Residual Sperm Research Articles

Significación clínica de la presencia de espermatozoides en los seminogramas posvasectomía. Análisis de una serie de 2.168 pacientes

IntroductionVasectomy is a safe and effective technique to achieve azoospermia, although the failure rate of the technique is less than 1%. Sterility is not immediate so the post-vasectomy seminogram continues o be essential to ensure the success of the technique. The aim of this trial is to establish the attitude when dealing with immobile residual sperm patients. Material and methodsCross-sectional analysis of 2,168 vasectomies performed between January 2010 and March 2017. The first post-vasectomy seminogram was performed at 3 months. Those patients with azoospermia did not undergo further controls. Patients with immobile sperm (<100,000/ml o>100,000/ml) were considered potentially fertile and were followed with monthly seminograms until azoospermia was obtained. ResultsOf a total of 1,807 patients were included; 1,297 of these had azoospermia at 3 months seminogram and 501 patients had immobile residual sperm. Only 24 patients of this last group showed more than 100.000 sperm/ml; 9 cases showed mobile sperm. All patients who presented immobile residual sperm underwent serial seminograms. Azoospermia was achieved in an average time of 4,5 months in a rage of 4-10 months, regardless of the initial sperm count. An average of 2,5 tests were performed on each patient. All of the patients with mobile sperm required a reintervention. ConclusionAll patients with immobile sperm on the first post-vasectomy seminogram will achieve azoospermia regardless of the initial count. Therefore, serial controls until a negative seminogram is obtained are unnecessary.

Circulatory exosomal tRF-Glu-CTC-005 and tRF-Gly-GCC-002 serve as predictive factors of successful microdissection testicular sperm extraction in patients with nonobstructive azoospermia

To identify circulating plasma exosomal transfer RNA-derived fragments (tRFs) as the predictive factors of successful microdissection testicular sperm extraction (micro-TESE) in patients with nonobstructive azoospermia (NOA). Case and control prospective study. Academic research laboratory. Twelve patients with NOA with successful sperm retrieval by micro-TESE, 18 patients with NOA with failed sperm retrieval by micro-TESE, and 12 normozoospermic fertile controls. Blood samples were collected from participants. The abundance of tRFs normalized as counts per million of the total aligned reads with the next-generation sequencing system; candidate tRF levels were validated through quantitative reverse transcription polymerase chain reaction; predictive accuracy was evaluated by the receiver operating characteristic area under the curve analysis. The nomogram was built for ranking. The plasma circulating exosomal tRF-Gly-GCC-002 and tRF-Glu-CTC-005 manifested the most confident differential expression between patients with NOA with successful sperm retrieval by micro-TESE and patients with NOA with failed sperm retrieval by micro-TESE. The target gene prediction of these 2 tRFs followed by the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis indicated the functional enrichment of neuroendocrine protein metabolism and striatum/subpallium development. The herpes simplex virus 1 infection pathway was also involved. The receiver operating characteristic area under the curve (AUC) analysis demonstrated a promising predictive accuracy: tRF-Gly-GCC-002, AUC of 0.921, and tRF-Glu-CTC-005, AUC of 0.954. A regression model was built and presented with the nomogram for further assessment. This study described the exosomal tRF-Gly-GCC-002 and tRF-Glu-CTC-005 expression values, indicated a promising predictive effect for accessibility of sperm retrieval through micro-TESE from patients with NOA, and highlighted tRF-Gly-GCC-002 and tRF-Glu-CTC-005 as useful biomarkers in patients with NOA seeking invitro conception with their residual sperm.

Induction of precocious females in the protandrous barramundi (Lates calcarifer) through implants containing 17β-estradiol - effects on gonadal morphology, gene expression and DNA methylation of key sex genes

Sex control is vital for the efficient breeding of aquaculture species. Barramundi (Lates calcarifer) is a protandrous sequential hermaphrodite that naturally sex change from male to female over its lifespan. The induction of precocious female barramundi will permit breeding of males and females of the same-generation individuals in selective breeding programs, increasing the rates of genetic gain, while reducing infrastructure costs. Accordingly, the efficacy of two dosages of 17β-estradiol (E2) delivered via implants to induce the precocious female barramundi were evaluated. Six-month-old male barramundi (405 ± 50 g body weight (BW)) were given a single cholesterol-based pellet implant containing either 0 mg E2 kg−1 BW (untreated control), 4 mg E2 kg−1 BW (‘low dose’), or 8 mg E2 kg−1 BW (‘high dose’). Changes in gonadal morphology and liver condition of implanted males, along with RT-qPCR and bisulfite amplicon sequencing to quantify expression profiles and DNA methylation of key male-female sex-related genes were then examined after 9 weeks post-implantation. Results showed that at 9 weeks post-implantation, in the ‘high dose’ E2 treatment group, 78% (7/9) of fish sex-changed completely to female, signified by gonads containing oocytes (20–30 μm) and no observed residual sperm. Comparably, 44% (4/9) of fish in the ‘low dose’ E2 treatment group had sex-changed, while remaining fish showed complete testicular regression with gonads containing only undifferentiated germ cells. In the ‘high dose’ E2 treatment, upregulation of female-biased genes (cyp19a1a and foxl2) and downregulation of male-biased genes (dmrt1, cyp11b and esr1) were observed. Increased gene expression was accompanied by decreased DNA methylation in cyp19a1a, but no significant changes in DNA methylation of foxl2 or esr1 were observed. The success of artificially-induced sex change in barramundi provides an important tool that is critical to improving selective breeding of this species.

In vitro survival kinetics of microfluidic-sorted bovine spermatozoa.

The isolation and characterization of sperm subpopulations that can achieve fertilization is a major challenge of assisted reproduction methods. We focused on the microfluidic sperm sorter as a novel tool for collecting highly motile spermatozoa from heterogeneous semen samples. This study primarily aims to obtain baseline information on sorted spermatozoa according to its characteristics and in vitro life span. Frozen-thawed bull semen was subjected to microfluidic sperm sorting using diffuser-type microfluidic sperm sorter (DMSS). After sorting, samples were collected as the sorted spermatozoa and unsorted residual spermatozoa and incubated at 37°C for subsequent evaluation. The samples were assessed at different time points (0 or 1, 6, and 24h) in terms of motility, which was measured by computer-assisted sperm analysis (CASA), membrane integrity, mitochondrial function, and adenosine triphosphate (ATP) production after sorting (0h). To determine the characteristics and efficiency of DMSS sorting, the sorted spermatozoa were compared with samples collected using the swim-up method, a conventional method in motile sperm selection. A comparison between the sorted and residual spermatozoa demonstrated significantly higher motility parameters, membrane integrity, and mitochondrial function of the sorted spermatozoa until 6h after incubation. The time course decrement of membrane and mitochondrial status were subjected to curve fitting and theoretically supported. Sperm ATP production measured immediately after sorting showed higher ATP generation of the sorted spermatozoa compared with the unsorted, frozen-thawed spermatozoa. The motility parameters and mitochondrial activity of DMSS-sorted spermatozoa were higher than the swim-up-collected spermatozoa (p<0.05). These results indicate that DMSS sorting can strictly select highly motile spermatozoa with the ability to maintain its membrane integrity and mitochondrial function related to ATP production. We speculate that the device that is able to sort high-quality spermatozoa can have great potential in assisted reproduction.

Can cover sheath model influence semen retention in AI-gun trials and pregnancy rates of cows inseminated at a fixed-time?

The achievement of satisfactory results in fixed-time artificial insemination (FTAI) protocols depends on several factors, such as the quality of the materials used to perform the artificial insemination (AI). In this context, three experiments were conducted to evaluate the effect of the cover sheath model on the residual retention of semen during AI and the pregnancy rates of cows submitted to FTAI. In Experiment 1, 400 straws of cryopreserved bovine semen were thawed and repacked in samples with similar weight and volume characteristics. Each new dose of semen (n = 300) was mounted using one of three AI cover sheath models (n = 100 semen/sheath doses): the main brand in the global market (G1), the main model in the Brazilian market (G2), and a model marketed as having a low residual semen retention rate (G3), to determine the percentages of seminal elimination (PSEs) during AI. In Experiment 2, 464 Nelore cows were synchronized through intravaginal progesterone implants and inseminated using the same cover sheaths as those tested in Experiment 1, which were grouped into classes (conventional: G1 and G2 [n = 225]; and low reflux: G3 [n = 239]). In Experiment 3, 859 Nelore cows were synchronized and inseminated as described above, using the same cover sheaths previously tested. After each AI, cover sheaths were visually inspected for the presence of residual semen and pregnancy diagnosis was performed 35 days after FTAI (Experiments 2 and 3). The PSEs in Experiment 1 were 91.7%, 90.6% and 96.5% for groups G1, G2 and G3, respectively (P = 0.05). The pregnancy rates in Experiment 2 were 53.33% and 58.16% for the conventional and low reflux model, respectively (P > 0.05), but there were differences (P = 0.05) among the cover sheath models employed for AI (G1 - 57.72% [71/123]; G2 - 48.04% [49/102]; G3 - 58.16% [139/239]). Additionally, a lower semen reflux incidence (P < 0.05) was observed in the low residual sperm retention model (0.72%) in Experiment 3, compared to the main international (10.42%) and national (22.99%) market models. In conclusion, residual semen retention occurs regardless of the cover sheath model used for AI. However, the PSE is influenced by the model and quality of cover sheath used. According to the results, the cover sheath represents a factor that may compromise the pregnancy rates of synchronized cows for FTAI.

The effect of sperm miR-34c on human embryonic development kinetics and clinical outcomes

AimsWe aimed to investigate the effect of sperm miR-34c on early human embryonic development kinetics and clinical outcomes of in vitro fertilization (IVF) patients. Materials and methodsAfter oocyte insemination, residual sperm specimens were collected from 58 patients undergoing IVF. miR-34c expression levels in sperm, oocytes, zygotes, and embryos/blastocysts were detected with qRT-PCR, and embryonic development kinetics were observed using time-lapse technology. To confirm the role of miR-34c in regulation of early embryonic development, miR-34c siRNA was injected into zygotes obtained from in vitro-matured oocytes. A ROC curve was used to determine the cutoff value. Comparisons of embryonic development kinetics and clinical outcomes were performed according to the cutoff value. Key findingsThe miR-34c expression level was highest in 3PN zygotes, but was not expressed in human oocytes. In the miR-34c siRNA group, embryonic development kinetic parameters t2, t3, t4, and t5 were significantly prolonged, but the cleavage rate and high-quality embryo rate were lower than in the control group. The levels of sperm miR-34c were negatively correlated with t5 and positively correlated with rates of blastocyst formation, high-quality blastocysts, and pregnancy. The miR-34c levels and the blastocyst formation rate were higher in the pregnancy group (p < 0.05). Logistic regression analysis showed that sperm miR-34c level was significantly correlated with pregnancy (OR: 5.056, 95% CI: 1.560–16.384; p = 0.007). SignificanceThe sperm miR-34c expression level is associated with embryonic development kinetics and clinical outcomes. Thus, miR-34c expression is beneficial to embryonic development and may be used as an indicator of IVF outcomes.

In vivo cellular evidence of autophagic associated spermiophagy within the principal cells during sperm storage in epididymis of the turtle.

The epididymis plays a significant role as a quality control organ for long-term sperm storage, maturation, and fertilizing ability and perform filtration function to eliminate abnormal or residual spermatozoa by phagocytosis. However, the role of autophagy in spermiophagy during sperm storage in turtle epididymis still needs to be studied. In this study, we reported in vivo spermiophagy via the cellular evidence of lysosome engulfment and autophagy within the principal cells during sperm storage in the turtle epididymis. Using immunofluorescence, Lysosome associated membrane protein-1 (LAMP1) and microtubule-associate protein light chain 3 (LC3) showed strong immunosignals within the apical cytoplasm of epididymal epithelia during hibernation than non-hibernation. Co-immunolabeling of LAMP1 and LC3 was strong around the phagocytosed spermatozoa in the epididymal epithelia and protein signaling of LAMP1 and LC3 was confirmed by western blotting. During hibernation, ultrastructure showed epididymal principal cells were involved in spermiophagy and characterized by the membrane’s concentric layers around phagocytosed segments of spermatozoa, degenerative changes in the sperm head and lysosome direct attachment, and with the existence of cellular components related to autophagy (autophagosome, autolysosome). In conclusion, spermiophagy occurs by lysosomal engulfment and autophagic activity within the principal cells of the turtle epididymis during sperm storage.

Histological evidence of the lunar reproductive rhythm of Atlantic sea urchin Arbacia punctulata in the Gulf of Mexico

ABSTRACT Natural phenomena act as external cues that stimulate the reproductive activity of many marine organisms. In this study, we report the gonadal maturation in relation to the lunar cycle of wild-caught Atlantic sea urchin, Arbacia punctulata (Echinodermata: Arbaciidae, an edible marine invertebrate), in the Gulf of Mexico. Weekly changes in gonadal development were observed histologically according to the lunar cycle during the summer months from May to July. Histological observations revealed that testes were developed synchronously around the new moon in June and July. Several testes collected in the same moon phase contained residual sperm (~7%) or testicular lobules/acini fully occupied by mature sperm (~46%). Cyclic ovarian development was also observed during the summer months, some ovaries collected around the new moon in June and July contained oocytes at maturation stage (ova: ~39-71%) or empty follicles, suggesting that the Atlantic sea urchin spawns according to the lunar cycle in the Gulf of Mexico. To the best of our knowledge, this is the first report on the lunar reproductive rhythm of Arbacia species in the Gulf of Mexico.

Effective removal of residual sperm from rearing tank using a foam separation device

Effective removal of residual sperm from rearing tank using a foam separation device

Hyaluronic Acid Binding Sperm Selection for assisted reproduction treatment (HABSelect): study protocol for a multicentre randomised controlled trial

IntroductionThe selection of a sperm with good genomic integrity is an important consideration for improving intracytoplasmic sperm injection (ICSI) outcome. Current convention selects sperm by vigour and morphology, but preliminary...

Developmental Phases of the Seminal Vesicle related to the Spermatogenic Stages in the Testicular Lobules of Neptunea (Barbitonia) cumingii (Gastropoda: Buccinidae).

Cytological changes of the epithelial cells according to the developmenatal phases of the seminal vesicle related to the spermatogenic stages in the testicular lobules during spermagenesis in male Neptunea (Barbitonia) cumingii (Gastropoda: Buccinidae) were investigated monthly by electron microscopical and histological observations. N. (B) cumingii is dioecious, and an internal fertilization species. The male genital organ is located near the tentacles. The spermatozoon is approximatley 50 μm in length. The axoneme of the tail flagellum consists of nine pairs of microtubles at the periphery and one pair at the center. The process of germ cell development during spermatogenesis can be divided into five succesive stages: (1) spermatogonia, (2) primary spermatocytes, (3) secondary spermatocytes, (4) spermatids, and (5) spermatozoa. A considerable amount of spermatozoa make their appearance in the testicular lobules (or acini) and some of them are tranported from the testis towards the seminal vesicles until late July. In this study, the developmental phases of the epithelial cells of the seminal vesicles of N. (B.) cumingii could be classified into four phases: (1) S-I phase (resting), (2) S-Ⅱphase (early accumulating), (3) S-Ⅲ phase (accumulating), and (4) S-IV phase (spent). However, in case of N. (B.) arthritica cumingii, the developmental phases of the seminal vesicle were devided into three phases: (1) resting, (2) accumulating and (3) spent. Granular bodies in the inner layer of the seminal vesicles are involved in resorption of digestion of residual spermatozoa.

CTCF contributes in a critical way to spermatogenesis and male fertility.

The CCCTC-binding factor (CTCF) is an architectural protein that governs chromatin organization and gene expression in somatic cells. Here, we show that CTCF regulates chromatin compaction necessary for packaging of the paternal genome into mature sperm. Inactivation of Ctcf in male germ cells in mice (Ctcf-cKO mice) resulted in impaired spermiogenesis and infertility. Residual spermatozoa in Ctcf-cKO mice displayed abnormal head morphology, aberrant chromatin compaction, impaired protamine 1 incorporation into chromatin and accelerated histone depletion. Thus, CTCF regulates chromatin organization during spermiogenesis, contributing to the functional organization of mature sperm.

피뿔고둥 Rapana venosa (Valencienes) 정소소엽 내에서의 생식세포 발달과 정자형성과정 중 출현하는 비정형 세포들의 미세구조적 연구 및 저정낭의 발달단계에 따른 상피세포들의 주기적 변화

피뿔고둥, Rapana venosa의 정소소엽 내에서 정자형성과정 중에 생성되는 정상적인 생식세포의 발달과 함께 섞이어 일정 시기에만 출현하는 비정형세포들을 미세구조적으로 관찰하였고, 또한 정소 발달단계에 따른 저정낭 내층 상피세포들의 주기적 변화를 조직학적 관찰에 의해 조사하였다. 생식세포의 발달단계는 정원세포기, 정모세포기, 정세포기, 정자기로 나누어지며, 정모세포기는 다시 제1 정모세포기와 제2 정모세포기로 세분할 수 있어, 총 5 단계로 구분할 수 있었다. 정상적인 웅성생식세포들의 분화와 발달과정은 다른 복족류 종들과 유사하였다. 정자는 길이가 대략 <TEX>$50{\mu}m$</TEX> 정도이었다. 정자의 미부 편모의 악소님 (axoneme) 은 주변에 9쌍의 미세소관들과 중앙에 1쌍의 미세소관들로 구성되어 있다. 즉, 9+2 구조를 이루고 있다. 특히, 대형 복족류 중 뿔소라과 피뿔고둥의 경우는 예외적으로 다른 이매패류나 두족류 등과 달리 정소소엽 내에서 정자형성과정 중에 정상적인 생식세포들 사이에서 총 4종 (Type IA, IB, Type IIA, IIB) 의 비정형세포들 (atypical cells) 이 함께 출현하는 특징을 보이고 있는데, 이러한 현상은 대형 복족류의 단지 소수의 종들에 한하여 출현하는 예외적인 특이한 현상이라 할 수 있다. 그러나 비정형세포들은 저정낭의 여러 단계 중 내층 상피세포들 내에서는 발견되지 않았다. 추측컨대 몇 가지 비정형세포들은 리소좀-모양의 공포들이나 리소좀-모양의 소체들을 가지는데, 이들은 정소소엽 내에서 붕괴나 그 자신들의 흡수에 관여하는 것으로 추정된다. 상당량의 정자들이 정소소엽 내에서 형성되어, 그들의 일부는 7월 말까지 정소에서 저정낭으로 이동된다. 교미 성기는 6-7 월 사이이었다. 피뿔고둥 저정낭 발달단계의 주기적 변화는 (1) S-1 단계 (휴지단계), (2) S-II 단계 (축적단계), 그리고 S-III 단계 (배정단계) 의 3 단계로 구분되었다. Germ cell development and cyclic changes in the epithelial cells of the seminal vesicle of the male rapa whelk, Rapana venosa, were investigated by cytological and histological observations. The process of germ cell development can be classified into five stages: (1) spermatogonial, (2) primary spermatocyte, (3) secodary spermatocyte, (4) spermatid, and (5) spermatozoon. In particular, four atypical cells (Type IA, IB, IIA and IIB cells) occur among normal germ cells in the acini during spermatogenesis. Presumably, the atypical cells, which have lysosome-like vacuoles or lysosome-like bodies in the cells, are involved in breakdown and absorption themselves in the acini. However, atypical cells were not found in the epithelial cells of the inner layer of the seminal vesicle. A considerable amount of spermatozoa are transported from the testis towards the the seminal vesicles until late July. The main coupulation period is between June and July. The process of the cyclical changes of the seminal vesicles can be classified into three phases: (1) resting, (2) accumulating, and (3) spent. Yellow granular bodies are involved in resorption or digestion of residual spermatozoa.

Aneuploidy Screening in Human Preimplantation Embryos Via Array Comparative Genomic Hybridization (ACGH) – Can Extraneous Sperm Affect Diagnosis?

ICSI is often recommended for PGS in order to prevent contamination caused by DNA from residual sperm attached to the zona pellucida. Contamination can lead to PGS misdiagnosis. Due to its characteristics, array comparative genomic hybridization (aCGH) is presumed to be more impervious to contamination than other methods, perhaps allowing regular in vitro fertilization (IVF) using standard insemination for fertilization.

Correlation analysis of the progesterone-induced sperm acrosome reaction rate and the fertilisation rate in vitro.

In this study, we aimed to investigate whether progesterone-induced acrosome reaction (AR) rate could be an indicator for fertilisation rate in vitro. Twenty-six couples with unexplained infertility and undergoing in vitro fertilisation (IVF) treatment were involved. On the oocytes retrieval day after routine IVF, residual sperm samples were collected to receive progesterone induction (progesterone group) or not (control group). AR rate was calculated and fertilisation rate was recorded. The correlation between progesterone-induced AR and fertilisation rate and between sperm normal morphology and 3PN (tripronuclear) were analysed using the Spearman correlation analysis. The AR rate of progesterone group was statistically higher than that of the control group (15.6±5.88% versus 9.66±5.771%, P<0.05), but not significantly correlated with fertilisation rate (r=-0.053, P>0.01) or rate of high-quality embryo development (r=-0.055, P>0.01). Normal sperm morphology also showed no significant correlation with the amount of 3PN zygotes (r=0.029, P>0.01), rate of 3PN zygotes production (r=0.20, P>0.01), rate of 3PN embryo development (r=-0.406, P>0.01), fertilisation rate (r=-0.148, P>0.01) or progesterone-induced AR rate (r=0.214, P>0.01). Progesterone can induce AR in vitro significantly; however, the progesterone-induced AR may not be used to indicate fertilisation rate.

High-resolution helium ion microscopy of epididymal epithelial cells and their interaction with spermatozoa.

We examined the rat and mouse epididymis using helium ion microscopy (HIM), a novel imaging technology that uses a scanning beam of He(+) ions to produce nanometer resolution images of uncoated biological samples. Various tissue fixation, sectioning and dehydration methods were evaluated for their ability to preserve tissue architecture. The cauda epididymidis was luminally perfused in vivo to remove most spermatozoa and the apical surface of the epithelial lining was exposed. Fixed epididymis samples were then subjected to critical point drying (CPD) and HIM. Apical stereocilia in principal cells and smaller apical membrane extensions in clear cells were clearly distinguishable in both rat and mouse epididymis using this technology. After perfusion with an activating solution containing CPT-cAMP, a permeant analog of cAMP, clear cells exhibited an increase in the number and size of membrane ruffles or microplicae. In contrast, principal cells did not exhibit detectable structural modifications. High-resolution HIM imaging clearly showed the ultrastructure of residual sperm cells, including the presence of concentric rings on the midpiece, and of cytoplasmic droplets in some spermatozoa. Close epithelium-sperm interactions were also detected. We found a number of sperm cells whose heads were anchored within the epididymal epithelium. In certain cases, the surface of the sperm cytoplasmic droplet was covered with vesicle-like structures whose size is consistent with that of epididymosomes. In conclusion, we describe here the first application of HIM technology to the study of the structure and morphology of the rodent epididymis. HIM technology represents a major imaging breakthrough that can be successfully applied to study the epididymis and spermatozoa, with the goal of advancing our understanding of their structure and function.

Spermatangium formation and sperm discharge in the Japanese pygmy squid Idiosepius paradoxus

In cephalopods, sperm discharge is an important event not only for sperm transfer but also influencing sperm storage capacity of attached spermatangia (everted spermatophores). To investigate sperm discharge from spermatangia and the condition of naturally attached spermatangia in Japanese pygmy squid (Idiosepius paradoxus) we (i) investigated the morphology of spermatophores and spermatangia, and the process of spermatophore evagination and sperm discharge from spermatangia obtained in vitro; (ii) observed spermatangia that were naturally attached to female squids at 6, 12, 18, 24 and 48h after copulation to investigate alterations in naturally attached spermatangia with time. The spermatophore of I. paradoxus is slender and cylindrical and consists of a sperm mass, a cement body and an ejaculatory apparatus, which is similar to those of loliginid squids. The spermatangium is fishhook-shaped, its distal end being open and narrow. After the spermatangium is formed, the sperm mass gradually moves to the open end of the spermatangium, from where sperm are released. Sperm discharge is a rapid process immediately after the beginning of sperm release, but within 5min changes to an intermittent release of sperm. Although the volume of residual spermatozoa differed among spermatangia that were naturally attached to a single individual, the probability that spermatangia would be empty increased with time. Most naturally attached spermatangia discharged almost all of their spermatozoa within 24h after copulation, and no spermatangia were attached to females 48h after copulation. These results suggest that sperm transfer from the spermatangium to the seminal receptacle must occur within 24h, and that the spermatangium functions as a transient sperm storage organ in pygmy squids.

Ultrastructure of the semicystic spermatogenesis in the South American freshwater characid Hemigrammus marginatus (Teleostei, Characiformes)

Semicystic, a rare type of spermatogenesis, was detected in the characid Hemigrammus marginatus and characterized by cysts hatching during the spermatid phase and maturation of the spermatozoa being completed at the lumen of the anastomosed seminiferous tubules. Primary spermatogonia, or type A, are distributed along the entire length of the seminiferous tubules, in an unrestricted spermatogonial pattern. H. marginatus spermiogenesis is included in type I, mainly characterized by presence of nucleus rotation. During this process, a vesicle resembling the acrosomal vesicle is visualized at the anterior region close to the nucleus of the early spermatids, however this structure did not remain in the spermatozoa. In H. marginatus, the spermatozoon is uniflagellated, primitive, type I aquasperm, with a rounded head, a short midpiece and a long flagellum with the axoneme in a 9 + 2 microtubules arrangement and no lateral fins. Residual spermatozoa are reabsorbed by Sertoli cells. Unusual biflagellate spermatozoa with three long cytoplasmatic projections originating in the midpiece are rarely observed and have not been registered in other characiforms. Ultrastructural characteristics of the spermatogenesis and spermatozoa observed in the present work provide important subsidies to systematic and phylogeny studies of Characidae fishes included in Incertae sedis groups, such as H. marginatus.

The effect of diets with different levels of protein and energy on the process of final maturation of the gametes of Rhamdia quelen stocked in cages

The influence of the interaction between the nutrition of tropical freshwater broodfishes when stocked in an intensive system (cages) and their reproductive cycle has received little attention, and the results obtained until now have been very conflicting. One of the main nutritional demands for any species of fish is the source of energy and protein, which is essential for its maintenance, growth and reproduction. This study was conducted with 6300 “jundiá”, Rhamdia quelen juveniles (47.0 ± 5.7 g and 17.3 ± 2.1 cm), at Itaipu Power Station, in the town of Santa Helena, Paraná, Brazil, in order to assess the different levels of crude protein (CP) and digestible energy (DE) (A: 25% CP; B: 30% CP; C: 35% CP, with 3250 kcal g − 1 and D: 25% CP; E: 30% CP; F: 35% CP, with 3500 kcal g − 1 ) during their first reproductive cycle. Six treatments and three replicates were applied to a total of 18 cages (5.0 m 3) with 70 fishes m − 3 each, from January to December/2006. Fishes from each cage were randomly captured, measured ( Lt, cm), weighed ( Wt, g), and had their gonads (Wg, g) weighed to determine the condition factor ( K) and gonado-somatic ( GSI) indices. The histomorphological appearance (light microscopy) of the gonads was not affected by the diets of treatments A, B, D, E, or F. However, the highest mean value of GSI was 15% in treatment C (35% CP and 3250 kcal g − 1 ), differently from the other treatments ( p < 0.05). Nevertheless, it could be noticed that in treatment C the mean values of K were very similar for both sexes and had significant differences during the experiment ( p < 0.05). The asynchronous ovary contained oocytes at all stages of development and had potential to perform fractional spawning up to four times a year (treatment C), with the first maturation beginning in July/06 (28.6 ± 1.4 cm), and then in September (33.2 ± 0.5 cm), October (32.5 ± 2.7 cm) and December (34.4 ± 3.4 cm). The fishes that had received the other diets exhibited only three spawning peaks/year, in September (28.0 ± 4.8 cm), October (28.4 ± 7.6 cm) and December (32.9 ± 2.0 cm). Nevertheless, the males started their maturation process when they were two months old (17.6 ± 1.9 cm), and continued progressively during spermatogenesis, reaching the peak of maturation at the beginning of spermiation. During the regression phase, residual spermatozoa could be observed together with the decrease of the mean values of GSI.

Examinations of polyvidone-iodine as an irrigation solution for spermatic duct occlusions.

In vitro experiments with human sperm as well as animal experiments were carried out to test the applicability of polyvidone-iodine as an irrigation solution for spermatic duct occlusions. The substance polyvidone-iodine was initially chosen on the basis of spermiograms because of its ability to immobilize spermatozoa, and then its spermicidal effect was clearly proven by means of eosin vital staining. The aim of the animal experiments was to examine the substance for possible histotoxic side effects and to ascertain the retention of residual sperm in the ampulla area of the vas deferens. Polyvidone-iodine fulfills all the required criteria as an irrigation solution: it is definitely spermicidal with no long-term histotoxic side effects; in addition, it acts as a disinfectant and is easily obtainable as well as inexpensive. Postoperative azoospermia is attained in animal experiments after four weeks at the most.