Capture, health, and morphological assessment of free-ranging mantled howler monkeys (Aloutta palliata) in Nicaragua. G. Peter, Parke-Davis Research Ann Arbor, MI, R. Sohn, Wildlife Veterinary Consulting, Salt Lake City, UT, and L. Winkler, University of Pittsburgh, Titusville, PA

ABSTRACT

This paper describes the preliminary results of an inter-disciplinary project on free-ranging mantled howler monkeys on Ometepe Island in Lake Nicaragua. This project is unique in being the first in Nicaragua to assess the health of these biogeographically isolated monkeys. Eleven mantled howler monkeys (10 adults - 5F, 5M and 1 subadult F) in a group composed of 13 adults were captured, marked and released over a 4 day period in late July, 1998. Capture was affected with the PneuDart©Γ capture system delivering 120 mg of a combination of tiletamine and zolazepam (~20 mg/kg). Unconsciousness occurred rapidly (1-4 minutes) with a mean recovery time of 2.53 ± 0.56 hours. During the period of restraint, each animal was given a thorough physical examination, blood feces, and hair obtained, and information gathered on temperature, heart and respiration rates, body weight, morphology (sitting height, tail length, upper and lower limb components (Schultz 1929)), dental morphology, and estimates of age and reproductive status. Overall the health of these animals was excellent. Mean body weights were 4.86 ± .34 (females) and 6.23 ± .10 (males). Palpation indicated three females were pregnant. Hematology performed in the field revealed median values for WBC (6,660), RBC (4.09x106), and PCV (40%). ELISA for measles and IFA for EEE, VEE, yellow fever, dengue, herpes plattyrhinus, and leptospirosis indicated no exposures. Plasma, serum, blood smears and clots were retained for later analyses (DNA isolation, serum chemistries, differential blood counts, blood parasites exam). Feces were also collected in 10% formalin for parasitic ova examination and Giardia antigen detection. Results of these analyses will be reported. Our results will be compared and discussed in relation to other capture projects from other locales (Glander et al., 1991).

MATERIALS AND METHODS

A base of operations was established near the home range of the troop. Care was taken to locate a spot where there was shade throughout the day and breezes so as to keep both the animals and staff cool. A processing area was established where all the equipment and supplies could be arrayed and organized to facilitate rapid and efficient conduct of the dart preparation, sample collection, measurement, and tagging procedures.

Darting procedure - Monkeys were captured with the use of a PneuDart' CO2-powered rifle delivering a dart loaded (Click here to see Figure 1) with 120 mg of Telazol" (tiletamine and zolazepam). We estimated the dose to be equivalent to ~20 mg/kg. A net team of 4-6 individuals accompanied two dart teams consisting of a shooter and a spotter (Click here to see Figure 2). Once a desired troop member was identified, the dart team would observe the animal until a safe shot was presented (hindquarters visible and vital areas preferably obscured by limbs or facing away). The spotter's responsibility was to watch the flight of the dart to assure a hit and the subsequent movement of the monkey. The timing of anesthetic events began (see Table 1) with the shot. Once the monkey came to rest, the dart teams directed the immediate and accurate placement of the large hammock net by observing the possible path of the monkey's descent from several angles. Patience and an occasional fortuitous wind or shaking of limbs were needed until the monkey fell into the net.

Examination and sample collection - Once on the ground, the animal was immediately examined by a veterinarian who ausculted the thorax and assessed the subject for possible trauma. All personnel directly handling the monkeys wore latex exam gloves and a mask to protect both parties. The monkey was rushed immediately to the staging area on the beach, a run of only a few minutes. Ophthalmic ointment was placed in the eyes to prevent corneal drying. A complete physical examination was performed at this time (Click here to see Figure 3). Heart and respiration rates and body temperature were monitored periodically through out the ensuing collection and tagging procedures. If the body temperature of the animal exceeded 103o F they were immersed in the lake (Click here to see Figure 4). Personnel rotated to the table to perform assigned procedures, commonly more than one at a time to assure the most information could be gathered efficiently. Body weights were obtained with the use of a net and a Pesola" 20 kg scale (accurate to 0.2 kg) suspended from a tree (Click here to see Figure 5). Morphometric measurements were taken with a Helios" needle point calipers (accurate to 0.01 cm) and a cloth tape (Click here to see Figure 6). Dental examination to estimate age was performed prior to acquiring latex dental casts. Feces were obtained directly from the rectum, placed in 100% ethanol, and handled to prevent contamination by foreign DNA. Additional feces were placed in 10% buffered formalin and an anal tape was taken for parasitological examinations. Blood samples were drawn by femoral venipuncture (Click here to see Figure 7) and placed in both EDTA and clot tubes in an ice chest. Dart injection sites were identified and assessed for trauma. Each animal received an intramuscular injection of 50,000 U/kg of penicillin G benzathine/penicillin G procaine (Ambipen") as a prophylactic therapy. Colorful nylon dog collars perforated with several heat-seared holes approximately 1" apart along their length were carefully sized by cutting to length and affixed using a pop rivet tool to the necks of females and ankles of males (the large hyoid apparatus in males precluding neck placement). After sizing, the frayed ends of the collars were melted with a lighter flame to prevent unraveling. In addition, metal ear tags were placed in the outer edge of the pinna. Following these procedures, the animals were returned to capture site and observed until recovery from anesthesia defined as the ability to climb into the understory.

Sample handling and analysis

In the field - White blood cell (WBC) and red blood cell (RBC) counts were performed using a Unopette" microcollection system and a stage hemocytometer. Packed cell volume (PCV) was obtained by centrifuging a heparinized capillary tube. Both thin and thick blood smears were made and fixed by air-drying following immersion in 100% methanol. Plasma and serum samples were obtained by centrifugation of the EDTA and clot tubes, respectively, and the remaining blood clots were saved. Samples were frozen for transport.

In the lab - Differential white blood cell counts were performed by hand from thin blood smear slides stained with azure B/methylene blue/eosin Y and clinical chemistry analyses were performed on a Vitros" chemistry analyzer, both by the Clinical and Molecular Pathology Laboratory, Parke-Davis1. Microscopic examination of the thick smears for blood parasites and the feces for parasite ova and the performance of an ELISA for giardia antigen in the feces were performed by the Animal Health Diagnostic Laboratory2. Serologic analyses for exposure to viral and bacterial disease were performed through the services of MA Bioservices3.

RESULTS

Darting - With practice the teams became very efficient. Monkeys were safely caught in the net after the first day. Only one animal experienced an unbroken fall after running through the troop and confusing observers. It sustained no injuries in the fall. The weight-adjusted doses of Telazol" delivered, assuming each injection of 120 mg was entire, are reported in Table 1. The mean dose was 23.6 ± 6.3 mg/kg (n=11) for all animals and 22.0 ± 3.3 mg/kg (n=10) for adult animals. Unconsciousness occurred very rapidly within 1 - 4 minutes, but the time from injection to netting (T1) averaged 12.7 ± 10.1 minutes. The average time to recovery as defined as the ability to climb (T2) was 150.5 ± 35.4 minutes for animals receiving an initial dose only (n=9). Two animals (#6 and #11) receiving an additional 40 mg of Telazol" recovered in 190 and 266 minutes. Only one monkey was darted which was not retrieved from the canopy. This was a subadult that received a glancing shot and did not receive a full dose. The only injuries noted were small lacerations at the dart injection site of which the larger few were cleansed with betadine and alcohol and sutured with an absorbable suture using a subcuticular pattern.

Examination, measurements and analysis of samples - The physical examination revealed animals in very good condition with adequate body fat. Palpation of the abdomen revealed three possibly pregnant females (see Table1). Other physiologic data are summarized and reported in Table 1. Three animals, including both of the animals receiving supplemental doses, attained body temperatures above 1030 F and were immediately immersed in the lake (Figure 4). Hematology and clinical serum chemistry values are summarized in Tables 2 and 3. ELISA for measles and IFA for Herpes platyrrinae, Herpes saimirii, eastern and Venezuelan equine encephalitis, yellow fever, dengue, and leptospirosis were negative. Fecal and anal tape examinations for internal parasite ova revealed all animals carried an unspeciated pinworm. External parasitism was limited to only a couple of unidentified ticks and, as a likely result of mutual grooming, there was evidence of louse eggs on the fecal exam. Morphological measurements are summarized in Table 4. Blood and fecal DNA analyses are reported by Dr. Linda Winkler in Poster # 3.

Table 1: Anesthetic & Physiologic Data

#

Sex

Wt(kg)

Initial Dose

T1

T2

Heart Rates

Body Temp. Range

Respiration Rate

Pregnancy Status

(mg/kg)

(min)

(min)

(Beats/min)

(° F)

(Breaths/min)

1

E

4.5

26.7

20

148

210, 211

99

N.R.

-

2

G

5.8

20.7

30

129

138, 180

101.3-101.4

N.R.

n/a

3

E

5.4

22.2

3

165

136, 136

99.8-100.7

N.R.

+

4

E

3

40 **

23

136

132

100.3-100.8

24

subadult

5

G

6.3

19.0

5

196

116, 120

101.9-102.3

24

n/a

6

E

5

24.0

4

190 *

128

103.2-103.9

N.R.

+

7

G

6.8

17.6

5

190

100, 100

100.8-101.0

18

n/a

8

E

4.9

24.4

N.R.

120

148, 150

103.9

36

+

9/10

E

4.5

26.7

4

90

124, 140

101.0-101.0

N.R.

-

11

G

6.2

19.4

22

266 *

178, 120

103.2-103.4

N.R.

n/a

12

G

6.2

19.4

11

181

126, 136, 144

100.9-101.0

30, 28

n/a

mean (all)

22 ▒ 3.3 **

12.7 ▒ 10.1

150.5 ▒ 35.4 *

141 ▒ 28

101.5 ▒ 1.5

26.2 ▒ 6.7

n/a

mean (G)

4.8 ▒ 0.3

mean (E)

6.2 ▒ 0.1

n =

10

10

9

11

11

5

Table 2: Hematology Results

Parameter

Mean (n=11)

S.D.

Median

Range

RBC (106/ml)

4.28

0.65

4.09

3.27-4.56

PCV (%)

40

3

40

36-45

WBC (#/ml)

6703

2226

6660

2442-10434

Neutrophil (#/ml)

2687

1014

2684

1319-5125

(%)

41

8

40

27-57

Lymphocyte (#/ml)

3048

1350

2833

879-6365

(%)

45

9

43

31-61

Monocyte (#/ml)

735

364

809

147-1265

(%)

11

5

12

4-16

Eosinophil (#/ml)

201

135

170

0-422

(%)

2.8

1.4

3

0-5.0

Basophil (#/ml)

33

42

24

0-104

(%)

0.5

0.5

0.5

0-1.0

Table 3: Serum Chemistry Results

Parameter

Mean (n=11)

S.D.

Median

Range

BUN (mg/dL)

6.7

2.1

6

4-11

Creatine (mg/dL)

1

0.2

1

0.7-1.2

Na (mEq/dL)

139

2

139

137-142

K (mEq/dL)

10.5

3.6

9.4

5.3-14

Cl (mEq/dL)

98

3

96

95-103

Ca (mg/dL)

11.6

2.3

9.9

9.2-14

P (mg/dL)

4.7

1.4

5.6

2.5-6

AST (U/L)

176

59

179

102-292

ALT (U/L)

38

14

40

11-51

TBIL (mg/dL)

0.8

0.9

0.6

0.2-3.3

ALKP (U/L)

138

158

159

10-521

GLU (mg/dL)

82

25

83

40-116

CHOL (mg/dL)

146

22

141

121-176

Total Protein (g/dL)

7.3

0.5

7.3

6.7-8.6

Albumin (g/dL)

4.7

0.5

4.5

4.3-5.9

Globulin (g/dL)

2.6

0.3

2.6

2.2-3.1

A/G ( :1 )

1

0.3

1.9

1.4-2.4

LDH (U/L)

1586

708

1294

877-3017

CK (U/L)

303

78

312

172-467

Table 4: Morphological Measurements

#

Sex

Wt(kg)

Sitting

Tail Length

Upper Arm

Forearm

Hand

Hand

Thumb

Thigh

Lower Leg

Foot

Foot

Hallux

Ht(cm)

Dermato-glyphics*

Length

Length

Length

Breadth

Length

Length

Length

Length

Breadth

Length

1

E

4.5

36.2

49.5 (20.4)

14.4

14.1

10

3.5

2.8

19.4

16.5

12

4.1

2.5

2

G

5.8

48.9

63.1 (22.8)

15.6

13.5

11.2

3.8

3.1

16.1

16.7

14.1

5.8

3.1

3

E

5.4

44.9

56.1 (21.9)

15.7

16.1

9.9

3.5

2.7

16

15.1

13.3

4.9

4.4

4

E

3

37.8

52.4 (18.6)

12.2

14.3

9.2

2.9

2.6

13.6

13

11.8

4.5

2.8

5

G

6.3

43.9

57 (21.2)

17

16.2

12.1

4.3

3.3

16.2

16.5

14.3

5.8

3.2

6

E

5

41.8

55.6 (19.6)

15

14.2

10.7

4.1

2.5

13.7

15.7

12.7

4.3

3.2

5.3

7.5

7

G

6.8

44.4

58.7 (22.6)

15.1

14.8

11.3

4.2

2.7

15.2

15.9

13.8

5.3

3.5

7.1

10.6

8

E

4.9

42

57 (21.1)

14.2

14.5

9.9

3.4

2.9

14.7

13.8

12.2

4.3

2.5

5.2

9.7

9/10

E

4.5

42.3

56.8 (23.9)

16.65

15.1

9.7

3.7

2.6

15.2

16.3

13.1

4.4

3.2

6

10.1

11

G

6.2

49

56.3 (22)

16.2

16.5

11

4

3.0

15.1

16.1

13.8

5.8

2.1

7.2

10.6

12

G

6.2

49.2

63.1 (23.7)

15.7

15.1

12

3.6

3.0

16.3

16.7

14.2

5.3

3.7

6.1

10.1

x E

4.8

40.8

54.6 (20.9)

14.7

14.7

9.9

3.5

2.7/5.5

15.4

15.1

12.5

4.4

3.1/9.9

x G

6.2

47.1

59.6 (22.5)

15.9

15.2

11.5

4

3.0/6.8

15.8

16.4

14

5.6

3.1/10.2

DISCUSSION

The goal of chemical immobilization is to effectively restrain the subject at a sufficient depth of anesthesia to allow the intended procedures while keeping them light enough to recover quickly and return safely to their habitat. Telazol is widely used in the immobilization of wild and exotic species (Schobert 1987) especially nonhuman primates (Eads 1976). Previous captures of howling monkeys (Crissey&Edwards 1989; Glander 1991) describe Telazol" as a safe and effective capture drug which provides rapid immobilization, good muscle relaxation, and sufficient duration. Rapid immobilization is important in preventing the co-mingling of the injected animal with the remainder of the troop and confounding the problem of accurately setting the net. Good muscle relaxation is a necessary characteristic in a capture agent for animals with a prehensile tail. Glander (1991) found that phencyclidine derivatives alone immobilize these animals but the prehensile tail does not relax and the monkeys remain suspended in the canopy. Telazol" has a wide safety margin as illustrated in this study by the uneventful recovery of a subadult who potentially received 40 mg/kg without adverse effects. The fact that this animal had one of the shorter recovery times, however, suggests the possibility of a partial injection.

The darting success of this project was similar to other reports of howling monkey captures (Crissey&Edwards 1989; Glander et.al. 1991). Darting became more difficult with each successive day as the animals became noticeably wary as compared to the initial relaxed behavior. The availability of untagged targets diminished as well. Injury was kept to a minimum by thorough planning, coordinated teamwork, and fortuitous luck. Only one unbroken fall occurred and one animal was darted outside the targeted hindquarters. The injection site was high on the side of the abdomen posterior to the last rib and appeared to enter only muscle thereby effecting anesthesia. The 3/8-inch needle possibly minimizes risk to internal organs.

Both of the references above describe the use of Telazol" in howling monkeys. In mantled howling monkeys Glander etal (1991) reported a mean dose of 14.8 mg/kg resulted in immobility in 60 sec and a time to fall of 212 seconds. There was no record kept of recovery duration. Crissey and Edwards (1989) reported a study in the capture of red howling monkeys, Alouatta seniculus, where a mean dose of 23.3 mg/kg induced immobilization in 1-3 minutes producing a "sleep" time of 45 minutes as defined by the return of the righting reflex. We delivered an almost identical mean dose as this later study and the results are very similar to ours although we recorded recovery as the time to climb. The return to righting occurs sooner, which accounts for the difference in T2 and their "sleep time". Some of the variability in anesthetic events within our study is likely due to both injecting animals of varying weights, age, and sex with the same amount of drug and the possibility of partial injections inherent in darting procedures. The need for supplementation in this study may have been do to this partial injection. Supplemental dosing runs the risk of hyperthermia and prolonged recovery as seen in this study. Animal #11 is an example where the increased depth of anesthesia evident from the decrease in heart rate (180«120) led to a prolonged recovery and possibly contributed to hyperthermia. Dosing with ketamine to prolong the restraint as suggested by Glander et.al.(1991) is a common procedure when using Telazol" in primate medicine, but it requires transporting another drug. In cases of prolonged recovery, reversal of the benzodiazepine portion of Telazol" (zolazepam) with flumazenil at 1 mg/20-50 mg of zolazepam administered might be considered to decrease the sedation and shorten the recovery period (Karesh et.al. 1998).

Overall, the health of these monkeys was found to be excellent. Glander (1991) reported mean body weights of males and females at 6.53 kg and 4.02 kg, respectively. This compares well to our mean of 6.2 kg for male animals but our mean of 4.8 kg for females is higher possibly because 3 of 5 adult females were pregnant. Pregnancy and the presence of a subadult female in the data set is evident in the higher variation (S.D. 0.3) for the mean of females as compared to the males (S.D. 0.1) in our study. A high pregnancy rate also argues for a healthy population. In addition, serology for detection of antibodies to the pathogens listed above indicated no exposure among these monkeys. Although the tests used in these analyses were conducted with positive and negative controls using human sera, they have been used to detect positive sera in other New World primate species (owl and squirrel monkey) (personal communication). The results reported here can be considered accurate but the sensitivity of these assays may be less than ideal as it depends on the cross-reactivity of the reagents for human immunoglobulins to those of the howling monkey. The absence of giardia antigen in the feces and evidence of malaria and other blood parasites in the blood smears also support the good health and non-exposed status of this population. Pinworms, lice and an occasional tick are parasites commonly found in wild animal populations and are not overtly pathogenic in animals as healthy as these subjects. The lack of exposure to disease of this island population of monkeys may indicate them to be at risk for future exposure as the habitat becomes constrained by development. The increased opportunity for disease transmission from humans, other species, and within their own species increases as available habitat decreases.

CONCLUSION

The capture of wild animals sometimes becomes necessary in conservation biology studies. The risk of injury or mortality to an individual is justified by the value of the information gained to serve in the conservation of the species. Therefore, it is incumbent upon any capture and release effort to gather the maximal amount of information possible when risking the lives and well-being of any wild animal. This study establishes a baseline record for this population of animals as to their health status and provides some normative data for future comparisons.

Acknowledgements

The efficiency and success of this capture effort was the result of preparation, planning, and coordination. The capture of the majority of animals in a troop without a significant injury, not to mention mortality is a goal we had hoped to achieve. This goal was achieved with the help of many. Among those we would like to thank are the Molinas family for their invaluable assistance and cordial hospitality at the Ometepe Biological Field Station. We would also like to acknowledge the hard work of the students and graduate teaching assistants in the field.

PneuDart Inc., Williamsport, PA Telazol" Fort Dodge Animal Health, Fort Dodge, Iowa 50501 Ambipen" Butler Co., Columbus, Ohio 43228 Unopette" Becton Dickinson and Co., Franklin Lanes, NJ 07417 Vitros" chemistry analyzer, Johnson&Johnson, Rochester, NY

1Clinical and Molecular Pathology Laboratory, Parke-Davis, Ann Arbor, MI 2Animal Health Diagnostic Laboratory, Michigan State University, College of Veterinary Medicine, E. Lansing, MI 48909 3MA Bioservices, Rockville, Md. 20850

References cited Bezanson, M. 1999. Positional behavior and prehensile-tail use in Aluotta palliata. American Journal of Physical Anthropolgy, Supplement From 68th Annual Meeting.

Crissey, SD and Edwards, MS. 1989. Telazol" immobilization of red howler monkeys (Alouatta seniculus) under free-ranging conditions. Annual Conference of the American Association of Zoo Veterinarians. Greensboro, NC. p. 207

Eads, FE. 1976. Tilazol' (CI-744): A new agent for the chemical restraint and anesthesia of nonhuman primates. Veterinary Medicine May Issue p.648-652

Glander, KE, Fedigan, LM, and Fedigan, L. 1991. Field methods for capture and measurement of three monkey species in Costa Rica. Folia Primatologica 57:70-82

Karesh, WB, Wallace, RB, Painter, RLE, Rumiz, D, Braselton, WE, Dierenfeld, ES, and Puche, H. 1998. Immobilization and health assessment of free-ranging black spider monkeys (Ateles paniscus chamek). American Journal of Primatology 44:107-123

Schobert, E. 1987. Telazol" Use in wild and exotic animals. Veterinary Medicine October Issue p.1080-1088

Schultz, AH. 1929. The technique of measuring the outer body of humans and of primates in general. Contributions to Embryology, Carnegie Institute 20:213-257

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