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Journal of Zoo and Wildlife Medicine 36(3): 463-469, 2005 Copyright 2005 by American Association of Zoo Veterinarians

HEMATOLOGY OF FREE-LIVING MARSH DEER (BLASTOCERUS DICHOTOMUS) FROM SOUTHEAST BRAZIL

Mat?as Pablo Juan Szab? D.V.M., Ph.D., Eliana Reiko Matushima D.V.M., Ph.D., M?rcio Botelho

de Castro D.V.M., Ph.D., Danilo Alvaro Santana D.V.M., C?tia Dejuste de Paula D.V.M., M.Sc., and Jos? Mauricio Barbanti Duarte D.V.M., Ph.D.

Abstract: This work reports basic h?matologie values of a sample of a population of free-living marsh deer (Blas tocerus dichotomus) living by the Paran? River in Southeast Brazil. H?matologie values are presented separately for

male, female, and young animals as well as for anesthetized and nonanesthetized cervids. Nonanesthetized deer re

strained by physical means had significantly higher erythrocyte indices and total leukocyte counts. Comparisons of blood parameters of anesthetized animals of different ages and gender differed slightly, with only two significant differences observed: young animals had significantly higher red blood cell counts than adult males and a lower blood total protein content when compared to adult females. Results indicate that two main reference ranges for blood values should be considered for marsh deer, for blood obtained from anesthetized or physically restrained individuals.

Key words: Anesthetized, Blastocerus dichotomus, free-living, hematology, marsh deer, nonanesthetized.

INTRODUCTION

The marsh deer (Blastocerus dichotomus), the

largest cervid of South America, is an endangered

species throughout its range. The species is adapted to wetlands and other environments prone to flood

ing, which are among the areas most degraded by

hydroelectric power stations. Originally B. dicho

tomus was widespread in South America, but it has

suffered a considerable decline with population fragmentation9 and is now considered an endan

gered species.10 In Uruguay the marsh deer is ex

tinct.9 Efforts to preserve marsh deer are ongoing, but background information about its life in the wild is needed to optimize species conservation.

Basic physiologic data, such as the reproductive cy cle of males and females, social behavior, and dis ease parameters, are lacking. H?matologie values are important in assessing the general health of in

dividuals or populations of a species. Hematology results can be used in patient evaluation, in forming

diagnoses, and to evaluate the progress of certain

disease states.4

Basic h?matologie values are reported for a sam

pie of free-living marsh deer living by the Paran?

River in Southeast Brazil that were captured before

flooding of the marsh for a hydroelectric power sta

tion. H?matologie values are presented separately for male, female, and young animals and for anes

thetized and nonanesthetized cervids.

MATERIAL AND METHODS

Location and capture

Porto-Primavera hydroelectric power station is

located in Brazil by the Paran? River (22?25'77" S; 52?58'84" W) with S?o Paulo State to the southwest and Mato Grosso do Sul State to the east. The water

reservoir filled by the Paran? River flooded a 2,200 km2 area at the S?o Paulo and Mato-Grosso do Sul

border. From May 1998 to April 2001, marsh deer were captured for an extensive research program under the financial support of S?o Paulo State En

ergy Company (CESP). Wresting (bulldogging) technique, with the aid of a helicopter, was used for

the capture of deer as previously described.3 The

capture protocol described was submitted to and

approved by the National Institute of Environment

and Natural Resources of Brazil (IBAMA, Instituto Brasileiro do Meio Ambiente e dos Recursos Re

novaveis). After capture, some of the animals were re

strained without anesthesia for sample collection

and collaring before release. Other animals were

anesthetized at the capture site and transported to a

study base by helicopter. At the base the anesthe

tized animals were monitored and given a thorough

physical examination, and samples, including blood, were taken for various research projects. Af

ter examination, animals were sent to a quarantine

From the Faculdade de Medicina Veterinaria da Univ ersidade Federal de Uberl?ndia, Minas Gerais, 38400-902, Brazil (Szab?); the Departamento de Melhoramento Ge n?tico Animal, Faculdade de Ciencias Agrarias e Veteri

narias, Universidade Estadual Paulista, Jaboticabal, S?o

Paulo, 14884-900, Brazil (Duarte); the Departamento de

Patologia Animal, Faculdade de Medicina Veterinaria e

Zootecnia, USP, Sao Paulo, 05508-900, Brazil, (Matushi ma, Dejuste); and the Curso de Medicina Veterinaria, Universidade de Franca, Franca, Sao Paulo, 14404-600, Brazil (Castro, Santana). Correspondence should be di rected to Dr. Szab?.

463

464 JOURNAL OF ZOO AND WILDLIFE MEDICINE

Station and for inclusion in an ex situ conservation

program.

Animal anesthesia

Several anesthetic protocols were tested in the

field. For the h?matologie comparisons, only the

data from animals anesthetized with acepromazine 0.05 mg/kg (Acepran 1%, Univet S.A. S?o Paulo, S?o Paulo, 01523-000, Brazil), midazolam 1.0 mg/

kg (Dormonid, Roche Brasil, Rio de Janeiro, Rio de Janeiro, 22710-104, Brazil), and ketamine 7.0

mg/kg (Dopalen, Agribands do Brasil Ltda., Pau

linia, S?o Paulo, 13140-000, Brazil) were used.

Age estimation

Animal age was estimated by dental examination

performed by the same investigator (J.M.B.D.). An

imals less than 1 yr old were classed as young. There were insufficient deer in this group to merit

division of the sample into male and female.

Blood collection

Blood samples were collected from the jugular vein using BD Vacutainers with K3-EDTA (BD Va

cutainer?, BD Brasil, Juiz de Fora, Minas Gerais,

36081-000, Brazil). Samples were refrigerated until

processing in the laboratory. Blood was collected

either immediately after capture (nonanesthetized

animals) or 15-30 min after induction of anesthe

sia. Samples for this study (n =

97) were taken

from animals before flooding of the marsh or from

animals from nonflooded areas to avoid effects on

the sample caused by environmental change.

Analysis of blood samples Blood samples of marsh deer were analyzed, and

the effects of age, gender, and anesthetic use on

blood parameters were evaluated. Total blood cell

counts, stained blood smears, packed cell volume,

hemoglobin content, and total plasma protein esti

mation were obtained within 12 hr of collection.

Red blood cell (RBC) and white blood cell (WBC) counts were obtained with a Neubauer chamber

(Neubauer Improved, Precicolor HGB, Schwimm

badweg 12, D-35398 Giessen-Luetzellinden, Ger

many). The microhematocrit method was used for

packed cell volume (PCV) determination, and the cyanmethemoglobin method was used for hemoglo bin content measurement. Differential white blood

cell counts were performed on stained blood smears

(Quick stain?Pan?tico R?pido LB?Laborclin Produtos para Laboratorio?Pinhais, Paran?,

83321-210, Brazil). H?matologie indices including mean corpuscular volume (MCV) and mean cor

puscular hemoglobin concentration (MCHC) were

calculated. Total plasma protein (TPP) was esti

mated by refractometer.4

Statistical analysis Data from deer populations grouped according to

age, gender, and anesthetized vs. nonanesthetized were analyzed either by unpaired r-test or by one

way ANO VA test followed by Tukey's Multiple Comparison Test. Differences were considered sig nificant when P values were less than 0.05.

RESULTS

Preliminary analysis showed major differences in

h?matologie profile between anesthetized and non

anesthetized animals, so data from these two groups are presented in separate tables. Nonanesthetized

adult females (physically restrained) had signifi cantly higher RBC, WBC, monocyte, neutrophil, and lymphocyte cell counts but lower basophil counts relative to anesthetized females (Fig. 1).

Nonanesthetized adult males had significantly high er red and white blood cell, mature neutrophil, and

lymphocyte counts, higher packed cell volume, and

lower basophil counts relative to adult anesthetized

males (Fig. 2). H?matologie parameters of anesthetized female,

male, and young animals were very similar: only minor differences were observed between animals

of different age or gender. Young animals had sig

nificantly higher RBC counts than adult males and a slight but significantly lower total protein when

compared to adult females (Table 1). No significant differences could be detected between nonanesthe tized adult males and females (Table 2). All cap tured young animals were anesthetized. Hemoglo bin levels, TPP, and MCHC could not be deter

mined for physically restrained (nonanesthetized) animals.

DISCUSSION H?matologie parameters of free-living marsh

deer frequently varied considerably within the groups analyzed. The variation was not unexpected because the study included animals captured over

a long period of time and exposed to different en

vironmental stimuli, though they were apparently

healthy at the time of capture. Care must be taken

if h?matologie data of free-living and captive ani

mals are to be compared. In the wild, marsh deer

would be expected to be exposed to more varied

environmental stimuli relative to those in a captive situation. Consequently, differing h?matologie pa rameters could be expected in free-living and cap tive animals.

Generally, it was the nonanesthetized, physically

SZAB? ET AL.?HEMATOLOGY OF FREE-LIVING MARSH DEER 465

20n

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Red Blood Cells

AAAA

n OpOfl_

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Figure 1. Comparison of red and white blood cell, mature neutrophil, basophil, lymphocyte, and monocyte counts of anesthetized (n

= 15) and nonanesthetized (n

= 17) female adult marsh deer. Asterisks indicate significant difference

between means at P < 0.05.

restrained deer that had higher blood values. Non

anesthetized females and males had significantly

higher RBC, WBC, neutrophil, and lymphocyte blood counts but lower basophil counts relative to

anesthetized counterparts. Nonanesthetized females

had higher monocyte counts, and nonanesthetized

males had higher packed cell volumes. There is no

obvious explanation for these differences, as non

anesthetized animals were under capture stress, but

the other group was under the effect of anesthetic

drugs. An important contribution to the observed

changes in red blood cell counts and packed cell

volume of nonanesthetized, physically restrained

deer may be splenic contraction in response to cat

echolamine release during acute stress.6 Catechol

amines can also cause a transient leukocytosis with

466 JOURNAL OF ZOO AND WILDLIFE MEDICINE

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Lymphocytes

anesthetized non-anesthetized

0.6

Q.5H > O a.

0.4

0.3

Packed cell volume

anesthetized non-anesthetized

Figure 2. Comparison of red and white blood cell, mature neutrophil, lymphocyte, and basophil blood cell counts

and packed cell volume of anesthetized (n =

15) and nonanesthetized (n = 17) male adult marsh deer. Asterisks

indicate significant difference between means at P < 0.05.

neutrophilia, monocytosis, and lymphocytosis by

shifting cells from the marginal pool into the cir

culating pool.4 Basophil counts must be interpreted with caution because they account for less than 2%

of the leukocytes in most species, and for precise

counts one must count thousands of cells.8 Thus,

explanations for the lower basophil counts of phys

ically restrained deer would be considered specu lative.

Anesthesia may also account for some of the ob

SZABO ET AL.?HEMATOLOGY OF FREE-LIVING MARSH DEER 467

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468 JOURNAL OF ZOO AND WILDLIFE MEDICINE

Table 2. Means, standard deviations (:! adult female and male wild marsh deer.b

SD), and ranges for blood values in nonanesthetized (physically restrained)

Blood parameters

Female (n = 25) Male {n = 17)

Mean ? SD Range Mean ? SD Range

RBO (X 1012/L) PCV

MCV (fL) WBC (X 107L) Neutrophil, band (X 109/L)

Neutrophil, mature (X 107L)

Eosinophils (X 109/L)

Basophil (X 107L) Lymphocyte (X 107L)

Monocyte (X 109/L)

11.5 ? 4.1 0.45 ? 0.05 45.0 ? 23.1 15.3 ? 4.2 0.0 ? 0.0 4.5 ? 2.3 1.9 ? 1.0

0.1 ? 0.2 8.4 ? 3.0 0.4 ? 0.3

3.2-18.4 0.30-0.54 26.5-140.6

8.0-23.1 0.0-0.2 1.5-10.9

0.1-4.1 0.0-0.7 3.2-17.1 0.0-1.3

11.3 ? 3.5 0.46 ? 0.04 46.7 ? 22.1 17.0 ? 5.9 0.0 ? 0.0 4.7 ? 1.8 1.9 ? 1.3

0.2 ? 0.2 9.6 ? 3.8 0.5 ? 0.7

3.9-15.3 0.37-0.53 27.1-107.7

9.7-33.0 0.0-0.0 1.9-7.8

0.19-4.9 0.0-0.7 2.9-18.5 0.0-2.6

a RBC, red blood cell; PCV, packed cell volume; MCV, mean corpuscular volume; WBC, white blood cell. b No significant differences (P < 0.05) were detected between nonanesthetized adult female and male wild marsh deer.

served differences between the two groups. Ace

promazine is known to reduce red and white blood

cell counts through hemodilution secondary to hy

potension, white blood cell margination along ves

sel walls, and splenic sequestration of erythro

cytes.7 Higher RBC, WBC, neutrophil, monocyte, and lymphocyte counts were also observed in phys

ically restrained sika deer (Cervus nippon yesoen

sis) when compared to anesthetized deer.5 It is im

possible to determine the extent of alteration in

blood parameters caused by physical restraint and

stress or anesthesia at this time.

Comparison of blood parameters within the anes

thetized marsh deer showed only minor differences

with age and gender. Young animals had signifi

cantly higher RBC counts than adult males but low

er total protein compared to adult females. Higher RBC counts in young animals were observed in

captive juvenile reindeer (Rangifer tarandus) also,1 but the meaning of this is unclear. Lower protein content is probably related to lower immunoglob ulin levels in the blood of young animals exposed to fewer environmental antigens than adult deer.

H?matologie reference ranges for free-living marsh deer are reported for the first time. Blood

parameters were found to vary significantly with

the method of restraint. It has been suggested pre

viously that two reference ranges for blood values

should be considered for wild animals2: one for

blood obtained from anesthetized animals and an

other for blood collected from nonanesthetized,

physically restrained animals. Other factors, such as

nutrition, circadian influences, physiologic status, time of year, and captivity might also influence h?

matologie parameters.

Acknowledgments: We thank CESP (Companhia

Energ?tica de S?o Paulo) for financial support and logistic collaboration. Permission for animal cap ture was kindly given by IB AMA. We are indebted to all the technicians and volunteers who helped

capture and maintain the animals.

LITERATURE CITED

1. Catley, A., R. A. Kock, M. G. Hart, and C. M.

Hawkey. 1990. Haematology of clinically normal and sick

captive reindeer (Rangifer tarandus). Vet. Rec. 126: 239 241.

2. Cross, J. P., C. G. Mackintosh, and J. F. T. Griffin. 1988. Effect of physical restraint and xylazine sedation on

haematological values in red deer (Cervus elaphus). Res. Vet. Sei. 45: 281-286.

3. Duarte, J. M. B., M. L. Merino, S. Gonzalez, A. L. V Nunes, J. M. Garcia, M. P. J. Szab?, J. R. Pandolfi, I. G. Arantes, A. A. Nascimento, R. Z. Machado, J. P. Ar

aujo, Jr., J. L. Cat?o-Dias, K. Werther, J. E. Garcia, R. J. S. Girio, and E. R. Matushima. 2001. Order Artiodactyla, Family Cervidae (Deer). In: Fowler, M. E., and Z. S. Cu bas (eds.). Biology, Medicine, and Surgery of South American Wild Animals. Iowa State University Press, Ames, Iowa. Pp. 402-422.

4. Jain, N. C. 1993. Essentials of Veterinary Hematol

ogy. Lea & Febiger, Philadelphia, Pennsylvania. P. 417. 5. Maede, Y., Y Yamanaka, A. Sasaki, M. Suzuki, and

N. Ohtaishi. 1990. Hematology in sika deer (Cervus nip pon yesoensis Heude, 1884). Nippon Juigaku Zasshi

52(1): 35-41. 6. Marco, I., and S. Lavin. 1999. Effect of the method

of capture on the haematology and blood chemistry of red deer (Cervus elaphus). Res. Vet. Sei. 66: 81-84.

7. Muir, W. W, and D. Mason. 1996. Cardiovascular

System. In: Thurmon, J. C, W J. Tranquilli, and G. J. Benson (eds.). Lumb and Jone's Veterinary Anesthesia, 3rd ed. Williams & Wilkins, Philadelphia, Pennsylvania. Pp. 62-114.

8. Scott, M. A., and S. L. Stockman. 2000. Basophils

SZAB? ET AL.?HEMATOLOGY OF FREE-LIVING MARSH DEER 469

and mast cells. In: Feldman, B. F., J. G. Zinkl, and N. C. Jain (eds.). Schalm's Veterinary Hematology, 5th ed. Lip pincott, Williams & Williams, Philadelphia, Pennsylvania. Pp. 308-317.

9. Tomas, W M., M. D. Beccaceci, and L. Pinder. 1997. Cervo-do-Pantanal (Blastocerus dichotomus). In:

Duarte, J. M. B. (ed.). Biologia e Conserva?ao de Cerv?

d?os Sul-Americanos: Blastocerus, Ozotoceros e Mazama.

Funep, Jaboticabal, S?o Paulo, Brasil. Pp. 24-40. 10. Wemmer, C. 1998. Status survey and conservation

action plan: Deer. IUCN/SSC Deer Specialist Group, Gland, Switzerland.

Received for publication 27 May 2004