In the field of assisted reproductive techniques (ART), computer-assisted sperm analysis (CASA) systems have proved their utility and potential for assessing sperm quality, improving the prediction of the fertility potential of a seminal dose. Although most laboratories and scientific centers use commercial systems, in the recent years certain free and open-source alternatives have emerged that can reduce the costs that research groups have to face. However, these open-source alternatives cannot analyze sperm kinetic responses to different stimuli, such as chemotaxis, thermotaxis or rheotaxis. In addition, the programs released to date have not usually been designed to encourage the scalability and the continuity of software development. We have developed an open-source CASA software, called OpenCASA, which allows users to study three classical sperm quality parameters: motility, morphometry and membrane integrity (viability) and offers the possibility of analyzing the guided movement response of spermatozoa to different stimuli (useful for chemotaxis, thermotaxis or rheotaxis studies) or different motile cells such as bacteria, using a single software. This software has been released in a Version Control System at Github. This platform will allow researchers not only to download the software but also to be involved in and contribute to further developments. Additionally, a Google group has been created to allow the research community to interact and discuss OpenCASA. For validation of the OpenCASA software, we analysed different simulated sperm populations (for chemotaxis module) and evaluated 36 ejaculates obtained from 12 fertile rams using other sperm analysis systems (for motility, membrane integrity and morphology modules). The results were compared with those obtained by Open-CASA using the Pearson’s correlation and Bland-Altman tests, obtaining a high level of correlation in all parameters and a good agreement between the different used methods and the OpenCASA. With this work, we propose an open-source project oriented to the development of a new software application for sperm quality analysis. This proposed software will use a minimally centralized infrastructure to allow the continued development of its modules by the research community. In the field of the biology of reproduction, it is very important to develop new methods that allow us to predict the breeding capacity of males of certain species. In recent years, the use of Computer Assisted Sperm Analysis systems (that allow the user to analyze certain parameters of sperm that have been shown to be related to fertility), has been fundamental in advancing this knowledge. In addition, new parameters are being investigated that are believed to help characterize seminal quality. However, most of the software used to analyze these parameters is commercial and private, and the user is not allowed to add new features. Although there were already open source alternatives to analyze certain parameters of seminal quality, in this work we have integrated several tools into one, with an easy-to-use interface for the user, which we hope will help the scientific community to develop new functionalities and methods. The tool is free and open-source.

Some breeds of sheep are highly seasonal in terms of reproductive capability, and these changes are regulated by photoperiod and melatonin secretion. These changes affect the reproductive performance of rams, impairing semen quality and modifying hormonal profiles. Also, the antioxidant defence systems seem to be modulated by melatonin secretion, and shows seasonal variations. The aim of this study was to investigate the presence of melatonin and testosterone in ram seminal plasma and their variations between the breeding and non-breeding seasons. In addition, we analyzed the possible correlations between these hormones and the antioxidant enzyme defence system activity.

The reproductive seasonality of sheep suggests that melatonin receptors may be present in ram spermatozoa. The present study confirms the presence of melatonin MT1 and MT2 receptors. The MT1 receptor was detected using immunocytochemistry, with four sperm subpopulations identified based on the following labelling patterns: (1) one small subpopulation with labelling over the entire head and tail; (2) one of two main subpopulations that exhibited reactivity at the equatorial, post-acrosomal, neck and tail regions; (3) another main subpopulation with equatorial and tail labelling only; and (4) a subpopulation in which staining was detected only in the tail. Immunocytochemistry revealed the presence of the melatonin MT2 receptor, with intense staining on the acrosome, post-acrosomal region and neck and tail regions of all cells, but not in the equatorial region. Western blot identification of ram protein extracts revealed a 39-kDa band compatible with both MT1 and MT2 receptors, a 75-kDa band compatible with MT1/MT2 heterodimerisation, a 32-kDa band compatible with MT1 receptor activation and a double band of 45–55 kDa that is compatible with MT2 receptor homodimerisation or heterodimerisation with other G-proteins. In conclusion, we provide evidence of the presence of MT1 and MT2 receptors in ram spermatozoa, although the biochemical pathway triggered by these receptors and their function in terms of fertility remains to be elucidated.