reported forgreen turtles in the Caribbean (Bjorndal, 1980); 2) NDF of 77% for themixed (omnivorous) diet suggests that intake does not necessarilyoccur rapidly. However, the animal might be compensated withnutrients for growth supplied by the high protein component of thediet and efficiency of the fibre content digestibility; 3) An IPT between14 and 32 days seems to be a reasonable time for an ectothermicanimal to efficiently digest meals composed of different types of food.Further research combining microbiological techniques, moleculartags, stable isotopes and satellite tracking, will contribute to furtherunderstanding the foraging ecology and importance of GorgonaNational Park as a transitional habitat for growth, performance andproductivity of East Pacific green turtles.

Acknowledgements

We thank the US National Fish and Wildlife Foundation (NFWF),The Rufford Small Grants Foundation (UK), the Colombian National

Parks Administrative Unit (UAESPNN), Centre for Research andEnvironmental Development (CIMAD, Colombia), the InternationalCentre for Tropical Agriculture (CIAT), the Zoo of Cali (Colombia), theR.E. Train Education for Nature (EFN), the University of West Indies(Barbados), and theWider Caribbean Sea Turtle Network (WIDECAST),for their financial, logistic and human contributions to this project.Our special gratitude to A. Pavia, M.J. Restrepo, J. Hoyos, A. Ruiz, J.Puertas, J. Rodriguez, L. Merizalde and other volunteers for theirenergy and enthusiasm proved while catching and processing turtlesduring the fieldwork. We also thank Dr. Fernando Gast from theColombian Biodiversity Research Institute Alexander von Humboldtand Dr. Fernando Castro from the Department of Zoology of the ValleUniversity for their support during this investigation. This study wasconducted under research permit DTSO 0029 from the UAESPNN(Colombia) and ethics approval BSCI /2003/04 from Monash Uni-versity (Australia). [SS]

Appendix A. Supplementary data

Supplementary data associated with this article can be found, inthe online version, at doi:10.1016/j.jembe.2008.04.009.

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