Research Article |
Corresponding author: Marcio Frazão Chaves ( marciochaves@ufcg.edu.br ) Academic editor: Mauricio O. Moura
© 2017 Marcio Frazão Chaves, Geraldo Jorge Barbosa de Moura, Fernanda das C.M.A. Tenório, Josemberg da S. Baptista, Clovis J.C. Lapa Neto, Valeria W. Texeira, Álvaro A.C. Texeira.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Chaves MF, Moura GJB, Tenório FCMA, Baptista JS, Lapa Neto CJC, Texeira VW, Texeira ÁAC (2017) Influence of rainfall and temperature on the spermatogenesis of Leptodactylus macrosternum (Anura: Leptodactylidae). Zoologia 34: 1-7. https://doi.org/10.3897/zoologia.34.e20782
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In the semi-arid environment, the reproductive success of anurans depends on adaptations in their life cycle, which synchronizes with ideal environmental conditions to maximize the number of offspring. In this study changes in the histological and morphometric aspects of the testes of Leptodactylus macrosternum Miranda-Ribeiro, 1926 are characterized, to evaluate the influence of rainfall and temperature on them. Specimens were collected at Horto Florestal Olho d’Água da Bica – HFOB (06°49’20”S, 36°15’85”W) area, municipality of Cuité, state of Paraíba, Brazil. Search for specimens was active, happened at night, and amounted to 15 days from January to December 2013. The densities of spermatids (primary and secondary), sperm and area of the seminiferous locules of the testes were registered. The influence of climate variables (rainfall and temperature) on the density of primary and secondary spermatids, sperm and locular area were verified using Simple Linear Regression. Primary spermatids had the lowest density in July (57.90 ± 51.54 mm2), with a peak in November (300.32 ± 117.35 mm2); secondary spermatids had the lowest density in December (287.87 ± 79.05 mm2), with a peak in May (135,727.00 ± 301.13 mm2); sperm was in the lowest density in July (237.37 ± 121.10 mm2), with a peak in June (2,270.45 ± 602.62 mm2) and the locular area had the lowest density in December (40,292.9 ± 8,174.20 µm2) and highest density in June (338,875.01 ± 2,262.10 µm2). A notable decrease in sperm density was evident between June and July. That, associated with the observation of a larger locular area in June, allowed us to identify as June as the peak of spermatogenesis and the following month as the most potentially reproductive. The density of secondary spermatids(r = 0.02), sperm (r = 0.21) and locular area (r = 0.01) showed dependency on rainfall whereas only sperm (r = 0.09) showed dependency on temperature. Therefore, we can state that the reproductive cycle of L. macrosternum is potentially continuous, with a reproductive peak in July.
Anuran, semi-arid, spermatids, spermatozoa
In anurans, the control of spermatogenesis and consequently the reproductive cycles, are mediated by the interaction between the animal’s endogenous rhythms and the influence of environmental factors (
The reproductive cycles of anurans are classified based on the sum of the histological features of their gonads and their behavioral patterns as: continuous, potentially continuous and discontinuous (
Typical of semi-arid regions, potentially continuous cycles are determined by irregular and limited rainfall (
Leptodactylus macrosternum Miranda-Ribeiro, 1926 is widely distributed in semi-arid environments. It displays a generalist behavior and is well adapted to disturbed areas (
Night expeditions were undertaken every 15 days, from January to December 2013, to verify the reproductive activity of L. macrosternum males. According to
Males of L. macrosternum were collected manually at Horto Florestal Olhod’Água da Bica-HFOB (06°49’20”S, 36°15’85”W), located in the municipality of Cuité, Paraíba. The area is situated in the Brazilian Caatinga domain and is characterized by its hot, dry climate with temperatures ranging between 17 and 28 °C, an average monthly rainfall of 76.35 mm and relative humidity of approximately 70%. The hydrography of the Caatinga is peculiar, with temporary rivers, ponds, streams, reservoirs, wetlands and as slope areas from which the sources of water flow. The Horto Florestal encompasses 70 ha of bush and arboreal caatinga (
The monthly weather temperature (°C) and rainfall (mm) data were obtained from the Agência Executiva de Águas do Estado da Paraíba (
The captured males of L. macrosternum were taken to the Biosystematics Amphibian Laboratory (LABAN) of the Universidade Federal de Campina Grande, Cuité Campus, Paraíba, where they were euthanized with an overdose of lidocaine 5%.
The testes were fixed in Bouin solution (a mixture of 71% picric acid, 24% pure formaldehyde and 5% of glacial acetic acid) for 24 hours, then dehydrated in increasing alcohol series (70%-30 min, 80%-30 min, 90%-30 min, 95%30 min, 100%-30 min and 100%-30 min) (
Stereological analysis to determine the density of spermatids and spermatozoa was based on the precepts of Mandarin-de-Lacerda (
The values for the density of cells in the reproductive lineage and the area of the seminiferous locules were compared between months using the Kruskal-Wallis test and, when necessary, supplemented by the Dunn test.
The dependency between the climate variables (rainfall and temperature), the density of the population of reproductive lineage cells, and the area of the seminiferous locules were verified using a Simple Linear Regression test. All analyzes were based on
The highest rainfall rates were concentrated from April to July 2013. The lowest rainfall levels at the region were recorded in October and November (Figs
Twenty-nine L. macrosternum males were collected in May (5 samples), June (11 samples), July (3 samples), August (4 samples), November (4 samples) and December (2 samples) 2013, coinciding with rainfall in the region. Between January and April, and September and October 2013, no individuals of L. macrosternum were found during field trips.
All cell types in the testes of L. macrosternum collected during the sample period were easily verified, with each locule featuring multiple cysts of spermatogenic cells (Fig.
Primary spermatogonia (Fig.
Primary spermatocytes (Fig.
Primary spermatids (Fig.
The monthly density values of the primary spermatids (St1), secondary spermatids (St2), sperm (Ez) and variation of the locular area of seminiferous tubules showed significant differences (p < 0.05, Kruskal-Wallis) over the months of collection (Table
As shown in Table
Monthly values (mean and standard error) for density of primary spermatids (St1), secondary spermatids (St2), sperm (Ez) (mm2) and locular area (μm2) of the testes of Leptodactylus macrosternum species collected in the HFOB area, municipality of Cuité, state of Paraíba, Brazil, in the period from May to December 2013. * Significant differences (p < 0.05, Kruskal-Wallis).
St1 | St2 | Ez | Locular Area | |
May | 176.29 ± 46.64 | 1357.27 ± 30113* | 1121.97 ± 189.2* | 227688.7 ± 1274.1 |
Jun | 93.18 ± 58.35 | 1301.81 ± 117.64* | 2270.45 ± 602.62* | 338875.01 ± 226.1* |
Jul | 57.90 ± 51.54* | 633.33 ± 619.74 | 237.37 ± 121.1* | 96968.4 ± 464.2* |
Aug | 83.19 ± 37.12 | 770.45 ± 298.04 | 1007.57 ± 370.52 | 206367.2 ± 719.3 |
Nov | 300.32 ± 117.35* | 628.4 ± 3090.36 | 602.27 ± 212.83 | 281801.9 ± 4736.6 |
Dec | 139.06 ± 26.22 | 287.87 ± 79.05* | 356.06 ± 179.46 | 40292.9 ± 8174.2* |
Relations between the density of primary spermatocytes (St1), secondary spermatocytes (St2), sperm (Ez) and the area occupied by the seminiferous locules (LA) in male Leptodactylus macrosternum species in relation to rainfall and temperature in the Horto Florestal Olhod’Água da Bica (HFOB) area in the municipality of Cuité, state of Paraíba, Brazil, in the periods from May to August and November to December 2013. * = Represents significant differences.
Rainfall | Temperature | |||
r | p | r | p | |
St1 | 0.03 | 0.40 | 0.01 | 0.77 |
St2 | 0.02 | 0.04* | 0.05 | 0.20 |
Ez | 0.21 | 0.04* | 0.09 | 0.03* |
AL | 0.01 | 0.04* | 0.01 | 0.75 |
Histologic sections of Leptodactylus macrosternum testes, stained in HE, collected between the months of May to August and November to December in the Horto Florestal Olhod’Água da Bica (HFOB) area in the municipality of Cuité, state of Paraíba, Brazil. (3) section at 40x: (TA) tunica albuginea, (Med) mediastinum; (4) section at 40x TA: (St1) primary spermatid, (ST2) secondary spermatid, (Sp1) primary spermatocyte; (5) section at 40x TA: (Sg1) primary spermatogonia, (SG2) secondary spermatogonia, (Sp2) secondary spermatocyte, (Z) sperm; (6) section at 40x: (*) lumen with sperm. Scale bars: 3 = 500 µm, 4–6 = 50 µm.
Our results corroborate the results of Santos and Oliveira (
In our findings, the cell characteristics of L. Macrosternum, and the process of spermatogenesis, with variation in the quantities of cysts and especially the spermatozoa within the locules, are similar to L. podicipinus (
This morphology, added to a constant production and partial interruption when conditions are unfavorable for reproduction, features an acyclic spermatogenesis process. This appears to be a characteristic of anurans from the Fuscus group of Leptodactylidae (
Therefore, the characteristics observed in the gametogenic process of L. macrosternum classify this type of reproductive cycle as “potentially continuous”. This type is typical of seasonal, tropical environments (
With regards to the successive histological changes of the analyzed testes, the monthly density values of the primary spermatids (St1), secondary spermatids (St2), sperm (Ez) and variation of locular area of the seminiferous tubules varied significantly between the months of collection, besides portraying a chronological sequence that are compatible with the evolutionary stages of spermatogenesis (Table
In tropical regions of seasonal climate, the reproductive activity of anurans is associated with the rainy season (
In this study, the density of the secondary spermatocytes (St2), the density of sperm (Ez) and the monthly variation in the locular area of the seminiferous tubules (LA) (Table
The monthly temperature variation in the HFOB area only showed significant relationships with sperm density (Ez) (Table
Our findings confirm the studies of Prado Uetanabaro (
To SISBIO-RAN for the permission to collect and transport biological material (#44134-1).To the Foundation for Science and Technology of the state of Pernambuco-FACEPE (0287-2.05/14) for the financial aid during the execution of this study.