close

Вход

Забыли?

вход по аккаунту

?

j.theriogenology.2017.10.032

код для вставкиСкачать
Accepted Manuscript
Feasibility of semen collection in red-winged tinamou (Rhynchotus rufescens) by
manual stimulation and sazonality implications
Cristiane Sella Paranzini, Luiz Eduardo Cruz dos Santos Correia, Laiza Sartori de
Camargo, Kelry Mayara Silva, Tatyana Mendes de França, Josineudson Augusto de
Vasconcelos Silva, II, Nabor Veiga, Fabiana Ferreira de Souza
PII:
S0093-691X(17)30516-2
DOI:
10.1016/j.theriogenology.2017.10.032
Reference:
THE 14315
To appear in:
Theriogenology
Received Date: 1 June 2017
Revised Date:
21 October 2017
Accepted Date: 23 October 2017
Please cite this article as: Paranzini CS, dos Santos Correia LEC, de Camargo LS, Silva KM, de França
TM, de Vasconcelos Silva II JA, Veiga N, de Souza FF, Feasibility of semen collection in red-winged
tinamou (Rhynchotus rufescens) by manual stimulation and sazonality implications, Theriogenology
(2017), doi: 10.1016/j.theriogenology.2017.10.032.
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to
our customers we are providing this early version of the manuscript. The manuscript will undergo
copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please
note that during the production process errors may be discovered which could affect the content, and all
legal disclaimers that apply to the journal pertain.
REVISED HIGHLITED
ACCEPTED MANUSCRIPT
1
Brief observations on the feasibility of semen collection in red-winged tinamou
2
(Rhynchotus rufescens) by manual stimulation and sazonality implications
3
Cristiane Sella Paranzini1, Luiz Eduardo Cruz dos Santos Correia2, Laiza Sartori de
5
Camargo1, Kelry Mayara Silva1, Tatyana Mendes de França3, Josineudson Augusto II
6
de Vasconcelos Silva3, Nabor Veiga4, Fabiana Ferreira de Souza1*
RI
PT
4
7
8
1
9
State University, UNESP, Botucatu, Brazil
10
2
11
Jaboticabal, Brazil
12
3
13
UNESP, Botucatu, Brazil
14
4
15
Botucatu, Brazil
SC
Department of Animal Reproduction and Veterinary Radiology, FMVZ, São Paulo
M
AN
U
Department of Animal Science, FCAV, São Paulo State University, UNESP,
Department of Animal Nutrition and Breeding, FMVZ, São Paulo State University
16
TE
D
Department of Animal Production, FMVZ, São Paulo State University, UNESP,
*Corresponding author:
18
Fabiana F Souza, Departamento de Reprodução Animal e Radiologia Veterinária,
19
FMVZ, Universidade Estadual Paulista, UNESP
20
Rua Prof. Doutor Walter Mauricio Correa, s/n
21
Bairro: Unesp Campus de Botucatu, 18618-681 - Botucatu, SP, Brasil
22
Telephone number: +55(14)3880-2237
23
Email: [email protected]
AC
C
EP
17
24
25
26
1
ACCEPTED MANUSCRIPT
Abstract
28
This study aimed to report in detail, the technique and challenges of cloacal massage, to
29
collect and evaluate semen from red-winged tinamou (Rhynchotus rufescens) keep in
30
captivity, performed by only one technician. Sixty-four semen collection attempts, from
31
16 adult males, during breeding season and 16 attempts form these same 16 males in
32
non-breeding season, were performed. Prior to collection, all animals were conditioned
33
to cloacal massage for 6 weeks and the ejaculates were succeed with viable spermatozoa
34
and then, evaluated for feces, urine and mucus contamination, volume, concentration,
35
sperm vigor, motility, morphological defects and acrosome integrity. Semen collection
36
success rate was 63% in breeding season and 2 (5%) samples were discarded by grade 5
37
contamination. Only 3 ejaculates from 16 tinamou were obtained in non-breeding
38
season. Sperm concentration and acrosome integrity was higher (p = 0.00) in breeding
39
season, and the percentage of total sperm morphological defects, were high in both in
40
breeding and out breeding season. Overall, we concluded that the red-winged tinamou
41
breeding season, is linked to photoperiod (spring and summer), and at this period time,
42
semen can be obtained by cloacal massage collection satisfactorily, allowing its use in
43
reproduction biotechnologies and sperm cryopreservation.
44
45
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
27
Keywords: Bird; Tinamidae; Semen Collection; Season; Sperm Quality
46
47
1.
Introduction
48
The interest in breeding wildlife with commercial potential, such as tinamou
49
(Rhynchotus rufescens), is a viable alternative for this species conservation with
50
economic profitability. Knowledge on seminal characteristics and understanding of its
51
reproductive biology [1] is essential.
2
ACCEPTED MANUSCRIPT
The chosen semen collection technique varies by the research group experience.
53
Different techniques, such as collection direct from the vas deferens [2], mannequin
54
mate [1], artificial cloaca [3], cloacal massage [4], and electroejaculation [5]are
55
described as methods of obtaining semen from poultry since 1930. Manual stimulation
56
is a recommended method for wild birds and was described as a massage of the soft part
57
of abdomen, beneath the pelvic bones [4]. As advantage, cloacal massage presents total
58
control of the technician on the ejaculation moment and disadvantages, the need of
59
conditioning the birds and samples contamination with urine and feces. To the authors´
60
knowledge, in red-winged tinamou, cloacal massage technique for semen collection,
61
wasn´t previously described [4,6–8]. Thus, this study aimed to describe this technique
62
in detail handled by only one operator, different from the published literature on
63
domestic and wild birds. On the other hand, the semen collection allows seminal
64
evaluation and helps the identification of subfertile and infertile individuals, allowing to
65
classify male according sperm quality, for assisted reproduction programs [9].
66
Therefore, this study aimed to report in detail, the technique of semen collection by
67
cloacal massage in red-winged tinamou (Rhynchotus rufescens).
68
EP
TE
D
M
AN
U
SC
RI
PT
52
2.
Material and Methods
70
The tinamou used in this experiment were allocated at Fazenda Lageado (São Paulo
71
State University, UNESP, Botucatu, Brazil) separated in boxes. Sixteen males (5 years
72
to 9 years-old) were used. The study was registered on Brazilian Biodiversity
73
Authorization and Information System (Sistema de Autorização e Informação em
74
Biodiversidade – SISBIO number 50367 ),conducted in accordance with ethical
75
guidelines recommended by National Council for Control of Animal Experimentation
AC
C
69
3
ACCEPTED MANUSCRIPT
and College of Animal Experimentation, and approved by the Institution’s Animal Care
77
and Experimentation Ethics Committee on protocol number 98/2016.
78
After collection by cloacal massage, semen was evaluated for volume (µL),
79
concentration (x109 spermatozoa/mL), vigor (score 0 to 5), motility (%), morphological
80
defects (minor and major defects %) and acrosome integrity (%). It was kept in a 37° C
81
in dry bath until performed all semen sample analysis. Ejaculate volume was
82
determined by micropipette. Then semen was diluted in 0.9% saline solution in a
83
proportion of 1: 50 (2 µL: 98 µL) and deposited in a lamina with a cover sheet, and
84
evaluated to motility and vigor by a light microscope under 400X magnification.
85
Acrosome integrity was evaluated using 10 µL of this dilution added to 10 µL of the
86
POPE dye [10] and incubated for 70 seconds. Smear was prepared and evaluated under
87
a light microscope. Were counted 100 cells and classified as intact acrosome (lilac
88
stained) or not (pink/no stained) (figure 1) [11]. For sperm concentration, 1 µL of the
89
ejaculate was diluted in 499 µL of buffered formol saline solution (1: 500) and counted
90
in a Neubauer chamber. The remainder dilution was stored at 4° C for morphology
91
analysis under differential phase interference microscopy (1,000X magnification). The
92
sperm morphological defects were classified as head, intermediate piece and tail
93
defects.
94
The variables were described in mean and standard deviation. Shapiro-Wilk test was
95
used for variables normality and analyzed by t test with significance level p ≤ 0.05,
96
using R software version 3.2.2., to compare the seminal parameters in non- and
97
breeding season.
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
76
98
99
3.
Results
4
ACCEPTED MANUSCRIPT
The breeding season (September 2016 to February 2017) was identified by behavior, as
101
egg laying, vocalization, mating and dorsal region pecking of males and females (dorsal
102
areas of alopecia). Tinamou conditioning started in October 2016 by the contact of the
103
technician and the birds (1st week). In the 2nd week, the birds were restrained for cloacal
104
feather cut. By the 3rd week, cloacal and abdominal massages started (2 times/week).
105
For semen collection, the technician seated, supported the bird on one of his legs. With
106
the left hand, massage was performed on the tinamou back (caudally to the ribs and on
107
the testicles) making light pressure against the bird's body to help restraint, and with the
108
right hand, the abdominal region was stimulated between the pelvic bones to the cloaca
109
until the phallus was slightly engorged (figure 2 - A). At this point, the head of the
110
animal was trapped between the legs of the technician, leaving the bird upside down, for
111
better visualization of the phallus and semen collection accuracy. Then continuous
112
movements, with slight pressure, were made in the direction of the dorsal lips of the
113
cloaca until ejaculation (figure 2 - B). The technician should not allow phallus exposure
114
(figure 2 - C) until the ejaculation ends, as it has been noted that total exposure of this
115
organ, inhibits ejaculation. Already in the 4th and 5th weeks it was possible to observe
116
the elimination of mucous contents by the cloaca, containing spermatozoa, with rapid
117
movements (head and tail), but attached to the mucus. Due to the failure of the
118
collection, males were separated from the females and at the 6th week ejaculates were
119
obtained with viable spermatozoa. The ejaculate was collected in heated 1.5 mL
120
polypropylene microtubes with a flat surface cap (figure 2 – D) directly from the cloaca,
121
without the need of micropipette or microcapillary. Mucus secretion, urine or feces can
122
be eliminated after gentle cloacal massage and cleaning with a soft absorbent paper. The
123
contamination of the sample was graded (score from 1 to 5) by direct observation [1].
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
100
5
ACCEPTED MANUSCRIPT
During the breeding season, 64 attempts to collect semen were made (4 attempts per
125
animal, once a week), with success rate of 63% (40/64). From 40 samples, only 2 (5%)
126
were contaminated with feces and urine grade 5 and were discarded, 4 (10%) were
127
grade 2 and 1 (2.5%) sample, grade 1.
128
Other collection attempt was made soon after breeding season ends (March 17, 2017).
129
Only 3 (19%) ejaculates were obtained from 16 fowl and 1 sample had grade 2
130
contamination. Seminal parameters in breeding and non-breeding season were described
131
in Table 1. None of the birds died after their manipulation (stress).
132
In breeding season, the morphological analysis showed 65% defects (80% tail, 14%
133
intermediate piece and 6% head). Otherwise, in non-breeding season were observed
134
70% defects (6% tail, 7% intermediate piece and 87% head). The common sperm
135
defects found were looped and bent head, round-headed, folded intermediate middle
136
piece and strongly folded tail.
TE
D
137
M
AN
U
SC
RI
PT
124
4. Discussion
139
Because they are wild animals, it is important a continuous contact between technician
140
and birds, for adaptation and reducing seminal collection stress. Cloacal feather cutting
141
facilitates the direct collection from the cloaca and together with the massages (cloacal
142
and abdominal), make the fowl more tolerant and conditioned to the procedure.
143
This contact also aimed to observe individual behavior. We noted that some birds
144
needed longer stimuli, while the mean time of massage for each animal was around 2
145
minutes until total ejaculation. Some males vocalized during stimuli, and these males
146
ejaculated the greater seminal volume. It was possible to notice cloacal muscles
147
movement (contraction). Some fowl were so conditioned, that start to present cloacal
AC
C
EP
138
6
ACCEPTED MANUSCRIPT
contractions already at the restraining. Also, we observed that the stimulus was correct,
149
because the bird allows cloacal manipulation without struggle.
150
During the 4th and 5th week of conditioning, mucoid secretion was obtained with live
151
spermatozoa. It´s believed that this mucous secretion comes from secretory anal glands.
152
The literature reports a waxy, greasy or mucous material in ratites, some large raptors,
153
and a few passerines. Even the presence of mucoid secretion has not been reported in
154
many other species, secretory anal glands have been found in 72 species in 42 families
155
[12]. It is unclear if it was vestigial spermatozoa due to previous copulation, due to the
156
lack of technician experience or the lack of tinamou familiarity to the procedure. At this
157
point, it was decided to separate the male from female, and apparently it had a positive
158
influence, because at the 6th week, viable spermatozoa were acquired. The cloacal
159
mucoid gland function is unclear, however, this secretion compromises seminal quality
160
[6].
161
Seminal color is a good index of sample contamination with urine and feces. The color
162
without contamination ranges from opaque white, white transparent to yellowish. In
163
contaminated
164
white/yellow/green. Because it is a very subjective analysis, the score (1-5) will depend
165
on the experience of the observer. Samples with contamination 4 and 5 degree were
166
discarded because semen analysis showed ejaculate of poor quality, which make it
167
unfeasible to biotechnologies applicability. Urine contamination leaves the semen pH
168
acidic and reduces sperm viability, which could be bypassed by washing it with neutral
169
extender, but its benefits, is under discussion [13]. In endangered species this shall be
170
used as attempt to help its preservation.
TE
D
M
AN
U
SC
RI
PT
148
the
color
range
from
white/green/brownish
to
AC
C
EP
samples,
7
ACCEPTED MANUSCRIPT
As this study shows, semen concentration was higher in breeding than non-breeding
172
season. It was already predictable due the changes in the libido, testicular size, LH and
173
testosterone levels during non-breeding season [14].
174
Semen volume collected was greater than others studies [5,10], however concentration,
175
motility, vigor, morphological defects and acrosome integrity were similar in breeding
176
season. Although the last author [15] used the same tinamous’ group that we use in this
177
study, we believe that there is a difference in the volume of ejaculate, due to the type
178
and intensity of the stimulus used at the time of collection, and because the technique
179
that they use was cloacal digital manipulation, and we performed abdominal and cloacal
180
massage, followed by cloacal digital manipulation.
181
Because of the semen high concentration, it was necessary a dilution (1:50) for
182
immediately motility and vigor evaluation. According to Cavalcante [16], the red-
183
winged tinamous ejaculate dilution with 0.9% NaCl, a commercial turkey extender, and
184
other two extenders, for fresh analysis, did not differ. The differences were found after
185
24 and 48 h of refrigeration. 0.9% saline solution (380 mOsm) is considered an isotonic
186
solution, since the red-winged tinamous semen osmolarity is around 300 mOsm [16].
187
Reproductive features vary in the birds genera in the same order and there is differences
188
in the seasonality expression [17]. In this study, the seasonality of tinamou in São Paulo
189
(latitude: 22º 53' 09" S; longitude: 48º 26' 42" W; altitude: 804 m) state in 2016/17 was
190
elucidated by behavioral patterns and semen analysis. Too, by the necropsy of some
191
males tinamou, we could notice the difference of the testicle size in breeding and no-
192
breeding season (figure 3). There were scarce studies about sperm parameters of the
193
red-winged tinamou (Rhynchotus rufescens), and this study contributed to understand
194
some their reproductive characteristics and behavior. The difficult of tinamous’ semen
195
parameters standardization is due to the lack of collection experience, seasonality,
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
171
8
ACCEPTED MANUSCRIPT
inbreeding and fowl training. Those facts interfere directly in the ejaculate quality.
197
Consequently, semen biotechnology development and breeding program success is
198
hampered.
199
Percentage of morphological defects found in breeding season in this study was very
200
high compared with a study in other tinamous’ species [15], that found around 30%. In
201
breeding season, folded tail was the main morphological defect and in non-breeding
202
season was those associated to the spermatozoa head, which are related with the
203
spermatogenesis. Sperm morphological defects in mammals, can be related to aging
204
[18]. Red-winged tinamou reaches its sexual maturity around ten months [19], and to
205
our knowledge, there are no scientific records about their lifespan. It´s believed that
206
tinamou species can live up to 10 years [20]. This life expectancy might increase in
207
captivity. Previous study conducted by Góes et al. [11], that studied these same fowls,
208
two years before (3 -4 years age), showed similar results (60%). By this, we believe that
209
these features could be related with high endogamy degree, as they are being raised in
210
captivity since 1987, without the introduction of new fowls, as it was previous studied
211
in other animal species, endogamy coefficient showed a strong inverse relationship with
212
ejaculate quality [21]. In our knowledge there are scarce studies mentioning tinamous
213
morphological spermatozoa defects [15] and considering the role sperm morphology
214
plays in fertility, further studies about morphological defects are essentials.
215
Acrossomal integrity has higher indices in breeding season than non-breeding season.
216
It´s an indication, together with low concentration, that the sperm capacity to fertilize an
217
egg, is righer in breeding season.
218
This paper describes, to our knowledge, for the first time, the successful cloacal
219
massage, to collect and evaluate semen from red-winged tinamous, performed by only
220
one technician. Overall, we concluded the red-winged tinamou breeding season, which
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
196
9
ACCEPTED MANUSCRIPT
is linked to photoperiod (spring and summer), occurs from September to March in the
222
south hemisphere. Semen collection by cloacal massage can be performed by only one
223
technician. The red-winged tinamou are easily adaptable to handling, and semen
224
collection can be facilitated by the experience and perception of the technician. The
225
absence of females in male donors of semen group, for a few days, before the collection,
226
increases the libido and the sperm quality. Feces and urine contamination occurs due the
227
lack of bird adaptation to technique, technician experience, and the semen contaminated
228
should be discarded, but can be used in endangered species. Semen characteristics,
229
excluding morphological defects, allow to use the samples in reproduction
230
biotechnologies as sperm cryopreservation in commercial breeding and to species
231
conservation.
M
AN
U
SC
RI
PT
221
232
5. Conclusion
234
Cloacal massage performed by one person is possible in red-windged tinamou, but its
235
seminal quality is strong related to breeding season.
236
TE
D
233
Acknowledgment
238
We would like to thank the São Paulo State University (UNESP), School of
239
Agricultural and Veterinarian Sciences, Jaboticabal, Brazil, for giving the tinamous
240
used in this study; and NUPECCE´s (Núcleo de Pesquisa e Conservação de Cervídeos)
241
staff; Prof. Dr. Frederico Ozanam Papa for allowing the use of his laboratory for semen
242
morphological analysis. The study was supported by the Brazilian Coordination for the
243
Improvement of Higher Education Personnel (CAPES).
AC
C
EP
237
244
245
Conflict of interest
10
ACCEPTED MANUSCRIPT
246
None of the authors of this paper has any financial or personal relationship with any
247
other person or organisation that might inappropriately influence or bias the content of
248
the paper.
249
Author contribution
251
All authors made a substantial contribution to the execution of this study. Paranzini,
252
Ferreira, Correia, J. Silva and Veiga designed the study. Paranzini, conducted seminal
253
collection with França, K. Silva and Correia support. Paranzini, Ferreira and Camargo
254
were responsible for seminal analysis. França and Correia were responsible for
255
management, handling, separating and feeding, all animals including weekend and
256
holidays. Paranzini with Ferreira helps, drafted the paper. All authors contributed to the
257
manuscript revisions.
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
250
11
ACCEPTED MANUSCRIPT
258
References
259
[1]
Dogliero A, Rota A, Lofiego R, Mauthe von Degerfeld M, Quaranta G. Semen
evaluation in four autochthonous wild raptor species using computer-aided sperm
261
analyzer. Theriogenology 2016;85:1113–7.
262
doi:10.1016/j.theriogenology.2015.11.023.
1913:371.
264
265
[3]
[4]
[5]
Samour JH. Semen Collection , Spermatozoa Cryopreservation , and Artificial
Insemination in Nondomestic Birds. J Avian Med Surg 2004;18:219–23.
270
[6]
Gee GF, Bertschinger H, Donoghue AM, Blanco J, Soley J. Reproduction in
TE
D
271
Burrows WH, Quinn JP. A Method of Obtaining Spermatozoa from the Domestic
Fowl. Poult Sci 1935;14:251–4. doi:10.3382/ps.0140251.
268
269
Ishsikawa H. The lift duration of cock spermatozoa outside the body. Proceeding
World Poult. Congr., 1930, p. 91.
266
267
Ivanov E. Experience sur la fecondation artificielle des oiseaux. C R Biol
SC
[2]
M
AN
U
263
RI
PT
260
Nondomestic Birds: Physiology, Semen Collection, Artificial Insemination and
273
Cryopreservation. Avian Poult Biol Rev 2004;15:47–101.
274
doi:10.3184/147020604783637435.
[7]
semen analysis for evaluation of Rosy-faced lovebird (Agapornis roseicollis)
276
semen collected in different periods of the year. Theriogenology 2015;83:103–6.
277
doi:10.1016/j.theriogenology.2014.08.013.
278
279
Dogliero A, Rota A, von Degerfeld MM, Quaranta G. Use of computer-assisted
AC
C
275
EP
272
[8]
Dogliero A, Von Degerfeld MM, Rota A, Pregel P, Quaranta G. Comparison of
280
different methods of semen cryopreservation in Melopsittacus undulatus. Anim
281
Reprod Sci 2017;185:53–65. doi:10.1016/j.anireprosci.2017.08.002.
282
[9]
Fischer D, Neumann D, Wehrend A, Lierz M. Comparison of conventional and
12
ACCEPTED MANUSCRIPT
283
computer-assisted semen analysis in cockatiels (Nymphicus hollandicus) and
284
evaluation of different insemination dosages for artificial insemination.
285
Theriogenology 2014;82:613–20. doi:10.1016/j.theriogenology.2014.05.023.
[10] Pope CE, Zhang M, Dresser LB. A simple staining method for evoluating
287
acrosomal statue of cat sperm. J O Zoo Wildl Med 1991;22:87–95.
RI
PT
286
[11] Góes PA de A, Cavalcante AC da S, Tavian AF, Felipe L, Santos EC, Nichi M,
289
et al. Análise espermática de perdizes (Rhynchotus rufescens) criadas em
290
cativeiro e suplementadas com selênio. Brazilian J Vet Res Anim Sci
291
2011;48:370–7.
293
294
[12] Oliveira CA De, Mahecha GAB. Morphology of the copulatory apparatus of the
M
AN
U
292
SC
288
spotted tinamou. Ann Anat 2000;182:161–9.
[13] Umapathy G. Semen Characteristics of the Captive Indian White-Backed Vulture
(Gyps bengalensis). Biol Reprod 2005;73:1039–45.
296
doi:10.1095/biolreprod.105.043430.
297
TE
D
295
[14] Noirault J, Brillard J-P, Bakst MR. Effect of various photoperiods on testicular
weight, weekly sperm output and plasma levels of LH and testosterone over the
299
reproductive season in male turkeys. Theriogenology 2006;66:851–9.
300
doi:10.1016/j.theriogenology.2005.11.025.
302
303
[15] Felipe L, Santos EC, Tavian AF, Góes PAA, Moraes VMB, Tonhati H, et al.
AC
C
301
EP
298
Effect of crude protein levels and organic selenium supplementation in the diets
fed during the breeding season on reproductive parameters of red-winged
304
tinamous ( Rhynchotus rufescens ). Brazilian J Poult Sci 2010;12:63–71.
305
doi:10.1590/S1516-635X2010000100010.
306
307
[16] Cavalcante AKS. Parâmetros reprodutivos de perdizes machos ( Rhynchotus
rufescens ) criadas em cativeiro : comparação entre os índices reprodutivos de
13
ACCEPTED MANUSCRIPT
308
309
animais acasalados e inseminados. São Paulo University, 2006.
[17] Bublat A, Fischer D, Bruslund S, Schneider H, Meinecke-tillmann S, Wehrend
A, et al. Theriogenology Seasonal and genera-specific variations in semen
311
availability and semen characteristics in large parrots. Theriogenology
312
2017;91:82–9. doi:10.1016/j.theriogenology.2016.11.029.
313
RI
PT
310
[18] Kidd SA, Eskenazi B, Wyrobek AJ. Effects of male age on semen quality and
fertility: A review of the literature. Fertil Steril 2001;75:237–48.
315
doi:10.1016/S0015-0282(00)01679-4.
316
SC
314
[19] Marques MVR. Tinamiformes (Codorna, Inhambu, Macuco, Jaó, e Perdiz). In:
Cubas ZS, Silva JCR, Catão-Dias JL, editors. Tratado de Animais Selvagens. 2a
318
ed., São Paulo: Roca; 2014, p. 2740.
319
M
AN
U
317
[20] Burton M, Burton R. Tinamou. In: Burton M, Burton R, editors. Int. Wildl.
Encycl. Sweetfish - tree snake. 3rd ed., Tarrytown, New York: Marshall
321
Cavendish; 2002, p. 590.
TE
D
320
[21] Roldan ER, Cassinello J, Abaigar T, Gomendio M. Inbreeding, fluctuating
323
asymmetry, and ejaculate quality in an endangered ungulate. Proc Biol Sci
324
1998;265:243–8. doi:10.1098/rspb.1998.0288.
326
327
328
AC
C
325
EP
322
329
330
331
332
14
ACCEPTED MANUSCRIPT
333
Figure 1. Pope´s coloration for Tinamous´ acrossomal integrity. A. Intact acrossoma. B.
334
Acrossoma not intact. C and D. Detached acrossome.
335
Figure 2. Handling and restraint Tinamous (Rhynchotus rufescens) for semen collection.
337
A. Dorsal stimuli and abdominal massage. B. Cloacal massage, with phallus exposure
338
(arrow). C. Tinamous (Rhynchotus rufescens) phallus exposure (arrow) during semen
339
collection. D. Microtube with tinamous’ (Rhynchotus rufescens) ejaculates.
RI
PT
336
SC
340
Figure 3. Tinamous (Rhynchotus rufescens) testicle. A. Non-breeding season. B/C. In
342
breeding season/ measure in mm
AC
C
EP
TE
D
M
AN
U
341
15
ACCEPTED MANUSCRIPT
1
Table 1.
2
Semen parameters (mean ± standard deviation) from 16 red-winged tinamous
3
(Rhynchotus rufescens) collected by manual stimulation in breeding and non-breeding
4
season.
Non-breeding season
(n = 38)
(n= 3)
Volume (µL)
58.4 ± 44.5
55.5 ± 13.0
0.42
Concentration (x109 sperm/mL)
1.62 ± 0.73
0.72 ± 0.05
0.00
Motility (%)
73.0 ± 14.0
Vigor (0 to 5)
3.7 ± 1.0
Acrosome integrity
92.0 ± 3.2
79.0 ± 2.2
0.01
70.0 ± 13.0
75.0 ± 16.0
0.28
p value*
defects (%)
3.5 ± 0.7
0.42
EP
TE
D
*significance level p ≤ 0.05
0.37
AC
C
5
75.0 ± 7.0
M
AN
U
Total sperm morphological
SC
Semen parameter
RI
PT
Breeding season
1
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
ACCEPTED MANUSCRIPT
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
ACCEPTED MANUSCRIPT
AC
C
EP
TE
D
M
AN
U
SC
RI
PT
ACCEPTED MANUSCRIPT
ACCEPTED MANUSCRIPT
Highlights
•
Breeding season in red-winged partridges is linked to photoperiod (spring and
summer) that occurs from September to March in the south hemisphere
Semen collection by cloacal massage can performed by only one technician
•
Semen concentration and acrosome integrity has better parameters in breeding
than out-breeding season
SC
The endogamy is related with decreased sperm quality
AC
C
EP
TE
D
M
AN
U
•
RI
PT
•
Документ
Категория
Без категории
Просмотров
3
Размер файла
1 245 Кб
Теги
2017, 032, theriogenology
1/--страниц
Пожаловаться на содержимое документа