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At least 19 recordsLinked to original sources

Epididymal epithelium immortalized by simian virus 40 large T antigen: a model to study epididymal gene expression.

Primary cultures of the differentiated, adult epididymal duct epithelium were immortalized by retroviral transduction with the simian virus (SV)40 large T antigen. The canine epididymis was chosen here as a model with high human relevance, representing a convenient and acceptable source of differentiated epididymal tissue and, compared to other animal models, expressing a relatively large number of gene products which are also expressed by the human epididymis. To determine whether the immortalized canine epididymal (IMCE) cells retained a phenotype comparable to the original tissue, epithelial cytokeratins, various epididymal transcription factors as well as mRNAs encoding abundant epididymal secretory proteins, were studied as molecular markers. All IMCE populations obtained after transduction were of epithelial origin. The nuclear androgen receptor (AR) and the polyoma enhancer activator (PEA3), as well as the epididymal mRNA encoding the canine counterparts of human HE1, HE4 and HE5/CD52 epididymal mRNA, were retained in all populations tested. The majority of tested clones were oestrogen receptor ERalpha-positive, but ERbeta-negative, while one ERalpha-negative cell population was positive for ERbeta. The IMCE populations described thus represent useful permanent tools for studying gene expression of the epididymal duct epithelium, and for other types of experiments, examples including drug effects and toxicity on the epididymis.

Animals↗

Bacterial epididymitis in the rat: a model for assessing the impact of acute inflammation on epididymal antibiotic penetration.

A rat model of bacterial epididymitis was developed and characterized for use in assessing the impact of acute epididymal inflammation on antibiotic penetration into the epididymis. A 0.2 ml. intratesticular injection of a 0.5 McFarland standard suspension of E. coli resulted in histologically confirmed acute epididymitis in all animals studied. Inflammatory changes were detectable as early as 24 hours following inoculation and were progressive to the last assessment point at 11 days. Early testicular infarction was observed in association with epididymal inflammation. Serial transcrotal ultrasounds of infected animals showed progressive increase in epididymal size and a late decrease in testicular size. Serum and epididymal drug concentrations were assayed following a single dose of the antibiotic amdinocillin. Fifteen minutes following the peak serum level, the drug concentration in infected epididymis was 2.3-fold higher than the contralateral, non-infected epididymis. These data suggest that acute inflammation enhances antibiotic penetration into the infected epididymis. The model described provides a rapid, reproducible method to study epididymal drug delivery in normal and diseased states.

Amdinocillin↗

Chronic epididymitis (epididymal nodule) mimicking an adenomatoid tumor--case report with review of literature.

AIM: To describe and review the differential diagnosis of epididymal nodules and chronic epididymitis so as to have a broad view of this pathology. We have suggested a possible diagnostic algorithm for the workup of an epididymal nodule. METHODS/RESULTS: We have reported a case of a 35-year-old patient with a symptomatic palpable nodular mass in the tail of the left epididymis. Fine needle aspiration suggested an adenomatoid tumor while the surgical excision histology was reported as chronic epididymitis. CONCLUSIONS: Epididymal nodules are frequently encountered in the epididymis. Their differential diagnosis includes chronic granulomatous epididymitis, adenomatoid tumor and benign paratesticular neoplasms. We consider that it is important to distinguish epididymal nodules from benign inflammatory lesions and the threshold for a surgical excision should be low as it is therapeutic and provides a definite diagnosis. Whilst the FNAC alone may not be always adequate to confirm diagnosis, an epididymectomy may be curative as well as diagnostic in selected cases.

Adenomatoid Tumor↗

[Acute epididymitis and antibiotherapy. Measurement of the epididymal diffusion of pefloxacin].

Acute epididymitis is a common infection in the young sexually active adult. Etiologically, the organisms most frequently found are Chlamydia trachomatis, Neisseria gonorrhea and gram negative bacilli. Pefloxacin is a novel quinolone whose antibacterial spectrum and bactericidal activity allow it to be considered for use in the treatment of orchitis and epididymitis Prior to clinical study the authors investigated the degree of epididymal diffusion of Pefloxacin. Ten subjects underwent extraction of an epididymal sample by direct access to the epididymis through a transverse scrotal incision under peridural anesthesia. Three days before surgery, Pefloxacin was administrated at the rate of 400 mg every 12 hours by the oral route. On the day of the operation, the subject receive a 400 mg infusion over one hour, 2 hours before epididymal biopsy. Blood specimens were also extracted. Pefloxacin assays were performed by HPLC following a method derived from that of Montay. The trough concentration of Pefloxacin inhibiting 90% of sensitive strains (MIC 90) is less than or equal to 2 micro-g/ml. The majority of sensitive organisms have minimum bactericidal concentrations equal to twice the MIC 90. The epididymal concentrations that the authors measured are situated at values of 8.15 to 21.80 miro-g/g tissue (mean 13.44). These data allow the use of Pefloxacin to be considered an option in the treatment of epididymitis.

Acute Disease↗

Studies on the binding of a 32K rat epididymal protein to rat epididymal spermatozoa.

A glycoprotein of molecular weight 32K has been isolated and purified from the rat caudal epididymal fluid by gel filtration, ion-exchange and affinity chromatography. The highly purified protein was labeled with radioactive iodine and the binding of the 125I-labeled 32K rat epididymal protein (REP) to washed rat caudal epididymal sperm was studied under various conditions. Scatchard plots of the binding data revealed two binding kinetics. One bound with high affinity (KD = 2.6 X 10(-10) ) but low capacity. The other bound with lower affinity (KD = 2.2 X 10(-9)M) but high capacity. The rate of binding of the labeled protein to sperm was dependent on the temperature of the incubation medium. At the scrotal temperature of 33 degrees C, maximal binding was obtained after 40 min. However, at 22 degrees C equilibrium state was reached after 90 min and at 0 degrees C, the equilibrium rate was not reached even after 120 min of incubation. Binding showed dependence on extracellular pH (optimal pH at 4) and ionic strength of the incubation medium. High ionic strength was found to inhibit binding of the 125I-labeled 32K REP to rat caudal epididymal sperm. Specific binding was abolished by 100-fold molar excess unlabeled 32K REP or by native rat caudal epididymal fluid proteins, but not by albumin or ovalbumin. This indicates high specificity of binding. This study has provided direct evidence for the interaction of an epididymal protein with epididymal spermatozoa.

Animals↗

Localization of a maturation-dependent epididymal sperm surface antigen recognized by a monoclonal antibody raised against a 135-kilodalton protein in porcine epididymal fluid.

A specific 135-kDa protein was purified from porcine cauda epididymal fluid. Analysis of its N-terminal amino acid sequence revealed it to be a new protein. Stable clones of hybridomas that produced monoclonal antibodies against the purified 135-kDa protein were established. A clone, B-11, reacting both with epididymal fluid and with sperm plasma membranes was selected and used in this study. Immunoblotting analysis showed that B-11 reacted only with a 135-kDa protein among epididymal fluid proteins. In contrast, B-11 did not recognize a similar 135-kDa sperm protein but did strongly react with a 27-kDa protein among sperm membrane proteins, extracted by NP-40 in the presence of protease inhibitors. B-11 also reacted only with a 27-kDa protein fragment among trypsin digests of the 135-kDa epididymal protein. The 135-kDa protein was first detected, by ELISA or immunoblotting analysis, at the beginning of the corpus epididymis. Maximal levels were reached in the distal corpus and levels were slightly decreased in the cauda epididymis. On the other hand, the surface of caput sperm were found to contain small amounts of antigen(s), the concentration of which gradually increased during epididymal transit. In immunocytochemical studies, the antigen was detectable in the epithelial cells from the initial segment to the corpus of the epididymis but not in the caudal cells. In the lumen, the presence of the 135 kDa protein was apparent in the corpus (at a maximum in the middle and distal corpus) and to a lesser degree in the caudal lumen. The 27-kDa protein was distributed all over the equatorial region of the acrosome of less than 10% of caput epididymal sperm. As sperm passed through the corpus epididymis, the percentage of immunoreactive cells increased and the protein was restricted to specific domains of the sperm head. Thus, on the mature sperm, antigen was localized in a crescent-shaped area of the equatorial segment just behind the anterior part of the acrosome and on the apical rim of the sperm head. This is the first observation of a sperm surface antigen derived from an epididymal protein as a proteolytic fragment that interacts with specific regions of the sperm membrane during the process of spermatozoa maturation.

Age Factors↗

The culture of human epididymal epithelium and in vitro maturation of epididymal spermatozoa.

OBJECTIVE: To promote human sperm maturation in vitro. DESIGN: Spermatozoa from the proximal epididymis were coincubated with epididymal epithelial fragments. SETTING: Hospital and research institute. PATIENTS, PARTICIPANTS: Tissue samples were obtained from men undergoing epididymovasostomy procedures or vasectomy. INTERVENTIONS: Fragments of epididymal epithelium formed everted epithelial spheres that in the presence of androgen maintained cell integrity. Coincubation for up to 48 hours of caput epididymal spermatozoa with 3-day-old epithelial cultures from the cauda epididymis was undertaken. MAIN OUTCOME MEASURES: Morphology of epididymal epithelium was assessed by light and electron microscopy. Pulse labeling of tissue in vitro with 35S-methionine was performed with analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis technique and fluorography. Spermatozoa were assessed for progressive motility and their capacity to bind to salt-stored human zona pellucidae. RESULTS: Epididymal fragments formed everted epithelial spheres that maintained cell integrity and functional morphology for 5 to 7 days. Specific proteins were synthesized in culture, in particular, proteins of 20, 22, 40, and 66 kd. Coincubation of caput epididymal spermatozoa with cultures from the cauda epididymis induced a significant increase in progressive sperm motility and sperm binding to salt-stored human zona pellucidae compared with control cultures of epithelium incubated in the absence of androgens or overgrown with fibroblasts. CONCLUSIONS: Aspects of human sperm maturation processes can be mimicked in vitro using coculture techniques with epididymal epithelium. This method may be valuable for improving the fertilizing capacity of human spermatozoa retrieved from the proximal region of the excurrent ducts.

Cells, Cultured↗

Surgical and pathologic observations of epididymal tubules during microscopic epididymal sperm aspiration for intracytoplasmic sperm injection.

Microscopic epididymal sperm aspiration (MESA) for sperm retrieval and intracytoplasmic sperm injection (ICSI) is currently our routine treatment for selected male patients with obstructive azoospermia. In order to refine the surgical technique and obtain better quality sperm for our assisted reproductive technology program, we observed the epididymal tubules in 40 sessions of surgical exploration of the epididymis for sperm aspiration. Epididymal tubules with long-term obstruction could be divided into three groups on the basis of clinical observations and pathology findings: markedly dilated, mildly dilated, and nondilated. All of the markedly dilated epididymal tubules (grade III, n = 10) were azoospermic and ICSI could not be done. Epididymal sperm obtained from the mildly dilated tubules (grade II, n = 9) resulted in poorer fertilization (49%) and pregnancy (33%) rates than sperm obtained from nondilated epididymal tubules (grade I, n = 21, fertilization rate 72%, pregnancy rate 57%). These findings demonstrate that nondilated epididymal tubules are best for sperm retrieval and successful ICSI. We believe this observation will be a good surgical parameter for urologic surgeons performing MESA.

Epididymis↗

Potential contraceptive use of epididymal proteins: immunization of male rats with epididymal protein DE inhibits sperm fusion ability.

Rat epididymal protein DE associates with the sperm surface during maturation and participates in sperm-egg fusion. Immunization of male rats with DE raised specific antibodies and produced a significant reduction in the animals' fertility. The present study focused on determining the in vivo mechanism involved in fertility inhibition. Wistar males were injected with DE, and antibody levels and animal fertility were evaluated. Results revealed an association between the two parameters, since animals with absorbance values lower than 0.5 in ELISA presented high fertility rates (66%, 100%) while those with absorbance values higher than 0.5 exhibited the lowest fertility rates (0%, 33%). Histological studies showed no evidence of orchitis, epididymitis, or vasitis in DE-immunized animals. ELISA results revealed the presence of anti-DE antibodies in epididymal and vas deferential fluids. Indirect immunofluorescence and ELISA experiments indicated that these antibodies would not interfere with the synthesis or secretion of DE or with its association with the sperm surface. Finally, while epididymal sperm recovered from DE-immunized animals presented no changes in motility, viability, or ability to undergo capacitation and acrosome reaction, they exhibited a significant decrease in their ability to fuse with zona-free eggs, with no effect on their ability to bind to the oolemma. Together these results indicate that immunization of male rats with epididymal protein DE specifically interferes with the sperm fertilizing ability, supporting the use of epididymal proteins for contraceptive vaccine development.

Animals↗

Selected proteins of "prostasome-like particles" from epididymal cauda fluid are transferred to epididymal caput spermatozoa in bull.

During epididymal transit, spermatozoa acquire selected proteins secreted by epithelial cells. We recently showed that P25b, a protein with predictive properties for bull fertility, is transferred from prostasome-like particles present in the cauda epididymal fluid (PLPCd) to the sperm surface. To further characterize the interactions between PLPCd and epididymal spermatozoa, PLPCd were prepared by ultracentrifugation of bull epididymal fluid, then surface-exposed proteins were biotinylated and coincubated in different conditions with caput epididymal spermatozoa. Western blot analysis revealed that only selected proteins are transferred from PLPCd to spermatozoa. MALDI-TOF analysis revealed that these transferred proteins are closely related. The pattern of distribution of the PLPCd transferred varied from one sperm cell to the other, with a bias toward the acrosomal cap. This transfer appeared to be temperature sensitive, being more efficient at 32-37 degrees C than at 22 degrees C. Transfer of PLPCd proteins to spermatozoa was also pH dependant, the optimal pH for transfer being 6.0-6.5. The effect of divalent cations on PLPCd protein transfer to caput spermatozoa was investigated. Whereas Mg(2+) and Ca(2+) have no effect on the amount of proteins remaining associated with spermatozoa following coincubation, Zn(2+) had a beneficial effect. These results are discussed with regard to the function of PLPCd in epididymal sperm maturation.

Animals↗

The maintenance of motility and the surface properties of epididymal spermatozoa from bull, rabbit and ram in homologous seminal and epididymal plasma.

Epididymal spermatozoa from bull, rabbit and ram were incubated in homologous epididymal plasma or seminal plasma in a buffered saline-based medium with or without serum albumin. The spermatozoa were either diluted directly into the medium or were washed first. No effect of washing was observed on the subsequent reaction of the cells to the different media. A considerable proportion of the populations of epididymal spermatozoa survived (i.e. continued to exhibit motility) for up to 22 h at 30 degrees C in the simple saline-based medium. Initially epididymal plasma had a slight stimulatory effect on sperm motility in ram and bull but it had no effect on sperm survival in any of the 3 species. Seminal plasma stimulated motility markedly in ram initially, but in all 3 species seminal plasma was detrimental to survival: in ram even a 15-min exposure to the fluid reduced survival. Serum albumin also stimulated motility; it delayed, but did not prevent, the detrimental effect of seminal plasma, although it had no effect itself on survival. The effects of epididymal plasma, seminal plasma and serum albumin on surface properties of epididymal spermatozoa, i.e. agglutination, sticking-to-glass and eosinophilia, were also noted. These varied between species and there was no correlation between these effects and the effects on motility and survival.

Animals↗

Interruption of the luminal flow in the epididymal duct of the corpus epididymidis in the mouse, with special reference to differentiation of the epididymal epithelium.

The efferent duct or the epididymal duct at the border between the caput and corpus epididymidis was cut or ligated in mice at the following stages: birth, 20 days, and 60 days of age. The epididymal duct of the corpus epididymidis was observed at 60 days of age after neonatal or juvenile operation and 4 weeks after the adult operation. After efferent duct interruption, the epididymal duct in the corpus possessed abundant PAS-positive material with no spermatozoa in the lumen, and the principal cells of the epithelium contained PAS-positive inclusions which were considered to be formed due to absorption of the PAS-positive material secreted by the principal cells in the caput epididymidis. Especially after interruption at the border between the caput and corpus epididymidis in juvenile mice, the epithelium of the epididymal duct in the corpus was much taller with peculiar pale, vacuolated cells. These cells were considered as abnormally differentiated forms of the principal cells. The occurrence of the PAS-positive inclusions and peculiar pale cells varied in frequency according to the level and age of interruption of the epididymal duct. The results suggest that the luminal contents produced in the proximal region of the duct may exert an influence on the differentiation of the principal cells of the epididymal duct.

Age Factors↗

Microsurgical epididymal sperm aspiration versus epididymal micropuncture with perivascular nerve stimulation for intracytoplasmic sperm injection to treat unreconstructable obstructive azoospermia.

A novel sperm collection method by epididymal micropuncture combined with perivascular nerve stimulation has been developed to obtain as many clean sperm as possible for IVF for patients with surgically irreparable vasal obstruction. To assess whether the new technique could improve the fertilization and pregnancy rates obtained when attempting microsurgical epididymal sperm aspiration (MESA) to retrieve epididymal sperm from such patients, a prospective randomized comparative study was conducted. Twenty-nine cycles of conventional MESA with ICSI were performed on 25 couples with congenital bilateral absence of the vas deferens (CBAVD) and four failed vasovasostomy cases (group 1). Thirty cycles of epididymal micropuncture with nerve stimulation with ICSI were performed on 28 couples with CBAVD and two failed epididymovasostomy cases (group 2). The mean volume of epididymal fluid and sperm motility in group 2 was significantly higher than that in group 1 (p < .001). Both fertilization and pregnancy rates in group 2 were significantly higher than those in group 1 (p < .001 and p < .03). This novel epididymal sperm collection method for ICSI can provide significantly higher fertilization and pregnancy rates than conventional MESA for ICSI.

Cytoplasm↗

Comparison of the outcome of intracytoplasmic sperm injection using fresh and frozen-thawed epididymal spermatozoa obtained by percutaneous epididymal sperm aspiration.

The study was conducted to compare the outcome of intracytoplasmic sperm injection (ICSI) between fresh and frozen-thawed epididymal spermatozoa retrieved by percutaneous epididymal sperm aspiration (PESA) from obstructive azoospermic men. Overall, 53 PESA procedures were performed in 42 obstructive azoospermic men, followed by ICSI procedures with either fresh (n = 40) or frozen-thawed (n = 13) epididymal spermatozoa. Comparing all ICSI cycles with fresh and frozen-thawed epididymal spermatozoa, the fertilization rates (77.4 vs 86.8%) and the cleavage rate (91.3 vs 95.1%) were not statistically different. A total of 64 embryo transfers were performed: 48 embryo transfers after the use of fresh epididymal spermatozoa and 16 embryo transfers after the use of frozen-thawed spermatozoa. The overall pregnancy rate per embryo transfer was also similar between both groups (40.8 vs 40%). The implantation rate per embryo (18.2 vs 12.7%), clinical pregnancy per embryo transfer (36.7 vs 33.3%) and delivery/ongoing pregnancy rate (36.7 vs 33.3%) were not statistically different. In conclusion, there were no significant differences of the outcome of intracytoplasmic sperm injection using fresh and frozen-thawed epididymal spermatozoa obtained by PESA.

Adult↗

Interaction of the human epididymal protein CD52 (HE5) with epididymal spermatozoa from men and cynomolgus monkeys.

A monoclonal antibody (CAMPATH-1 G) against the human lymphocyte surface protein CD52, which is similar to the epididymal secretion HE5, was used to ascertain the presence of this protein on maturing primate spermatozoa by flow cytometry. The percentage of human viable spermatozoa stained specifically with this antibody increased from sperm in spermatocoeles (0.5%), to the efferent ducts (3.8%), corpus (47.2%), and cauda (85.7%) epididymidis. Positive cells revealed staining mainly over the whole tail and postacrosomal region of the sperm head. Spermatozoa (approximately 10%) from both the efferent ducts and corpus epididymidis took up additional antigen when incubated with human distal cauda epididymidal plasma as a source of CD52, and 12-22% of human testicular sperm (from spermatocoeles) took up CD52 from human seminal plasma. In the cynomolgus monkey, nonspecific binding of control IgG was greater than that in human males and net CD52 staining was measurable only on approximately 30% of corpus sperm where it was mainly on the principal piece. Neither caput nor cauda sperm took up human CD52 upon incubation with human seminal plasma, but an additional 27% of corpus sperm expressed CD52. Such uptake of CD52 was drastically reduced, or did not occur, when seminal plasma had been fractionated by filtration through 0.1 microns filters (filtrate II) or 300,000 Da cutoff filters (filtrate III), respectively. Western blots revealed that CD52 contents were much reduced in filtrate II and nondetectable in filtrate III of seminal plasma. Similar reduction of CD52 in the filtrate of cauda epididymidal plasma indicates the association of this epididymal secretion with large molecular factors and suggests their involvement as carriers in the in vivo transfer of the secretion onto the epididymal sperm surface. The in vitro uptake of CD52 by some but not all immature sperm and the detection by Western blotting of much less CD52 in the corpus than the cauda luminal plasma suggest that the acquisition of this epididymal secretion by spermatozoa depends on their maturation status as well as the availability of the protein in the epididymal lumen.

Animals↗

Epididymal sperm retrieval by epididymal micropuncture combined with intracytoplasmic sperm injection: difference between acquired and congenital irreparable obstructive azoospermia.

To evaluate the differences in fertilization and pregnancy rates following intracytoplasmic sperm injection (ICSI) of retrieved epididymal sperm between congenital (group 1) and acquired (group 2) unreconstructable obstructive azoospermic patients, we compared the outcome of the ICSI procedure between these two groups. Thirty-six patients with obstructive azoospermia received epididymal sperm retrieval by the micropuncture method for the ICSI procedure. Main parameters evaluated were epididymal fluid volume, sperm concentration, sperm-motility; fertilization rate and clinical pregnancy rate. There were no significant differences in epididymal fluid and sperm concentration between the two groups. However, the sperm motility in group 1 was significantly higher than that in group 2. The fertilization rates per couple in groups 1 and 2 were 78.2 and 82% (nonsignificant). The pregnancy rate per couple in group 1 was 37.5% (6/16), while that in group 2 was only 5% (1/20); this difference was statistically significant (p < 0.05). Using epididymal sperm for the ICSI procedure, the chances of pregnancy for couples with congenital absence of the vas deferens were significantly higher compared with couples with acquired unreconstructable obstructive male infertility.

Adult↗

Analysis of surface proteins of rat spermatozoa during epididymal transit and identification of antigens common to spermatozoa, rete testis fluid and cauda epididymal plasma.

Spermatozoa from the testis and cauda epididymidis of the rat were surface labelled with radioactive iodide, extracted with detergent, and the radioactive proteins separated by two-dimensional polyacrylamide gel electrophoresis. In some instances spermatozoa were also surface labelled with tritiated borohydride in the presence of galactose oxidase. Soluble proteins in blood serum, rete testis fluid and cauda epididymal plasma were also iodinated and separated by gel electrophoresis. In addition, aliquants of the radioactive sperm extracts, blood serum and reproductive tract fluids were each immunoprecipitated with polyspecific antisera directed against either testicular sperm membranes, caudal sperm membranes, blood serum, rete testis fluid or cauda epididymal plasma before gel electrophoresis. From the patterns of radioactive proteins detected on the resultant gels, a two-dimensional map was created for each of the sperm extracts and for the various fluids. Proteins which were nonhomologous between testicular and caudal spermatozoa were identified, as well as proteins which were common to spermatozoa and reproductive tract fluids. Epididymal transit was characterized by the loss of certain proteins from the sperm surface, including three borohydride-labelled proteins of Mr 130 000, and by the addition of others, most notably a highly abundant protein of Mr 42 000. Several of the proteins lost from spermatozoa accumulated in the epididymal plasma whilst some of those added to the sperm surface could be identified as direct secretory products of the epididymis. Rete testis fluid contained blood proteins in addition to others presumed to be testis-specific, whilst the composition of cauda epididymal plasma was markedly different from blood serum or rete testis fluid.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

In vitro culture of brushtail possum (Trichosurus vulpecula) epididymal epithelium and induction of epididymal sperm maturation in co-culture.

A medium modified from eutherian systems was used to culture epididymal epithelial cells of the brushtail possum (Trichosurus vulpecula) for more than 2 months. Epididymal tubule fragments from the caput, corpus and cauda epididymides were used to generate cell monolayers. All three epididymal cell culture systems supported maturational changes in marsupial spermatozoa and enabled immature possum spermatozoa to differentiate from a T-shape to a streamlined shape, accompanied by the development of progressive motility after co-culture with 7-day-old cultured epididymal cell monolayers. This epididymal cell and sperm co-culture system for marsupial species may facilitate the identification of specific epithelial factors that affect sperm maturation, particularly in a species in which morphological maturation is readily visible.

Animals↗