Copeia 2009, No. 4, 793–800
Temporal Germ Cell Development Strategy during Mixed Spermatogenesis
within the Male Mediterranean Gecko, Hemidactylus turcicus
(Reptilia: Gekkonidae)
Justin L. Rheubert1,2, Erik H. Poldemann2, Mallory E. Eckstut1,3,
Matthew H. Collier2, David M. Sever1, and Kevin M. Gribbins2
The testes of Hemidactylus turcicus are composed of seminiferous tubules lined with continuous germinal epithelia in
which multiple germ cell morphologies can be found during the active months of sperm development. Spermatogenesis
is quiescent during September, with only spermatogonia A and spermatogonia B present in the seminiferous epithelia
and minimal mitotic activity is observed. Recrudescence begins in October and the early stages of spermatogenesis
progress through November. The onset of spermiation is observed in December and continues through August with the
heaviest sperm release occurring in June and July. Multiple generations of late elongated spermatids are found in
association with early mitotic and meiotic cells during the months of December–August. This temporal germ cell
development strategy is similar to that described in other squamates and anamniotes and is different from the spatial
development exhibited by birds and mammals, in which germ cell populations collectively progress through the stages
of spermatogenesis. The reptilian temporal model of germ cell development within a structurally amniotic testis leads
to two hypotheses in character evolution: birds and mammals exhibit convergence of germ cell development strategy,
or the spatial development strategy is a synapomorphy in amniotes and reptiles represent an evolutionary reversal to
the strategy employed by anamniotes. These findings along with present and future data may allow for more concrete
phylogenetic analyses by creating more characters for phylogenetic matrices and may prove to be useful in future
histopathological studies.
D
IFFERENT germ cell development strategies have
been described for fishes and amphibians, birds and
mammals, and reptiles. In the temporal germ cell
development strategy exhibited by fishes and amphibians,
the testes are composed of tubules/lobules lined with cysts
where germ cells develop as a single population through the
phases of spermatogenesis. Maturation culminates in cyst
eruption and leads to a single spermiation event where
mature sperm are released into a centralized lumen (Lofts,
1964). The spatial germ cell development strategy, displayed
by birds and mammals, involves germ cells that develop and
maintain consistent and predictable spatial relationships
(stages) within the seminiferous epithelia of the seminiferous
tubules that make up their testes (Leblond and Clermont,
1952; Russell et al., 1990; Kumar, 1995). Within these stages,
germ cells late in development are consistently associated
with early mitotic and meiotic cells, and as many as three
stages of elongating spermatids are observed within a single
cross-sectional view of a seminiferous tubule allowing
portions of the seminiferous tubule to be characterized by
stages. Stages are also sequentially ordered along the length of
the seminiferous tubules, which leads to waves of constant
sperm release within varying regions of the seminiferous
epithelia lining these tubules (Russell et al., 1990).
The reptilian germ cell development strategy, previously
described as an intermediate, resembles the amphibian’s
germ cell development strategy within a testis that contains
seminiferous tubules lined with a continuous epithelium
similar to birds and mammals (Gribbins and Gist, 2003). In all
temperate reptiles studied to date, sporadic bursts of sperm
release are visualized during spermatogenically active periods, with a decrease in mitotic activity, spermiogenesis, and
spermiation occurring throughout the colder months of the
year (Sauria: Gribbins and Gist, 2003; Chelonia: Gribbins et
al., 2003; Crocodylia: Gribbins et al., 2006; Serpentes:
Gribbins et al., 2008). Spermiation occurs along large
portions of the seminiferous tubules, and no consistent
spatial relationships are observed between germ cells. This
represents a pleisiomorphic germ cell development strategy
taking place within a derived amniotic testis and may suggest
the decoupling of testicular organization and germ cell
development strategy during character evolution. This
decoupling of the two characters may suggest either convergence of the spatial germ cell development strategy in birds
and mammals or character reversal in modern reptiles with
their temporal germ cell development strategy. The exact
mechanism is dependent on the ancestral states as well as the
origin of the spatial germ cell development strategy exhibited
by birds and mammals.
No previous studies involving germ cell development
strategies within temperate reptiles included a representative
of the family Gekkonidae. Although investigations exist
describing the reproductive cycles within geckos (Rose and
Barbour, 1968; Ikeuchi, 2004; Ibarguengoytia and Casalins,
2007), these studies primarily focus on the seasonality of
reproduction and do not detail germ cell morphologies or the
germ cell development strategy. To date, data suggest that
geckos of the genus Hemidactylus from tropical and temperate
areas demonstrate seasonality in terms of reproduction
(Sanyal and Prasad, 1967; Rose and Barbour, 1968; Selcer,
1987; Shanbhag et al., 2000). Rose and Barbour (1968)
described H. turcicus as being reproductively active from
November through August in southern Louisiana. However,
their data use testicular mass to infer spermatogenesis and do
not focus on germ cell development (Hernandez-Gallegos et
al., 2002). Eckstut et al. (2009) reported that male H. turcicus in
1
Department of Biological Sciences, Southeastern Louisiana University, Hammond, Louisiana 70402; E-mail: (JLR) [email protected].
Send reprint requests to this address.
2
Department of Biology, Wittenberg University, Springfield, Ohio 45504.
3
School of Life Sciences, University of Nevada, Las Vegas, Nevada 89154.
Submitted: 6 February 2009. Accepted: 2 July 2009. Associate Editor: S. A. Schaefer.
DOI: 10.1643/CG-09-034
F 2009 by the American Society of Ichthyologists and Herpetologists
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Louisiana are reproductively and spermatogenically active
from December through August based on the presence of
sperm in the efferent ductules.
This study focuses on histological investigation of the
reproductive cycle within the testis of Hemidactylus turcicus
in southern Louisiana and will be the first study to describe
the germ cell morphologies and development strategy
within geckos. The Mediterranean Gecko is a small (10–
13 cm) nocturnal lizard native to the Mediterranean Basin
and Canary Islands (Conant and Collins, 1998) that has
been introduced to southern coastal areas of the United
States. Studies show that reptilian species from these coastal
areas exhibit a quiescent period within their testes during
the colder periods of the year (Fox, 1958; Gribbins et al.,
2008). This disruption in spermatogenesis may be attributed
to the lack of available resources during colder months.
These energy sources are thought to maintain higher energy
allocations in support of spermatogenesis (Olsson et al.,
1997). Germ cell morphology, development strategy, and
reproductive cycle of the Mediterranean Gecko will be
compared to that of other vertebrates and to the influence
of seasonality on an introduced species within southern
Louisiana. We hypothesized that H. turcicus will employ a
germ cell development strategy similar to other reptiles and
that environmental conditions will have a strong influence
on when spermatogenic activity occurs in their testes.
MATERIALS AND METHODS
Animal collection.—Adult male Hemidactylus turcicus (n 5 47)
were collected during the months of November 2006
through October 2007 from Hammond, Ponchatoula, and
Baton Rouge, Louisiana. Specimens were sacrificed using a
0.2–0.5 ml injection of sodium pentobarbital (1 g sodium
pentobarbital in 10% ethanol/40% propylene glycol solution). Testes were excised, fixed in Trump’s fixative (Electron
Microscopy Sciences, Hatfield, PA), cut into transverse
sections, and stored in 70% alcohol at 4uC.
Tissue preparation for light microscopy.—Testes were cut into
small (2–3 mm) cubes and dehydrated using a graded series of
ethanol solutions (70%, 85%, 2 3 95%, 2 3 100%). Tissues
were then incubated in 1:2 and 1:1 Spurr’s plastic:100%
ethanol solutions (Electron Microscopy Sciences, Hatfield, PA)
for 30 minutes before being infiltrated in pure Spurr’s plastic
overnight. Samples were then embedded and cured in fresh
Spurr’s plastic at 60uC for 48 hr in a Fisher isotemperature
vacuum oven (Fisher Scientific, Pittsburgh, PA). Using a dry
glass knife and a LKB-Ultramicrotome III (LKB Produkter AB,
Bromma, Sweden), sections (2–3 mm) were cut from plastic
blocks and placed on standard microscope slides. The tissues
were stained using a basic fuchsin and toluidine blue
composite stain as described by Hayat (1993).
Histological analysis.—All samples were examined using an
Olympus compound microscope (Olympus America, Center
Valley, PA) to ascertain cellular morphologies that occurred
during germ cell development. To determine the presence or
absence of spatial stages, cross-sectional areas of the
seminiferous tubules were randomly selected and analyzed
over all months to determine if consistent cellular associations are seen between germ cell cohorts. Photographs were
taken using a SPOT digital camera (Diagnostic Systems
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Laboratories, Webster, TX), and plates were constructed
using Adobe Photoshop CS (Adobe Systems, San Jose, CA).
Data analysis, measurements, and basic morphometrics.—
Twenty cross sections of seminiferous tubules representing
each month were randomly chosen, and the tubule
diameter and germinal epithelial heights of each were
measured using an ocular micrometer. Data analysis was
performed using SigmaStat version 3.5 (Systat Software, Inc.,
San Jose, CA) for Windows. Tubule diameter and germinal
epithelial height data were tested for normality and
homogeneity of variances using the Kolmogorov-Smirnov
and Bartlett’s tests, respectively, before statistical analyses
were performed (Flemming, 1993; Sokal and Rohlf, 1995).
These data did not meet assumptions of normality or
homogeneity of variance; thus, nonparametric KruskalWallis analyses of variance were used to test for significant
monthly variation in seminiferous tubule diameter and
germinal epithelial height. Post-hoc nonparametric multiple
comparison tests using Dunn-Sidak procedures were then
used to indicate significant differences among pairs of
monthly means (Glantz, 1992).
RESULTS
Premitotic cells.—The testes of Hemidactylus turcicus are
composed of seminiferous tubules that are lined with
continuous seminiferous epithelia consisting of developing
germ cells and supporting Sertoli cells. Spermatogonia A and
B (Fig. 1: SpA and SpB), characterized by randomly dispersed
chromatin, are present during all months of the year and
replenish the germ cell population through mitotic proliferation. Spermatogonia A are classically ovoid whereas
spermatogonia B are typically more round. These two cell
types are found in the basal compartments of the seminiferous epithelia. Sertoli cell nuclei are usually seen in
juxtaposition to these spermatogonia, and the base of each
sustentacular nucleus typically rests close to the basal
lamina of the germinal epithelium. Once spermatogonia B
complete mitosis, they are known as pre-leptotene cells and
they represent the first meiotic stage of germ cell maturation. These pre-meiotic cells are present during every month
of the year although there is a substantial decrease in their
number in August and September testes.
Meiotic cells.—Pre-leptotene cells (Fig. 1: PL), the product of
mitotic divisions of spermatogonia B, contain well-defined
nuclear membranes with intensely staining nucleoli. Preleptotene cells are the smallest of the developing germ cells
and are easily distinguished from the much larger spermatogonia. As prophase I continues, the chromatin becomes more
filamentous and darker staining in leptotene cells (Fig. 1: LP).
Also during chromatin condensation, nuclear size increases
as the cytoplasmic volume decreases; this inverse relationship
between cytoplasmic and nuclear volumes continues
through the rest of prophase I. Zygotene cells (Fig. 1: ZY)
are the largest of the meiocytes and have thicker chromatin
fibers compared to leptotene spermatocytes. Pachytene
spermatocytes (Fig. 1: PA) are approximately the same size
as zygotene cells, but they have more open nucleoplasm and
the chromatin fibers are thicker. In diplotene spermatocytes
(Fig. 1: DI) the nuclear membranes of each cell begin to
degenerate, and fully condensed chromosomal fibers align
just beneath the degenerating membrane.
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spermatids begin to stretch (Fig. 1: S5), marking the
beginning of elongation. Elongation continues in step 6
spermatids (Fig. 1: S6) as is evident by their more cylindrical
shape. In step 7 spermatids (Fig. 1: S7), elongation terminates and condensation of DNA climaxes leaving the
nucleus slightly bent, which results in concave and convex
surfaces to the nuclear shape. When step 7 spermatids
complete maturation, they are shed as mature sperm (Fig. 1:
MS), which have highly elongated, intensely staining nuclei,
well-developed flagellae, and minimal cytoplasmic material.
Fig. 1. Germ cell types found within seminiferous tubules of Hemidactylus turcicus. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; PL, pre-leptotene spermatocyte; LP, leptotene spermatocyte;
ZY, zygotene spermatocyte; PA, pachytene spermatocyte; DI, diplotene
spermatocyte; M1, meiosis I; M2, meiosis II; ss, secondary spermatocyte; S1, step 1 spermatid; S2, step 2 spermatid; S3, step 3 spermatid;
S4, step 4 spermatid; S5, step 5 spermatid; S6, step 6 spermatid; S7,
step 7 spermatid; MS, mature sperm.
Meiosis I (Fig. 1: M1) marks the first meiotic division
where fully condensed chromosomes align at the metaphase
plate. After the first meiotic divisions, the secondary
spermatocytes (Fig. 1: SS) show a marked decrease in cell
size with a newly formed and easily definable nuclear
membrane and diffuse chromatin. Secondary spermatocytes
begin the second meiotic division and fully condensed
chromosomes align at the metaphase plate for a second time
(Fig. 1: M2). The primary distinction between meiosis I and
II cells is the amount of chromosomal material aligning at
the metaphase plate and cell size. Meiosis II cells have half
the number of chromosomes and are smaller in size.
Spermiogenic cells.—After the second meiotic division the
haploid spermiogenic cells begin their differentiation into
mature sperm in a stepwise fashion that can be characterized
into seven steps based on the terminology of Russell et al.
(1990) for mammalian species. Step 1 spermatids (Fig. 1: S1)
mark the beginning of spermatid maturation, and these cells
have well-defined nuclear membranes and diffuse cytoplasm
surrounding the nucleus. During the latter stages of step 1
and the beginning of step 2 spermatid development (Fig. 1:
S2), acrosome vesicles come in contact with the apices of the
nuclei. The meeting points of acrosomes and the nuclear
membranes create deep depressions in the step 2 spermatid
nuclei. Step 3 spermatids (Fig. 1: S3) show an increase in
acrosome size and acrosome granules are typically seen
within each vesicle. The acrosomes of step 4 spermatids
(Fig. 1: S4) begin to flatten and widen the nuclear depression
in which they are found. The posterior nuclei of step 5
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Seasonal variation and germ cell development strategy.—Distinct monthly differences can be observed in the testes of H.
turcicus histologically and morphometrically. Monthly variations in seminiferous tubule diameter (Kruskal-Wallis: H 5
146.94, df 5 11, P , 0.001; Fig. 2A) and epithelial height
(Kruskal-Wallis: H 5 148.43, df 5 11, P , 0.001; Fig. 2B) show
significant trends over all sampled months. A quiescent
period occurs during September (Fig. 3A) when only spermatogonia A and B and Sertoli cell nuclei are found in the
seminiferous epithelium. The seminiferous tubules are very
small and in most cases it is hard to distinguish distinct
lumina. The absence of large numbers of spermatids and
spermatocytes in September seminiferous tubules results in
the smallest tubule diameters (90.67 mm; Fig. 2A: superscript
subset F) and epithelial heights (32.65 mm; Fig. 2B: superscript
subset C). Spermatogenic activity begins in October (Fig. 3B)
with a slightly less vacuolated seminiferous epithelium and
increased mitosis along with the early stages of meiosis
dominating the apical compartment of the seminiferous
epithelium. There are 3–4 layers of spermatogonia A and B,
which are the results of a boost in mitotic divisions. This trend
continues into November (Fig. 3C) and many of the early
meiotic cells have advanced into the latter stages of meiosis. A
few of the developing germ cells have advanced as far as the
early stages of spermiogenesis (cells that are located very close
to the lumen). December (Fig. 3D) seminiferous epithelia are
dominated by spermiogenesis and the first signs of spermiation are evident by the presence of mature sperm within the
lumina of seminiferous tubules. Morphometric data support
the histological trends observed from October–December.
Both seminiferous tubule diameters (144 mm, 178 mm,
213 mm; Fig. 2A) and epithelial heights (38 mm, 54 mm,
57 mm; Fig. 2B) increase sequentially over these months.
As spermatogenesis continues, mitosis and meiosis are
maintained at a steady rate through the months of January–
July. The steady flow of cells finishing meiosis allows the
seminiferous epithelium to undergo several waves of intense
spermiogenesis, which leads to bursts of spermiation during
these months. The increased rate of spermiogenesis is best
visualized from the January (78 mm), February (81 mm), April
(87 mm), May (78 mm), and July (88 mm) seminiferous
epithelial heights (Fig. 2B: superscript subsets A) compared
to epithelial height (57 mm; Fig. 2B: superscript subset B) in
December. January seminiferous epithelia (Fig. 4A) have
very large populations of round spermatids, which are the
most common cells of spermiogenesis at this time. February
tubules (Fig. 4B) reveal that most of these early spermiogenic cells have now entered the elongation phase of
spermiogenesis. Many of these cells have completed spermiogenesis and are being shed to the lumina of the tubules.
Again in March (Fig. 4C), a new generation of round
spermatids are produced from Meiosis II with a few
elongated cells remaining in the apex of the seminiferous
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leading to a significant drop in tubule diameter (139 mm;
Fig. 2A: superscript subset E) and epithelial height (37 mm;
Fig. 2B: superscript subset C). This release of residual cells
completing spermiogenesis is evident from the cells being
close to the epithelia and the lumen not flooded with sperm.
Once the last cohort of spermatids finish spermiogenesis and
are shed to the lumen as mature sperm, the testis enters
quiescence again in September (Fig. 3A). During these
spermiogenically active months, layers of three or more
generations of early and late developing spermatids are
associated with early stages of mitosis and meiosis. This
accumulation of spermatid generations prevents consistent
cellular associations between groups of germ cells within the
seminiferous epithelia.
DISCUSSION
Fig. 2. (A) Variation in seminiferous tubule diameter and (B) variation in
germinal epithelial height during the entire calendar year of the male
Mediterranean Gecko, Hemidactylus turcicus. Values represented on
these graphs are means 6 1 standard error. Different superscripts indicate
significant differences (P # 0.05; Dunn-Sidak multiple range test).
epithelium. This loss of elongating spermatids and the
Sertoli cell processes that support them leads to a significant
drop in the seminiferous epithelial height (55 mm; Fig. 2B:
superscript subset B) and tubular diameter (209 mm; Fig. 2A:
superscript subset C) compared to January (78 mm; Fig. 2B:
superscript subset A; 227 mm, Fig. 2A: superscript subset A)
and February (81 mm, Fig. 2B: superscript subset A; 241 mm,
Fig. 2A: superscript subset B). By April (Fig. 4D), elongation
again dominates the apices of the seminiferous epithelia, as
seen by a significant increase in both tubule diameter
(220 mm; Fig. 2A: superscript subset A) and epithelial height
(87 mm; Fig. 2B: superscript subset A), and a burst of
spermiation leads to large amounts of mature sperm in the
lumina of the tubules. This sequential increasing and
decreasing trend of epithelial heights (EH) and tubule
diameters (TD) during spermiogenesis continues for the
months of May (continuation of spermiogenesis: TD:
229 mm, EH: 78 mm; Figs. 2A, 5A: superscript subset A; Fig. 2B:
superscript subset A), June (decreased spermiogenesis/increased spermiation: TD: 167 mm, EH: 51 mm; Figs. 2A, 5B:
superscript subset D; Fig. 2B: superscript subset B), and July
(increased spermiogenesis: TD: 243 mm, EH: 88 mm; Figs. 2A,
5C: superscript subset B; Fig. 2B: superscript subset A). In
August (Fig. 5D), mitosis and meiosis are complete and cells
that are left over in spermiogenesis finish their maturation
and are shed to the lumina of the seminiferous tubules,
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The testes of Hemidactylus turcicus are composed of seminiferous tubules lined with continuous germinal epithelia where
germ cells mature and migrate towards centralized lumina
and are released upon maturation. This testicular structure is
consistent with previous reports in non-mammalian amniotic
testes and closely resembles birds and mammals (Yamamoto
et al., 1967; Pudney, 1995; Gribbins et al., 2003, 2006, 2008).
The major events of spermatogenesis take place during the
months of October through July, with a decrease in
spermiation and no new germ cell recruitment occurs in
August. The September germinal epithelium, which is highly
vacuolated, consists of only spermatogonia A and B found in
the basal cell layers, and no mature sperm are found in the
lumen. Morphometric data support these histological findings, and the testis is in quiescence in September. Recrudescence begins in October when mitotic divisions increase and
spermatogonia start to enter the early phases of meiosis.
During December, spermiogenesis is well established and the
first events of spermiation become evident in the lumina of
seminiferous tubules. December histological data are also
supported by large tubule diameter and epithelial height
values and the second largest Dunn’s superscript grouping.
During spermatogenic activity, the events of spermiogenesis seem to progress slower than mitotic and meiotic
divisions. The decrease in progression of spermiogenesis
occurs during elongation and is evident in months where
elongation dominates and as many as three or even four
spermatid generations are seen together with mitotic and
meiotic cells within a single seminiferous tubule crosssection. The extended time which it takes to complete the
events of spermiogenesis is consistent with what has been
observed in other saurians (Gribbins and Gist, 2003) and all
major reptilian orders (Chelonia: Gribbins and Gist, 2003;
Sauria: Gribbins et al., 2003; Serpentes: Gribbins et al., 2008;
Crocodylia: Wang et al., 2008). This germ cell development
mode is similar to the anamniotic temporal germ cell
development strategy (Gribbins et al., 2006; Van Oordt
and Brands, unpubl.).
Spermatogenic activity continues at an elevated rate for
the months of January through July. This means that
spermatogenesis is maintained over most of the year with
August and September being the only months during which
spermatogenesis is decreased. Spermatogenesis continuing
into the next calendar year and the increase in spermatogenic activity late in the summer or fall before hibernation
constitutes a mixed-type of reproductive cycle (Goldberg,
1972) in Louisiana Hemidactylus turcicus. The decreased
seasonal variation in the testis results in waves of spermio-
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Fig. 3. (A) The cell types observed in the September seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; white
arrow, Sertoli cell. (B) The cell types observed in the October seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; PL,
pre-leptotene spermatocyte; LP, leptotene spermatocyte; ZY, zygotene spermatocyte; VA, vacuole. (C) The cell types observed in the November
seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; PL, pre-leptotene spermatocyte; LP, leptotene spermatocyte; PA,
pachytene spermatocyte; DI, diplotene spermatocyte; ZY, zygotene spermatocyte; S1, step 1 spermatid. (D) The cell types present in the December
seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; SS, secondary spermatocyte; S1, step 1 spermatid; S4, step 4
spermatid; S5, step 5 spermatid; MS, mature sperm.
genesis, with the largest peaks of sperm release histologically
and morphometrically occurring January/February, April/
May, and July. This extended spermatogenic cycle is much
different than what is seen in typical temperate saurians (for
a review see Licht, 1984), such as Podarcis muralis (Gribbins
and Gist, 2003). In P. muralis from southwestern Ohio,
spermatogenesis begins in late summer, much like H.
turcicus, but sperm development is then suspended from
November through March. Thus, in more northern ranges,
seasonality may play a larger role in determining when
lizard species can maintain energetically expensive processes
such as spermatogenesis (Olsson et al., 1997; Zaidan et al.,
2003), which may be the case for P. muralis during their long
dormant winter hibernation in Ohio. This hypothesis that
seasonality affects spermatogenic activity has also recently
been reported in the Western Cottonmouth, Agkistrodon
piscivorus leucostoma (Gribbins et al., 2008). Western Cottonmouths in southern Louisiana show two peaks of
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spermatogenesis (spring and fall), while Eastern Cottonmouths in a more northern Alabama population lack the
spring peak of spermatogenesis (Johnson et al., 1982) and
can only support a fall release of sperm.
Previous researchers (Hernández-Gallegos et al., 2002)
have hypothesized that testicular mass can be an inaccurate
measure of spermatogenic activity; however, the present
data corroborate well with Rose and Barbour (1968), who
reported that testicular mass in H. turcicus was greatest from
November through August and smallest in September
within a Texas population. Furthermore, the heaviest
spermatogenic activity reported in the present study
parallels reproductive activity (March–July) of H. turcicus
within its native range (Lee, 1996). Also, New Orleans
populations of H. turcicus (Rose and Barbour, 1968) have the
largest number of sperm within their efferent ductules
during these months, supporting when spermiation is
occurring in these Louisiana populations.
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Fig. 4. (A) The cell types observed in the January seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; PA, pachytene
spermatocyte; DI, diplotene spermatocyte; M1, meiosis I; M2, meiosis II; S1, step 1 spermatid; S7, step 7 spermatid; MS, mature sperm. (B) The cell
types observed in the February seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; PL, pre-leptotene spermatocyte;
LP, leptotene spermatocyte; SS, secondary spermatocyte; S1, step 1 spermatid; S5, step 5 spermatid; S6, step 6 spermatid; MS, mature sperm; *,
apparent vacuole as a product of the dehydration process. (C) The cell types observed in the March seminiferous epithelia. Bar 5 20 mm. SpA,
spermatogonia A; SpB, spermatogonia B; DI, diplotene spermatocyte; M1, meiosis I; M2, meiosis II; S1, step 1 spermatid; S5, step 5 spermatid; S6,
step 6 spermatid; MS, mature sperm. (D) The cell types observed in the April seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB,
spermatogonia B; PL, pre-leptotene spermatocyte; LP, leptotene spermatocyte; ZY, zygotene spermatocyte; S1, step 1 spermatid; S2, step 2
spermatid; S4, step 4 spermatid; S6, step 6 spermatid; S7, step 7 spermatid; MS, mature sperm.
Eckstut et al. (2009) found that the efferent ductules
contained a residual amount of sperm in September and no
sperm in October through December in Louisiana Mediterranean Geckos. These findings are congruent with data
presented here because the residual sperm in September
ducts could be sperm that completed spermiogenesis in late
August. With the observations that spermatogenesis begins
in October and the first spermiation events are not seen
until December, it is possible that the efferent ductules are
devoid of sperm at that time due to individual variation and
the amount of time required to finish spermatogenesis and
transfer sperm through the seminiferous tubules. Eckstut et
al. (2009) also found that females contained sperm in the
oviducts from May through August, which coincides with
the greatest amounts of spermiation in this study and
recorded events of mating.
Testicular structure is thought to be a synapomorphy for
amniotes because all amniotes studied to date possess the
same seminiferous tubule morphology (Aves: Yamomoto et
al., 1967; Mammals: Russell et al., 1990; Squamata: Gribbins
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and Gist, 2003), which differs from the lobule/cystic testis of
anamniotes (Pudney, 1995). Testicular structure in amniotes,
however, is decoupled from the vertebrate germ cell development strategy. Reptiles possess the anamniote temporal
pattern of germ cell development (Gribbins and Gist, 2003;
Gribbins et al., 2003, 2006, 2008; Wang et al., 2008), whereas
birds and mammals have a more synchronized spatial pattern
of germ cell development (Leblond and Clermont, 1952; Tait
and Johnson, 1982; Russell et al., 1990; Kumar, 1995). The
paraphyletic status of reptiles (Pough et al., 2000) and the
uncertain placement of turtles (Cao et al., 2000; Rieppel,
2008) make the evolution of germ cell development strategies
within amniotes difficult to determine. The most parsimonious hypothesis (two steps) would involve retention of the
anamniote germ cell development in basal amniotes and
modern reptiles, and independent origin (convergence) of the
spatial development type in birds and mammals. In an
alternate hypothesis, the spatial development type could
have evolved in the ancestor to the Synapsida, but this would
require reversals to the ancestral (temporal type) state in the
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Rheubert et al.—Spermatogenesis in Hemidactylus turcicus
799
Fig. 5. (A) The cell types observed in the May seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; S1, step 1
spermatid; S3, step 3 spermatid; S5, step 5 spermatid; S6, step 6 spermatid; S7, step 7 spermatid; MS, mature sperm. (B) The cell types observed in
the June seminiferous epithelia. Bar 5 20 mm. SpA, spermatogonia A; SpB, spermatogonia B; SS, secondary spermatocyte; S1, step 1 spermatid; S2,
step 2 spermatid; S6, step 6 spermatid; S7, step 7 spermatid; MS, mature sperm. (C) The cell types observed in the July seminiferous epithelia. Bar 5
20 mm. SpA, spermatogonia A; SpB, spermatogonia B; PA, pachytene spermatocyte; SS, secondary spermatocyte; S1, step 1 spermatid; S3, step 3
spermatid; S7, step 7 spermatid; MS, mature sperm; SC, Sertoli cell nucleus. (D) The cell types observed in the August seminiferous epithelia. Bar 5
20 mm. SpA, spermatogonia A; SpB, spermatogonia B; S5, step 5 spermatid; S6, step 6 spermatid; MS, mature sperm.
crocodilians, lepidosaurians, and possibly turtles. We intend
to investigate basal mammalian groups (monotremes and
marsupials) in the future to determine what germ cell
development strategy they employ during spermatogenesis.
These data, along with understanding the process of spermatogenesis within other reptilian representatives, may help
elucidate our understanding of spermatogenesis within
vertebrates, particularly Amniota.
ACKNOWLEDGMENTS
Specimens were collected under permit #LNHP-06-02 and
LNHP-07-12 issued by Louisiana Department of Wildlife and
Fisheries and protocols were approved by Southeastern
Louisiana University’s Institutional Animal Care and Use
Committee. We thank E. Lemons, D. Siegel, R. Chabarria, A.
Hamilton, and J. Oaks for assistance with the collection and
processing of specimens. We also thank C. McMahan for
providing insight on earlier drafts of this manuscript. This
Copeia cope-09-04-20.3d 25/9/09 18:53:40
799
project was funded by competitive research grants through
Wittenberg University.
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