Computer assisted morphometric analysis of ram sperm heads: Evaluation of different fixative techniques

The recent development of automated systems for morphometric sperm head analysis has provided a series of objective parameters which have facilitated the standardization of morphological semen evaluation. This current work attempts to establish the optimum fixing conditions for the morphometric characterization of ram spermatozoa. Ejaculates were obtained from 5 Merino rams used for periodic collection of semen and were diluted at 1:50 with TEST medium. Air-dried smears were fixed either in ethanol-ether (1:1), 50% methanol, 2% glutaraldehyde or SUZA fixative, in which case the smear was pretreated with chloramine. The samples were then stained with commercial kit Hemacolor. Once the prepar…

Determining the relationship between bull sperm kinematic subpopulations and fluorescence groups using an integrated sperm quality analysis technique.

The aim of the present study was to determine whether there is an association between the kinematic sperm subpopulations and fluorescent groups in bulls using a new fluorescent staining method that allows classification of spermatozoa into groups depending on their acrosomal and membrane integrity, as well as functional status, without inhibiting sperm motility. Cryopreserved semen samples from 10 Holstein bulls were used in the study. A multiparametric analysis of results obtained by the ISAS 3Fun kit (Proiser) was performed. The different fluorescent groups were detected and their motility characteristics evaluated using ISAS software. Clustering procedures using the kinematic data resul…

Utilización del Integrated Semen Analysis System (ISAS)® para el análisis morfométrico espermático humano y su significado en las técnicas de reproducción asistida

Use of the integrated semen analysis system (ISAS ® ) for morphometric analysis and its role in assisted reproduction technologies

Effects of chitosan films on the growth of Listeria monocytogenes, Staphylococcus aureus and Salmonella spp. in laboratory media and in fish soup.

The objective of this study was to assess the antimicrobial effectiveness of chitosonium acetate films on the growth of Listeria monocytogenes, Salmonella spp. and Staphylococcus aureus. The samples were tested in both laboratory conditions using Tryptone Soy Broth (TSB) and in a real food system using fish soup. The study was carried out at different temperatures (4, 12, and 37 degrees C) in order to discern the influence of such variables. Moreover, a sensory evaluation of the final product was performed as a parameter of consumer acceptance. The results showed a significant reduction of the bacterial growth, which greatly depended on the bacteria type, the temperature of incubation and t…

Ejaculate fractioning effect on llama sperm head morphometry as assessed by the ISAS(®) CASA system.

Contents South American camelid sperm characteristics are poorly known compared with those of other domestic animals. The long-term duration of ejaculation makes difficult to gather all the seminal fluid, implying possible ejaculation portion losses. Thus, the aim of this research was to evaluate the characteristics of the morphology and morphometry of the spermatozoa change during ejaculation. The morphometric characterization was tested on nine specimens of the Lanuda breed, using a special artificial vagina. In five of the animals, a fractioning of the ejaculate was performed by taking samples every 5 min. for a total of 20 min. Air-dried seminal smears were stained with Hemacolor and mo…

Effect of hypoosmotic swelling test and water test on the distribution of sperm subpopulations in bull

Effect of Cetrorelix on sperm morphology during migration through the epididymis in the cynomolgus macaque (Macaca fascicularis)

The importance of the cynomolgus monkey as a model for human reproductive medicine prompted this examination of epididymal sperm morphology. Computer-aided sperm morphological analysis was used for the first time to provide morphometric data on sperm heads as they traversed the epididymal duct of Macaca fascicularis. The duct was divided into six regions, starting close to the testis (proximal) and ending close to the vas deferens (distal). To determine the androgen-dependence of the changes, one group of animals received a GnRH-antagonist (Cetrorelix, Asta Medica, Frankfurt, Germany) to induce testicular regression and lower epididymal androgens, while a control group received only vehicle…

Study of nuclear and acrosomal sperm morphometry in ram using a computer-assisted sperm morphometry analysis fluorescence (CASMA-F) method.

The aim of this study was to develop a new method that allows morphometric assessment of the sperm nucleus and acrosome in the ram using fluorescence microscopy and free software. The study was divided into three experiments. In the first experiment, semen smears from 20 ejaculates were fixed and labeled with a propidium iodide-pisum sativum agglutinin (PI/PSA) combination. Digital images of the sperm nucleus, acrosome, and whole sperm head were captured and analyzed using the ImageJ program. The computer-assisted sperm morphometry analysis fluorescence (CASMA-F) method used allowed the differentiation, capture, and morphometric analysis of most sperm nuclei, acrosomes, and whole heads with…

The Trumorph℗® system: The new univ the morphology of living sperm

Abstract Evaluation of sperm morphology is a fundamental component of semen analysis, but its real significance has been obscured by a plethora of techniques that involve fixation and staining procedures that induce artefacts. Here we describe Trumorph℗ ® , a new method for sperm morphology assesment that is based upon examination of wet preparations of living spermatozoa immobilized by a short 60 °C shock using negative phase contrast microscopy. We have observed samples from five animals of the following species: bull, boar, goat and rabbit. In every case, all the components of the sperm head and tail were perfectly defined, including the acrosome and midpiece (in all its length, includin…

CASA-Mot in mammals: an update

Sperm motility is one of the most widely used parameters of sperm quality. Computer-aided sperm motility analysis (CASA-Mot) systems were developed to reduce the subjectivity of sperm motility assessment, and have had broad scientific and practical acceptance. In this review, the sources of variation and current applications of this technology and its relationships with other sperm quality tests are described in detail. Despite remarkable advances in the technique, there is still great need for standardisation in many species, and the numerous factors that affect the results make it difficult to provide universally accepted criteria for classifying semen samples based on sperm motility char…

Computer assisted sperm morphometry in mammals: a review.

Computer-assisted sperm morphometry analysis (CASMA or ASMA) systems were developed to reduce the subjectivity of sperm morphology assessement. This review focuses on a complete description of the CASMA technique, including recent developments, factors of variation, results in the different species and possible applications. Techniques to study sperm morphometry include light microscopy, phase-contrast microscopy and, more recently, fluorescence microscopy. Most published studies on sperm morphometry have been centered on the whole sperm heads, although some of them also measured other parts of the sperm structure, such as the nucleus, acrosome, midpiece or flagellum. The independent study …

Regionalization of epididymal duct and epithelium in rats and mice by automatic computer-aided morphometric analysis.

Aim: To establish a rat and mouse epididymal map based on the use of the Epiquatre automatic software for histologic image analysis. Methods: Epididymides from five adult rats and five adult mice were fixed in alcoholic Bouin’s fixative and embedded in paraffin. Serial longitudinal sections through the medial aspect of the organ were cut at 10 µm and stained with hematoxylin and eosin. As determined from major connective tissue septa, nine subdivisions of the rat epididymis and seven for the mouse were determined, consisting of five sub-regions in the caput (rat and mouse), one (mouse) or three (rat) in the corpus and one in the cauda (rat and mouse). Using the Epiquatre software, several t…

Perioperative Outcome in Dogs Undergoing Emergency Abdominal Surgery: A Retrospective Study on 82 Cases (2018-2020)

Emergency abdominal surgery carries high morbidity and mortality rates in human medicine; however, there is less evidence characterizing the outcome of these surgeries as a single group in dogs. The aim of the study was to characterize the clinical course, associated complications and outcome of dogs undergoing emergency abdominal surgery. A retrospective study was conducted. Dogs undergoing emergency laparotomy were included in the study. Logistic regression analysis was performed to identify variables correlated with death and complications. Eighty-two dogs were included in the study. The most common reason for surgery was gastrointestinal foreign body. Overall 15-day mortality rate was 2…

A comparison of the effects of bilateral efferent duct ligation and of partial epididymectomy on the testes of rats.

Testis weight as a percentage of body weight did not change following bilateral ligation of the efferent ducts (EDL) close to the epididymis, whereas following removal of part of the epididymis between the site of ligation and a point close to the junction between the caput and corpus (PCE), testis weight first rose linearly until Day 4 and then showed an exponential decrease between Days 4 and 28. After EDL, the perimeter of the seminiferous tubules rose for the first 7 days and then remained elevated, whereas after PCE, there was a linear decrease between Days 4 and 28. Following EDL, the percentage of altered and degenerated tubular cross-sections rose to about 30% and 10%, respectively,…

Effect of chamber characteristics, loading and analysis time on motility and kinetic variables analysed with the CASA-mot system in goat sperm.

Several factors unrelated to the semen samples could be influencing in the sperm motility analysis. The aim of the present research was to study the effect of four chambers with different characteristics, namely; slide-coverslip, Spermtrack, ISAS D4C10, and ISAS D4C20 on the sperm motility. The filling procedure (drop or capillarity) and analysis time (0, 120 and 240s), depth of chamber (10 or 20μm) and field on motility variables were analysed by use of the CASA-mot system in goat sperm. Use of the drop-filling chambers resulted in greater values than capillarity-filling chambers for all sperm motility and kinetic variables, except for LIN (64.5% compared with 56.3% of motility for drop- a…

Male facial attractiveness and masculinity may provide sex- and culture-independent cues to semen quality

Phenotype-linked fertility hypothesis (PLFH) predicts that male secondary sexual traits reveal honest information about male fertilization ability. However, PLFH has rarely been studied in humans. The aim of the present study was to test PLFH in humans and to investigate whether potential ability to select fertile partners is independent of sex or cultural background. We found that on the contrary to the hypothesis, facial masculinity was negatively associated with semen quality. As increased levels of testosterone have been demonstrated to impair sperm production, this finding may indicate a trade-off between investments in secondary sexual signalling (i.e. facial masculinity) and fertilit…

Morphological characterization of ejaculated cynomolgus monkey (Macaca fascicularis) sperm

The aim of this study was to give reference values for the frequency of morphological sperm abnormalities present in the semen from non-experimental cynomolgus monkeys as well as for the dimensions of sperm heads. Spermatozoa from the liquid portion of electroejaculates from 14 cynomolgus monkeys were air-dried as smears, fixed, and stained with Harris's Haematoxylin and subjected to visual analysis of morphology and computer-aided analysis of ten morphometric variables. The majority (83%) of sperm were morphologically normal. Tail defects were the most common (11%), and showed the highest variation between individuals, the values ranging between 4 and 23%. Head abnormalities consisted of l…

Comparison of the brainstem auditory evoked responses during sevoflurane or alfaxalone anaesthesia in adult cats.

Abstract Objective To compare the effects of general anaesthesia using sevoflurane or alfaxalone on the brainstem auditory evoked response (BAER) test in adult healthy cats. Study design Prospective, clinical, ‘blinded’, crossover study. Animals Ten feral adult healthy cats. Methods Premedication consisted of dexmedetomidine (0.01 mg kg –1 ) intramuscularly (IM). The first general anaesthesia was induced and maintained with sevoflurane (treatment S) for physical examination, BAER test, complete blood tests, thoracic radiographs and abdominal ultrasound. The second general anaesthesia was induced with alfaxalone (treatment A) IM (2 mg kg –1 ) and maintained with alfaxalone (10 mg kg –1 hour …

Effect of genistein added to bull semen after thawing on pronuclear and sperm quality.

The aim of this research was to study the effect of different genistein treatments on bull sperm after thawing on pronuclear formation after in vitro fertilization (IVF) and on different sperm quality variables. Three experiments were performed. In Experiment 1, three treatments (Control, sperm incubation for 1 h at 37 degrees C with or without genistein) and two sperm concentrations during IVF (I or 3 x 10(6) sperm/mL) were evaluated to study the influence of genistein on pronuclear formation (PNF). Sperm incubation for 1 h before IVF reduced PNF regardless of sperm concentration. However, after sperm incubation and with 3 x 10(6) sperm/mL in IVF, the genistein treatment group had greater …