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Journal of Applied Animal Welfare Science

ISSN: 1088-8705 (Print) 1532-7604 (Online) Journal homepage: http://www.tandfonline.com/loi/haaw20

Training Large Macaws for Artificial InseminationProcedures

Frédérique Leblanc , Géraldine Pothet , Michel Saint Jalme , Mathieu Dorval& Dalila Bovet

To cite this article: Frédérique Leblanc , Géraldine Pothet , Michel Saint Jalme , Mathieu Dorval& Dalila Bovet (2011) Training Large Macaws for Artificial Insemination Procedures, Journal ofApplied Animal Welfare Science, 14:3, 187-210, DOI: 10.1080/10888705.2011.575741

To link to this article: http://dx.doi.org/10.1080/10888705.2011.575741

Published online: 17 Jun 2011.

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JOURNAL OF APPLIED ANIMAL WELFARE SCIENCE, 14:187–210, 2011

Copyright © Taylor & Francis Group, LLC

ISSN: 1088-8705 print/1532-7604 online

DOI: 10.1080/10888705.2011.575741

Training Large Macaws for ArtificialInsemination Procedures

Frédérique Leblanc,1 Géraldine Pothet,1

Michel Saint Jalme,1 Mathieu Dorval,1 and Dalila Bovet2

1Muséum National d’Histoire Naturelle (MNHN), UMR 7204,MNHN-CNRS-Paris VI, Ménagerie du Jardin des Plantes, Paris, France

2Laboratoire d’Éthologie et Cognition Comparées, Université ParisOuest Nanterre La Défense, Nanterre Cedex, France

For some endangered parrot species, captive breeding may be the only insurance for

their survival. However, many individuals in captivity do not reproduce. Artificial

insemination (AI) may help overcome reproductive failures or geographic distance.

For semen collection in birds, massage is the most commonly used method.

However, this process, which usually requires capture and restraint, involves risk of

stress and injuries. The aim of this experiment was to train large macaws to accept

the physical manipulations of their body parts needed for the artificial insemination

process. Within 15 weeks, a male and a female Buffon’s macaw (Ara ambiguus)

learned to accept handling without apparent stress. A pair of green-winged macaws

(Ara chloropterus) progressed more slowly and displayed some signs of stress. This

stress highlights the need to monitor the birds’ possible signs of discomfort during

the training in order to adapt the working protocol. These results demonstrate that

it is possible to train the AI behaviors, thus avoiding the capture, restraint, and

anesthesia. An added benefit to this is the potential for the training to provide a

form of behavioral enrichment.

Parrots are one of the most endangered groups of birds in the world (Juniper

& Parr, 1998; Snyder, McGowan, Gilardi, & Grajal, 2000). The situation in

Latin America and the Caribbean is especially serious: 65 of the 151 species of

parrots are endangered (BirdLife International, 2010). For some species, habitat

Correspondence should be sent to Michel Saint Jalme, Muséum National d’Histoire Naturelle,

57 rue Cuvier, 75005 Paris, France. Email: mstjalme@mnhn.fr

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188 LEBLANC ET AL.

destruction is the main cause of population decline, and reproduction in captivity

may be the only chance for survival (Snyder et al., 2000). Captive reproduction

requires taking into account the birds’ basic biological and behavioral require-

ments in order for them to successfully adapt to their environment and the need to

manage genetic diversity through international breeding programs (Saint Jalme,

2002). However, optimal genetic management may conflict with free choice of

the partner, which appears to be one of the keys of breeding success (Curio,

1998; Sutherland, 1998). Thus, reproduction in captivity, where mate choice

is limited due to low number of individuals, is particularly challenging for

species of endangered parrots such as Amazona sp. and macaws (Derrickson

& Snyder, 1992). Considering the failures of reproduction-related behavioral

incompatibilities between managed captive pairs, artificial insemination (AI)

may constitute a better alternative. Such an alternative could also allow the

formation of breeding pairs between birds in different breeding facilities. AI has

already been successfully carried out in a variety of bird species:

1. Falconiformes (Boyd, 1978);

2. Galliformes (Caïn, 1978; Saint Jalme, 2002; Saint Jalme, Lecoq, Seigneurin,

Blesbois, & Plouzeau, 2003);

3. Sphenisciformes (O’Brien, Oehler, Malowski, & Roth, 1999);

4. Gruiformes (Jones & Nicolich, 2001; Saint Jalme, Gaucher, & Paillat,

1994); and

5. Psittaciformes (Brock, 1991; Samour, Smith, Moore, & Markham, 1986).

The massage method is the most common procedure used for semen collection

for birds in the wild who are noncooperative and not imprinted on humans (Gee,

Bertschinger, Donoghue, Blanco, & Soley, 2004). Semen collection involves

massage to stimulate the birds with pressure applied on the sides of the cloaca

to extract the sperm.

Female insemination is more challenging. To introduce the sperm, the oviduct

must be localized, either by palpation—everting the cloaca—or by use of a

speculum of appropriate size; use of a speculum may require gaseous anesthesia.

AI is a labor-intensive process that requires caregivers to catch and restrain the

birds. This may pose a problem particularly for large psittacines, who can inflict

serious bites and therefore increase the risk of injuries for both bird and handler.

The only AI attempts documented with macaws describe the harvesting of

semen carried out under sedation with electroejaculation (Harrison & Wasmund,

1983), a technique that is not recommended because of the risks of burns and

convulsions (Gee et al., 2004). Restraint and sedation of nonhuman animals

are sources of stress and can affect their welfare (Morgan & Tromborg, 2007).

Although AI may be an important tool for reproduction in some species, it is

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ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 189

not currently used with large parrots because of the difficulty of the procedure,

the stress for the birds, and the risks of injuries for both birds and handlers.

In zoos and aquariums, training has now become a common practice, making

it possible to teach animals to voluntarily accept a variety of potentially stressful

procedures. This avoids the need for using excessive restraint or anesthesia. For

example, macaws have been successfully trained to accept tactile examinations,

blood collection, insertion of a thermometer in the cloaca, and inhalations of

medication (Heidenreich, 2004).

To teach cooperative husbandry behaviors, we applied principles of operant

conditioning with positive reinforcement. We used positive reinforcement, gener-

ally in the form of preferred food items, to shape and increase the probability of

the desired approximations until the target behavior was obtained (Heidenreich,

2007).

In order to train wild birds to accept the procedures necessary for AI, they

must first accept handling by trainers. Birds in general are very sensitive to stress

induced by handling (Straub, Forbes, Pees, & Krautwald-Junghanns, 2003).

Barnett, Hemsworth, Hennessey, McCallum, and Newman (1994) compared a

group of hens subjected to minimal contact with humans to a group subjected

daily to 15 min of additional contact mimicking usual husbandry procedures.

This additional contact caused adverse changes in cell-mediated immunological

responsiveness (thus probably involving a chronic stress response), even though

hens are domestic animals. Consequently, it is important to ensure that the

training carried out to avoid the need for restraint is not in itself a source of

stress that could reduce welfare or reproductive success (Mason, Clubb, Latham,

& Vickery; Swaisgood, 2007). Chronic stress may be revealed by changes in

behavior performed by animals, such as exploratory behavior, feeding, repro-

ductive behaviors, locomotion, apathy, vigilance, aggression, and stereotypic

behaviors (Mason et al., 2007; Morgan & Tromborg, 2007; Swaisgood, 2007).

Stereotypic behaviors in parrots can be locomotive (e.g., pacing, perch circles,

corner flips, and route tracing) or oral (e.g., wire chewing, sham chewing,

food manipulation, and dribbling; Garner, Meehan, Famula, & Mench, 2006).

Stereotypy is important to study because it is associated with frustration and poor

psychological welfare in captive animals (Garner, Meehan, & Mench, 2003).

The experiment was performed on a pair of Buffon’s macaws (Ara ambiguus)

and a pair of green-winged macaws (Ara chloropterus) held at the Ménagerie du

Jardin des Plantes, a zoo belonging to the National Museum of Natural History,

Paris, France. The goal of the experiment was to develop a program to train male,

large macaws so that they could learn to accept the semen collection procedure

without capture and restraint, thereby avoiding acute and chronic stress. For

the females, the objective was to train them to accept the introduction of a

speculum into the cloaca and to introduce their heads into a mask used for

gaseous anesthesia. This latter process was considered as an alternative method

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190 LEBLANC ET AL.

in case the behaviors necessary for insemination could not be trained. In order

to evaluate the subjects’ general welfare, the baseline behavior of birds was

recorded outside of training sessions throughout the program. We wanted to

answer the following questions:

1. Is it possible to train large psittacines to accept AI handling procedure?

2. Would this training be a source of stress?

3. Could the training provide some behavioral enrichment?

MATERIALS AND METHODS

Species, Sample, and Housing

The Buffon’s macaw is classified in the endangered category by the International

Union for Conservation of Nature, and the species is registered in Appendix I

of the Convention on Trade in Endangered Species (CITES). The nominal

subspecies Ara ambiguus ambiguus lives in Central America and ranges from

eastern Honduras to eastern Panama and northern Colombia. The subspecies Araambiguus guayaquilensis, the most threatened, is restricted to Ecuador where

the population is estimated at just 100 individuals (Snyder et al., 2000). The

three major threats to macaws are (a) the loss of their habitat; (b) the poaching

of nests for the pet trade; and (c) to a lesser extent, the poaching of birds

either for food or for the collection of feathers (Grajal, 2000). Unfortunately,

the Buffon’s macaw seldom reproduces in captivity (Juniper & Parr, 1998). The

green-winged macaw (Ara chloropterus) lives in eastern Panama and in South

America from Colombia to northern Argentina. The species is registered in

Appendix II of CITES. Both species are large birds, reaching 95 cm in overall

length and weighing 1,200 g to 1,300 g (Dunning, 2008).

Training was carried out with two pairs of macaws who had never successfully

reproduced. The first one was a pair of Buffon’s macaws (BM) living together

since the beginning of the experiment: a 7-year-old male and a 2-year-old female.

They were held by the Ménagerie for 2 months at the time the experiment began.

The second was a pair of green-winged macaws (GWM) living together at the

Ménagerie for 15 months before the beginning of this experiment: a male almost

3 years old and a female 22 years old. The 4 subjects were all born in captivity.

The two BM and the male GWM had been reared by their parents. The female

GWM was given to a zoo by a private individual when she was 9 years old, and

her rearing condition is unknown.

The aviary of the BM measured 4.6 m � 4.7 m � 5 m. An artificial nest was

fixed in it. Branches were installed at the start of training for both enrichment

and to facilitate the locomotion of the birds. The aviary of the GWM measured

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6.1 m � 4.7 m � 5 m. An artificial nest was installed 6 weeks after the experiment

began. Both aviaries were visible to the public; the visitors’ path was at a distance

of 1 m. The birds were fed twice a day: at 8:45 a.m. they were given extruded

pellets, and a fresh fruit meal was given to them at 12:00 p.m. Water was

provided ad libitum.

Training Process

Two trainers, a man and a woman, took turns training all 4 subjects. One of the

trainers had no previous contact with the birds prior to the experiment; the other

trainer had minimal contact during transport and medical vaccination.

Training was carried out inside the aviaries, 5 days a week. It was carried out

each day in two sessions of 10 min per bird at midmorning and early afternoon;

the birds continued to receive their customary diet. This training was done from

February 19, 2007, to June 28, 2007, for the GWM and from March 6, 2007,

to June 4, 2007, for the BM.

Verbal cues were used for the first two stages. Then, as a visual signal,

the index finger was shown to the bird during the third stage (as described in

Training Stage 3 following). A food reward preferred by the individual bird

(sunflower seeds, pieces of walnuts, hazelnuts, and extruded pellets) was given

after each correct response or behavior approaching the correct response at the

beginning of a new exercise. A conditioned reinforcer, “very good,” was used

to mark a successful behavioral approximation.

A secondary reinforcer or “bridge” was used between the emission of the

desired behavior and the delivery of rewards. This was a sharp snap emitted

by a clicker held with one hand. This sound constitutes an acoustic secondary

reinforcement, acquiring its reinforcing properties by immediately preceding

each food reward (McCall & Burgin, 2002). Even brief delays are detrimental

to learning; therefore, the function of training practice is to mark the precise

moment of the correct execution of exercises and to indicate that a reward will

follow (Heidenreich, 2007). We used both the verbal reinforcer, “very good,”

and the clicker because the birds were already used to this verbal expression

(sometimes said by the caretakers), but the clicker allowed a more precise

and more standardized signal as a bridge. If the parrot expressed a nondesired

behavior such as attempting to bite trainers, the bird was ignored: the trainer

turned round and ceased interacting with the bird for 10 s.

Training Stages

Stage 1: Approaching and accepting hand-feeding. In this stage, the

birds learned to gradually associate the sound of the clicker to the food reward.

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192 LEBLANC ET AL.

The trainer installed a wooden perch (1.2 m high) beside a large branch. The

trainer called each individual bird by name and presented some food, handheld.

If the bird did not approach, the trainer put the food on a branch situated between

bird and trainer, moved away, and again called the bird by name. As soon as the

bird took the food reward, the trainer activated the clicker. When the subject had

learned to reliably approach the trainer to a specific location, we began Stage 2

of the training process.

Stage 2: Go up the perch. The cue “place” was given. Initial approxima-

tions had the trainer holding the reward toward the bird above the perch. Next,

the reward was placed on the perch. In the final training approximations, the

subject was conditioned to move to the proper location without having to first

see the reward.

Stage 3: Accepting beak touching. A visual signal, the index finger, was

shown to the bird; this indicated to the bird that the trainer was going to attempt

to touch a body part. The bird was rewarded, with acoustic bridging, for staying

on the perch without escaping or displaying defensive or attacking reactions. The

trainer proceeded to approach the beak with the index finger and then calmly

touched the beak.

Stage 4: Accepting touches to the body including the wings, the back,

the tail, and the cloaca. The trainer started by touching the wings, ap-

proached with the index finger, then proceeded to touch gently, and finally

touched with more pressure. The trainer proceeded in the same way for the

touches of the back, the tail, and finally of the cloacal area.

Stage 5a: Accepting massage and for females the introduction of thespeculum. The trainer held the reward in front of the bird’s head and rein-

forced each long caress accepted with no muscle tension or avoidance behaviors

exhibited by the bird. For the females, the speculum was placed in contact with

the cloacal zone initially and then introduced into the cloaca itself. The female

was rewarded on a continuous reinforcement schedule during the introduction

of the speculum, which was lubricated beforehand with Vaseline.

Stage 5b: Accept the mask and a light pressure on the body (females

only). The verbal order, “touch,” was given. At the beginning of this stage, the

reward was put at the entry of a transparent anesthesia mask, and then it was

applied against the mask from the outside. This stage was trained at the same

time as the massage approximations.

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ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 193

Evaluation of the Performances

Success of training was evaluated for each stage before progressing to the next

step in the process: a stage was considered mastered if three orders had been

successively executed in the same session over the span of three consecutive

sessions.

During the training process the following parameters were recorded:

1. Number of sessions for engaging or for passing a stage;

2. Number of orders given; and

3. Number of behaviors executed per order, including distress calls, fear

postures, and threatening postures directed toward the trainers.

Participation of the birds in a given training session was measured using the

criteria, “taking food from the hand at least once.”

Behavioral Data Recorded Outside Training Sessions

The behavioral observations collected outside of the training sessions were

carried out from February to the end of June in three daily sessions of 20 min

each for the 2 individuals of each aviary: the first one starting at 9:15 a.m., the

second one after the morning training at 11:15 a.m., and the last one following

the afternoon training at 3:30 p.m. The order of observation of the two aviaries

was alternated each day. Behavioral data were collected with the scan sampling

method (Altmann, 1974) with scans taken every 30 s.

The behavioral categories recorded were resting, vigilance, foraging, loco-

motion, solitary activities others than stereotypic behaviors, possible stereotypic

behaviors, comfort behaviors, and social behaviors. The reproductive behaviors

were also quantified. These behaviors are described in more detail in Table 1.

For each session, the number of visitors passing in front of the aviary was also

noted.

Data Analysis

The behavioral data were used to calculate average frequencies of behaviors per

day for each stage of training. The frequencies of one given behavior for each

day were calculated by dividing the number of observations of this behavior in

that day by 120 (number of scan samples for 1 day). We then averaged these

frequencies per day for each stage of training.

The duration of each stage depended on the progression of the birds and

varied among subjects. Stage 0 corresponded to the period before the start of

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TABLE 1

Ethogram for Ara ambiguus and Ara chloropterus

Resting� Sleep: eyes are closed or beak is under the back or wings feathers

Vigilance� Perched on two feet: the bird is perched high, standing on two feet, head up, eyes open� Leaning on two feet: the bird is standing on two feet, body leaning toward the front, wings a little

spread, looking at a particular point� Perched on one foot: the bird is perched high, standing on one foot, head up, eyes open� Hanging: the bird is hanging from the wire mesh or from a rope by the feet and/or the beak, eyes open

Foraging� Eats food held in foot: the bird holds a piece of food in one foot and eats it� Pecks from feeder: the bird takes food from the feeder with beak� Looks for food: the bird walks looking on the ground and pecking in different directions� Pecks from ground: the bird pecks on food or vegetation that is on the ground� Eats: the bird chews some food or drinks the juice from a fruit� Asks for food: the bird lowers front of body, tilts head toward partner, who holds some food in beak

or foot� Steals food: the bird takes or tries to take food from partner’s foot� Gives food: the bird puts from beak some food in the partner’s beak, after being asked.� Drinks: the bird puts beak in water, and swallows it

Locomotion� Walks: the birds moves around using feet� Climbs: the bird climbs using beak and feet� Flies: the birds moves around using wings

Solitary activities other than stereotypies� Acrobatics: hanging to the ceiling’s mesh by one or two feet, or by one feet and beak, the bird swings

and gesticulates� Tears to shreds: the bird bites, tears to shreds, or tears off some bark, leaves, or twigs� Wings flap: the bird hangs on a branch or the mesh with feet and beak and flaps wings� Nods: the bird repeatedly nods head up and down� Bites metal: the bird bites the wire mesh or some wires� Bites a rope: the bird uses beak to manipulate a rope� Bites rock: the bird uses beak to manipulate a rock� Turns: turns around a branch� Plays alone: gives a peck to a rope or a hosepipe hanging and looks at it while it moves

Stereotypies� Intention of flying: the bird sits on a branch, extends neck, flaps wings slightly, then opens wings

without flapping and vocalizes. Often also shakes head to the right and the left (behavior observed in

the male BM)� Weak intention of flying: same as aforementioned, but the wings are not spread out and the bird does

not vocalize (behavior observed in the male BM and the female GWM)� Shakes head: bird shakes head from right to left many times in a row (behavior observed in the female

GWM)� Swings front of body: the bird sits on a branch, extends neck, and swings the front of body from right

to left in rhythm, many times in a row (behavior observed in the female GWM)

(continued )

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ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 195

TABLE 1

(Continued )

Comfort� Preening: the bird cleans and arranges feathers using beak� Beak cleaning: the bird cleans the inside of beak with feet� Beak scrubbing: the bird vigorously scrubs beak on a branch� Feet cleaning: the bird cleans feet with beak� Scratches: the bird scratches himself or herself with foot� Bites himself or herself: the bird bites body and vocalizes� Stretches: the bird stretches down one foot and the wing of the same side� Wings up: the bird raises wings without flapping� Shakes himself or herself: the bird shakes and fluffs feathers� Takes a bath: the bird sits in water and makes head and wing movements to clean himself or herself� Yawns: the bird opens beak wide and inhales deeply� Defecates: the bird discharge feces from the body

Social behavior

Affiliative behaviors� Allopreening: preening of the partner� Asking for preening: the bird leans toward partner and presents nape of neck� Goes toward: the bird goes toward partner, head forward, wings spread� Play together: both birds hold the same rope and pull together in opposite directions� Beak contact: the birds hold each other by their beaks� Slight peck: the bird gives a slight peck on partner’s head or beak� Bows: the birds leans head quickly a few times toward partner with high-pitched vocalization, then

turns a few times looking at partner� Hide together: the birds go together to a small cavity in a trunk, stay there for some time, then go out

of it together

Reproductive behaviors� Regurgitates: the bird regurgitates in partner’s beak, with spasmodic movements� Copulation: the male is on the female’s back and they make cloacal contact� Nest inspection: the bird goes in the nest

Agonistic behaviors� Rises: the bird rises and spreads wings a little, with fan-shaped tail, beak wide open, and sometimes

piercing vocalizations� Feet up: the bird lifts a foot at chest height toward the other, stretches neck, feathers fluffed or not,

often groaning� Wings spread: the bird spreads wings, head feathers fluffed and pupils contracted� Rushes: the bird fluffs feathers, spreads wings slightly, and walks fast toward the other with head

down and open beak� Strong peck: the bird gives strong pecks to the other, often both birds vocalize� Small peck: the bird gives a slighter peck to the other, both birds grunt� Peck exchanges: both birds give pecks to each other, beaks are wide open, both birds grunt or vocalize� Escapes: one of the birds rushes away when the other approaches� Shudders: the bird trembles convulsively� Stiffens: the bird flattens feathers, retracts neck, and stiffens

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the training. Stage 6 (attained only by the Buffon’s macaws) corresponded to

the period after the end of the training program.

The number of visitors passing was calculated as monthly averages based

upon the numbers recorded each day during the three observation sessions. The

differences among the average frequencies of the behaviors across training stages

and also in relation to the monthly averages for visitors were analyzed using the

Kruskall-Wallis test. If significant variations had been noted with the Kruskall-

Wallis test, these were then compared two by two with the test of Permutation

(StatXact software).

RESULTS

Training Stages

We performed 134 training sessions of 10 min each per bird for the BM

(15 weeks) and in 160 sessions (18 weeks) for the GWM. The female GWM

took food from the trainer’s hand from the first session and the male BM from

the second session. The female BM took food reliably from the hand from

Session 19. The male GWM only regularly accepted hand-feeding beginning in,

and continuing from, Session 33. Twenty-one sessions were necessary for the

female GWM to join the trainer near the perch, 24 for the male BM, 28 for the

female BM, and 52 for the male GWM (Table 2).

The male GWM systematically produced distress calls at the entry of the

trainers into the aviary during the first 3 weeks of training. The other birds

emitted distress calls only at the first entry. Only the male GWM and the

female BM expressed fearful body postures. The male BM displayed threatening

postures during the first 10 sessions, tending to precipitate toward the trainer.

We also noted conflicts between the birds: macaws not being rewarded during

training often stole or tried to steal their partner’s food. These conflicts could

impair the training. To limit as much as possible the stress linked to the training,

however, we preferred not to separate the members of a pair during the training

sessions.

The male BM learned to accept the massage with pressure on the sides of

the cloaca in 124 sessions (Table 2). The female BM learned to accept the

massage in 114 sessions and the introduction of a speculum into the cloaca in

134 sessions. She also learned to introduce her head into a mask for anesthesia

and to hold it inside for 1 min, while also accepting the light pressure of the

trainer’s hands on her body, in 122 sessions (Table 2). The male GWM learned to

accept touches of his wings in 143 sessions (Table 2). The female GWM learned

to accept the massage in 147 sessions and to put her head in the anesthesia mask

in 129 sessions (Table 2). For the GWM, the male never accepted massages, and

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TABLE 2

Number of Sessions for Engaging and for Validating a Stage for

Each Individual of the Two Pairs of Macaws: Buffon’s Macaw (BM)

and Green-Winged Macaws (GWM)

♂ BM ♀ BM ♂ GWM ♀ GWM

Stages of Training IS CS IS CS IS CS IS CS

Stage 1: Accepting food and going

near the perch

1 24 1 28 1 52 1 21

Stage 2: Going up the perch 25 45 29 45 53 87 22 65

Stage 3: Touch of beak 46 108 46 107 88 133 66 129

Stage 4: Touch of wing 69 108 79 107 110 143 104 129

Stage 5(a): Massage 109 115 110 114 129 147

Stage 5(a): Pressure on the cloaca 117 124

Stage 5(a): Speculum in the cloaca 131 134

Stage 5(b): Head inside the mask 85 104 129

Stage 5(b): Remain inside 99 107 129

Stage 5(b): Accept the touch 114 122

Note. IS (Initial Session): session number during which the corresponding order was given

for first time; CS (Criterion Session): session number during which the behavior was regarded as

learned according to the preset criteria (three orders successively obeyed during a session, for three

consecutive sessions).

training did not reach the stage of introducing the speculum into the female’s

cloaca.

The male BM had the highest participation of any subject throughout the

experiment and did not significantly vary (p > .05) according to the training

stages (Figure 1). The participation of the female BM was weak during the first

stages but increased by 150% between Stages 1 and 2 (p < .0001) and did not

significantly vary thereafter (Figure 1).

By comparison, participation of the male GWM was rather poor throughout

the program and did not significantly vary according to the training stages

(Figure 1). For the female GWM, the participation was high at the beginning of

the program and then decreased significantly (p D .048) between Stages 2 and

3, presumably when the physical contact criterion was introduced (Figure 1).

During training, aggressive behaviors toward trainers were sometimes ob-

served. The male BM sometimes rose up and raised his leg toward the trainer

when the visual signal of the touch was given. The reactions to the touch most

frequently performed were quivering and sometimes, in the case of GWM,

leaving the perch. The female GWM showed threats or fear when physical

contact was introduced to training; she often ruffled her head feathers (threat or

fear or both) when the visual signal of the touch was given. Fortunately, serious

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198 LEBLANC ET AL.

*p Kruskall Wallis.

FIGURE 1 Variation of average participation of each bird according to the training stages.

First stage: Approach the trainer, taking food in the hand and approach the perch. Second

stage: Go up the perch. Third stage: Accept to be touched on the beak. Fourth stage: Accept

to be touched on the wings, the back, the tail, and the cloaca. Fifth stage: Accept the massages

and for female the introduction of the speculum. BM D Buffon’s macaws, GWM D green-

winged macaws.

aggression in response to physical contact by the trainer (measured as attempts

to bite) was minimal: male BM (five total), female GWM (one).

Behavioral Data Recorded Outside Training Sessions

For the male BM (Figure 2a), all behavior categories, except locomotion, varied

significantly among training stages. Vigilance decreased between Stage 0 and

Stage 5 (p D .072) as did stereotypic behaviors (p < .0001) even though they

increased again between Stages 5 and 6 (p D .021), and an increase in social

behaviors was observed from Stage 0 to Stage 5 (p D .0002). The female BM

(Figure 2b) never presented any stereotypic behavior. Her vigilance increased

between Stages 0 and 1 (p D .049) and then decreased between Stages 1 and

5 (p D .001). Foraging activity increased between Stages 0 and 3 (p D .02),

then decreased between Stages 3 and 5 (p D .048). We observed an increase of

social activity (similar to the male BM) from Stage 0 to Stage 5 (p D .0001).

No variation of social activity behavior was observed in the GWM during

the training process. For the female, even though vigilance varied significantly

during the program, no general tendency was observed. The vigilance of the

male GWM (Figure 2c) increased between Stages 0 and 1 (p D .011) and then

remained at a relatively high level during all stages of training but decreased

between Stages 1 and 4 (p D .014). The male GWM never expressed stereotypic

behaviors. The stereotypic behaviors of the female GWM (Figure 2d) increased

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*p

<.0

5.

**

p<

.01

.*

**

p<

.00

1.

(a)

FIG

UR

E2

(a,b

);(c

,d).

Co

mp

aris

on

of

the

aver

age

freq

uen

cies

of

the

beh

avio

ral

cate

go

ries

for

each

stag

e,(a

)fo

rth

em

ale

BM

,(b

)fo

rth

e

fem

ale

BM

,(c

)fo

rth

em

ale

GW

M,

(d)

for

the

fem

ale

GW

M.

Fir

stst

age:

Ap

pro

ach

the

trai

ner

,ta

kin

gfo

od

inth

eh

and

and

app

roac

hth

ep

erch

.

Sec

on

dst

age:

Go

up

the

per

ch.

Th

ird

stag

e:A

ccep

tto

be

tou

ched

on

the

bea

k.

Fo

urt

hst

age:

Acc

ept

tob

eto

uch

edo

nth

ew

ing

s,th

eb

ack

,th

eta

il,

and

the

clo

aca.

Fif

thst

age:

Acc

ept

the

mas

sag

esan

dfo

rfe

mal

eth

ein

tro

du

ctio

no

fth

esp

ecu

lum

.B

MD

Bu

ffo

n’s

mac

aws,

GW

MD

gre

en-w

ing

ed

mac

aws.

(co

nti

nu

ed

)

199

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*p

<.0

5.

**

p<

.01

.

(b)

FIG

UR

E2

(Continued

).

200

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*p

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5.

(c)

FIG

UR

E2

(Continued

).

201

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*p

<.0

5.

**

p<

.01

.*

**

p<

.00

1.

(d)

FIG

UR

E2

(Continued

).

202

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ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 203

TABLE 3

Variations of the Monthly Average of the Number of Visitors Spending

Each Day in Front of the Macaws’ Birdcage Into 60 Min of Observation

Months Monthly Average of the Daily Number of Visitors ˙ SD

February 22 ˙ 3,21

March 27,05 ˙ 4,08

April 125,76 ˙ 12,77

May 108,42 ˙ 11,69

June 100,94 ˙ 10,26

between Stages 2 and 3 (p D .0005). Behaviors associated with reproduction

were observed in the pair of BM (Figure 3). They increased significantly between

Stages 0 and 5 (p D . 044) for the male and between Stages 0 and 4 (p D .026)

for the female. Very few behaviors that could be associated with reproduction

were observed in the pair of GWM (Figure 3).

The number of visitors (Table 3) passing each day in front of the macaws’

aviaries increased significantly from March to April (p < .0001) and then did

not vary significantly from April to the end of the training program.

DISCUSSION

Our training method using operant conditioning with positive reinforcement

(food) was effective in teaching macaws to accept increasing levels of handling

in a few months, with the aim of training behaviors that allow the collection of

the semen in males and AI of females. The behavioral observations suggest that

the training done here was not a significant source of stress for the 2 birds who

completed the entire training regimen. We found the following in both birds:

1. A decrease in, or no, stereotypic behaviors;

2. No increase or a decrease in vigilance after Stage 1;

3. An increase in social behaviors; and

4. No significant perturbation of other behaviors.

Although the first stage (approach—take from hand) was short for the 2 oldest

subjects (male BM and female GWM), it was longer and was associated with

behaviors related to fear and stress for the 2 youngest macaws (female BM and

male GWM). We observed important individual differences for the behaviors

learned as well as for the behaviors expressed outside the sessions during the

training program. These differences could be related to many factors such as

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*p

<.0

5.

FIG

UR

E3

Co

mp

aris

on

of

the

aver

age

freq

uen

cies

of

the

rep

rod

uct

ive

beh

avio

rsfo

rea

chst

age

for

each

of

the

4su

bje

cts.

BM

DB

uff

on

’s

mac

aws,

GW

MD

gre

en-w

ing

edm

acaw

s.

204

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ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 205

species, age, and life experience (rearing conditions—parents-reared vs. hand-

reared). These differences show the necessity of adapting the training to each

individual. We may have moved too fast to the physical-contact stages with the

female GWM; her participation decreased when the physical-contact criterion

was introduced. Because she often left the perch at the sight of the visual signal

preceding the touch, we should probably have slowed down the stage where

touch was introduced and added more reinforcement value for holding still

while food was offered and eaten. In this case, not requiring physical contact

for each training session may have helped to maintain the bird’s participation.

For the male BM, the first stage (approach—take from hand) was very quickly

acquired in spite of his not being familiar with hand-feeding before training

commenced. A long experience of captivity, feeding by caretakers, and human

proximity could explain this fast learning process. The participation of this male

was also very good for all the remaining training stages. The male BM learned

to accept the massage with pressure on the sides of the cloaca; however, no

semen collection was performed because the trainer was not experienced in the

AI process. Semen collection will be the project’s next step.

For the female BM, the first stage was much slower than for the male.

This could be related to the young age of the subject and less habituation to

human proximity. Nevertheless, her participation increased from the beginning

of the second stage and remained excellent without significant variation for the

remainder of the training stages. This female learned to accept the massage and

the introduction of a lubricated speculum into the cloaca. She also learned to

introduce her head into the anesthesia mask, to keep this position for a few

seconds, and to accept a light pressure of the trainer’s hands on her body during

this time.

The female GWM was the quickest of all subjects to go near the trainer and

take food from the hand. This could be explained by the relatively old age of

the subject (22 years) and a long experience with human proximity. However,

we observed a drop of the participation between Stages 2 and 3. That may be

related to the introduction of the touches at Stage 3. Before her acquisition by the

Ménagerie, this bird was kept by a private collector. This could have influenced

her behavior during the training. The influence of negative handling by humans

on later behavioral responses of animals is described by Breuer, Hemsworth,

and Coleman (2003). Hemsworth, Coleman, Cox, and Barnett (1994) discuss

the generalization of responses to other humans for the same stimulus. The

female GWM learned to accept some massage and to introduce her head into an

anesthesia mask; however, because of her behavior during the training (in many

sessions leaving the perch at the sight of the visual signal indicating to the bird

that she would be touched), the following training stages were not reached.

The first stage (approach—take from hand) was very long for the male GWM,

and his participation was weak during all stages of training. This could be

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206 LEBLANC ET AL.

related to his lack of socialization with humans. He seemed to be a fearful

bird who, at the beginning of the program, was startled by the clicker’s sound.

In addition, he was particularly sensitive to the presence of different trainers

(screaming and refusing at every change to participate in the training) and to

many possible sources of disturbances, such as the vocalizations of other macaws

or the presence of noisy visitors. He learned to accept a touch of his wings but

did not proceed beyond this step.

None of the following behavioral variations indicative of chronic stress were

detected in the male BM: alteration of exploratory, feeding, and reproductive

behaviors; locomotion; or an increase in apathy, vigilance, aggression, and

stereotypic behaviors (Morgan & Tromborg, 2007; Swaisgood, 2007). Moreover,

the reduction of stereotypic behaviors shown by the male throughout the program

indicates a possible improvement in the welfare of the bird; this may suggest that

the training represented a behavioral enrichment (Mason et al., 2007). However,

the observations of the BM were conducted just after their installation in the

aviary; this could have generated additional stress in the male; thus, the decrease

of stereotypic behaviors observed during the training could also be explained

by the progressive habituation of the bird to his new environment. The increase

in stereotypic behaviors recorded after Stage 5, when training had stopped,

supports the assumption that training could be considered a form of behavioral

enrichment. Nevertheless, the last increase of stereotypic behaviors could also

be linked to the end of the reproductive season (which was concomitant with

the end of the training program); this could also lead to an increase in inactivity

and, thus, boredom. Because this increase is very small, it also may not be

biologically relevant (even if it is statistically significant).

Concerning the female BM and the male GWM, increased vigilance (as

defined by the behaviors described in Table 1) was recorded for these 2 subjects

between the period preceding the start of training and the first stage and may

constitute a sign of stress (Morgan & Tromborg, 2007; Swaisgood, 2007), which

could be related to the initiation of the training schedule. This assumption

is supported by the weak participation and the fearful behaviors that these

birds expressed in the first stage of training sessions. However, the female

BM’s vigilance decreased from Stage 1 to Stage 5 with a parallel increase in

participation. Reduction in foraging behavior recorded from Stage 4 to the end

of the program may be related to the bird’s increased participation and the fact

that many food rewards were therefore consumed during the training sessions. In

this context, and in the absence of other corollary signs of behavioral changes,

this variation is probably not indicative of a stress response. For this bird, some

initial increase in stress was probably related to the first approaches of the trainer

and the trainer’s presence in the aviary. On the other hand, for the male GWM,

the relatively high level of vigilance and the weak participation during all stages

of training suggest a more chronic stress response.

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ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 207

For the female GWM, the increase of stereotypic behaviors observed during

the training process is likely to indicate elevated stress, according to Mason

et al. (2007); the increase seemed to be in direct response to the introduction of

the first touches of the beak. This increase of stereotypies, combined with the

fall of participation in the training observed between these two stages, supports

the assumption of increased stress in relation to the handling carried out during

training. However, Stage 3 began on April 7 and continued until May 7 for the

female GWM, when the monthly average number of visitors passing each day

in front of the macaws’ aviaries increased significantly. It is well known that the

visitors can also constitute a source of stress in captivity (Morgan & Tromborg,

2007).

The behaviors of the BM associated with reproduction increased during the

training program and then decreased after the training was stopped. However,

macaw reproduction is seasonal and the reduction in reproductive behavior

(recorded after the end of the training program for the male and starting from

the first massages for the female) could indicate the end of the season. In

contrast, the reproductive behaviors of the GWM did not develop over the

period. The reproductive behaviors of the male recorded during Stage 2 were

related to inspection of the nest in the week following its installation inside the

aviary. In this context, these inspections could be more related to exploratory

behavior than to reproductive behavior. Although the absence of reproductive

behavior in the GWM could indicate chronic stress, other explanations are

also possible. Large macaws are known to reach their sexual maturity only

around the age of 4 to 5 years (Quemin, 2003): the 3-year-old male was

probably not sexually mature, and the 22-year-old female had never reproduced.

The female’s rearing history was not known; hand-rearing or an absence of

socialization with conspecifics during maturation are known to have detrimental

effects on reproduction (Schmid, Doherr, & Steiger, 2005). Hand-rearing is one

of the possible causes of the difficulties in macaws reproducing in captivity

(Derrickson & Snyder, 1992). Finally, social familiarity and sexual compatibility

are important determinants of the parrots’ pairing process (Derrickson & Snyder,

1992). Thus, the behavioral incompatibility of the partners may also explain the

failure of these subjects’ reproductive behaviors to develop; in this case, the AI

process would probably be necessary.

CONCLUSION

Despite interindividual variation in responses to the training program, we suc-

ceeded in 15 weeks to teach 2 BM to accept various handling procedures

necessary for AI. In less than 4 months, the female accepted the introduction

of the speculum into the cloaca. This suggests that with additional training,

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208 LEBLANC ET AL.

the complete voluntary application of the insemination procedures is feasible

without having to use restraint or gaseous anesthesia. Moreover, the training

did not generate chronic stress, and the birds’ reproductive behaviors developed

over the period, so they were not disturbed by the training. Concerning the

2 GWM, the training produced some positive results; however, in particular

for the male, they were still insufficient with regard to the AI process. For the

female, behavioral indicators revealed signs of stress likely to be linked not

only with the training but also with other sources of stress in captivity such as

the bird’s individual experience or proximity of visitors. Extending the birds’

training periods should be considered.

Therefore, our experiment shows that it is possible to conduct AI procedures

by using training instead of traditional techniques (restraint or anesthesia). As

long as the birds are compatible with human interaction, this procedure offers

the added benefits of being not only a source of behavioral enrichment that may

improve the welfare of some birds but also a key element of assisted reproductive

approaches.

ACKNOWLEDGMENTS

This study was hosted at the Ménagerie du Jardin des Plantes of MNHN

at Paris, France. We thank the director, Jacques Rigoulet, for his hospitality

and zookeepers Betty Knopf, Marie Vergues, and David Buchemeyer for their

assistance in bird managing. We also thank Sarah-Jane Vick and anonymous

reviewers for their suggestions to improve the manuscript. The experiments

comply with the French laws concerning animal care.

REFERENCES

Altmann, J. (1974). Observational study of behaviour: Sampling methods. Behaviour, 34, 227–267.

Barnett, J. L., Hemsworth, P. H., Hennessey, D. P., McCallum, T. H., & Newman, E. A. (1994). The

effects of modifying the amount of human contact on behavioural, physiological, and production

responses of laying hens. Applied Animal Behaviour Science, 41, 87–100.

BirdLife International. (2010). BirdLife: State of the world’s birds. Retrieved from http://www.bird

life.org/datazone/species/index.html

Boyd, L. L. (1978). Artificial insemination of falcons. Symposia of the Zoological Society of London,

43, 73–80.

Breuer, K., Hemsworth, P. H., & Coleman, G. J. (2003). The effect of positive or negative handling

on the behavioural and physiological responses of nonlactating heifers. Applied Animal Behaviour

Science, 41, 87–100.

Brock, K. M. (1991). Semen collection and artificial insemination in the Hispaniolan parrot. Journal

of Zoo and Wildlife Medicine, 22, 107–114.

Caïn, J. R. (1978). Artificial insemination: A practical method for genetic improvement in ring-

necked pheasants. Symposia of the Zoological Society of London, 43, 81–88.

Dow

nloa

ded

by [

Dr

Ken

neth

Sha

piro

] at

13:

35 0

2 N

ovem

ber

2015

ARTIFICIAL INSEMINATION PROCEDURES FOR MACAWS 209

Curio, E. (1998). Behaviour as a tool for management intervention in birds. In T. Caro (Ed.),

Behavioural ecology and conservation biology (pp. 163–191). Oxford, UK: Oxford University

Press.

Derrickson, S. R., & Snyder, N. F. R. (1992). Potentials and limits of captive breeding in parrot

conservation. In S. R. Beissinger (Ed.), New world parrots in crisis: Solutions from conservation

biology (pp. 133–153). Washington, DC: Smithsonian Institution.

Dunning, J. B. (2008). CRC handbook of avian body masses (2nd ed.). Boca Raton, FL: CRC.

Garner, J. P., Meehan, C. L., Famula, T. R., & Mench, J. A. (2006). Genetic, environmental, and

neighbour effects on the severity of stereotypes and feather picking in orange-winged Amazon

parrots (Amazona amazonica): An epidemiological study. Applied Animal Behaviour Science, 96,

153–168.

Garner, J. P., Meehan, C. L., & Mench, J. A. (2003). Stereotypies in caged parrots, schizophrenia

and autism: Evidence for a common mechanism. Behavioural Brain Research, 145, 125–134.

Gee, G. F., Bertschinger, H., Donoghue, A. M., Blanco, J., & Soley, J. (2004). Reproduction in

nondomestic birds: Physiology, semen collection, artificial insemination and cryopreservation.

Avian and Poultry Biology Review, 15, 47–101.

Grajal, A. (2000). The Neotropics (Americas). In N. Snyder, P. McGowan, J. Gilardi, & A. Grajal

(Eds.), Parrots: Status survey and conservation action plan 2000–2004 (pp. 98–128). Cambridge,

UK: International Union for Conservation of Nature.

Harrison, G. J. & Wasmund, D. (1983). Preliminary studies of electroejaculation to facilitate manual

semen collection in psittacines. Proceedings of the Association of Avian Veterinarians Annual

Conference (p. 207). Krakow, Poland: Wyeclawska Dyurkarnia, Naukowa.

Heidenreich, B. (2004). Training birds for medical and husbandry behaviors. Proceedings of the As-

sociation of Avian Veterinarians Annual Conference (pp. 299–305). New Orleans, LA: Association

of Avian Veterinarians.

Heidenreich, B. (2007). An introduction to positive reinforcement training and its benefits. Journal

of Exotic Pet Medicine, 16, 19.

Hemsworth, P. H., Coleman, G. J. C., Cox, M., & Barnett, J. L. (1994). Stimulus generalization:

The inability of pigs to discriminate between humans on the basis of their previous handling

experience. Applied Animal Behaviour Science, 40, 129–142.

Jones, K. L., & Nicolich, J. M. (2001). Artificial insemination in captive whooping cranes: Results

from genetic analyses. Zoo Biology, 20, 331–342.

Juniper, T., & Parr, M. (1998). Parrots: A guide to the parrots of the world. Sussex, UK: Pica.

Mason, G., Clubb, R., Latham, N., & Vickery, S. (2007). Why and how should we use environmental

enrichment to tackle stereotypic behaviour? Applied Animal Behaviour Science, 102, 163–188.

McCall, C. A., & Burgin, S. E. (2002). Equine utilization of secondary reinforcement during response

extinction and acquisition. Applied Animal Behaviour Science, 78, 253–262.

Morgan, K. N., & Tromborg, C. T. (2007). Sources of stress in captivity. Applied Animal Behaviour

Science, 102, 262–302.

O’Brien, J. K., Oehler, D. A., Malowski, S. P., &, Roth, T. L. (1999). Semen collection, characteri-

zation, and cryopreservation in a Magellanic penguin. Zoo Biology, 18, 199–214.

Quemin, I. (2003). Le comportement des psittacidés et ses troubles [Psittacids’ behaviour and

disorders] (Hèse de doctorat Vétérinaire, University of Créteil, France).

Saint Jalme, M. (2002). Endangered avian species captive propagation: An overview of functions

and techniques. Avian Poultry Biology Review, 13, 187–202.

Saint Jalme, M., Gaucher, P., & Paillat, P. (1994). Artificial insemination in Houbara bustards

(Chlamydotis undula): Influence of the number of spermatozoa and insemination frequency on

fertility and ability to hatch. Journal of Reproduction and Fertility, 100, 93–103.

Saint Jalme, M., Lecoq, R., Seigneurin, F., Blesbois, E., & Plouzeau, E. (2003). Cryopreservation

of semen from endangered pheasants: The first step towards a cryobank for endangered species.

Theriogenology, 59, 855–888.

Dow

nloa

ded

by [

Dr

Ken

neth

Sha

piro

] at

13:

35 0

2 N

ovem

ber

2015

210 LEBLANC ET AL.

Samour, J. H., Smith, C. A., Moore, H. D., & Markham, J. A. (1986). Semen collection and

spermatozoa characteristics in budgerigars. Veterinary Record, 118, 397–399.

Schmid, R., Doherr, M. G., & Steiger, A. (2005). The influence of the breeding method on the

behaviour of adult African grey parrots (Psittacus erithacus). Applied Animal Behaviour Science,

98, 293–307.

Snyder, N., McGowan, P., Gilardi, J., & Grajal, A. (2000). Parrots: Status survey and conservation

action plan 2000–2004. Gland, Switzerland: International Union for Conservation of Nature.

Straub, J., Forbes, N., Pees, N., & Krautwald-Junghanns, M. E. (2003). Effect of handling-induced

stress on the results of spectral Doppler echocardiography in falcons. Research in Veterinary

Science, 74, 119–122.

Sutherland, W. J. (1998). The importance of behavioural studies in conservation biology. Animal

Behaviour, 56, 801–809.

Swaisgood, R. R. (2007). Current status and future directions of applied behavioural welfare and

conservation. Applied Animal Behaviour Science, 102, 139–162.

Dow

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ded

by [

Dr

Ken

neth

Sha

piro

] at

13:

35 0

2 N

ovem

ber

2015