Expanding the Limits of Computer-Assisted Sperm Analysis through the Development of Open Software

Abstract

Computer assisted sperm analysis (CASA) systems can reduce errors occurring in manual analysis. However, commercial CASA systems are frequently not applicable at the forefront of challenging research endeavors. The development of open source software may offer important solutions for researchers working in related areas. Here, we present an example of this, with the development of three new modules for the OpenCASA software (hosted at Github). The first is the Chemotactic Sperm Accumulation Module, a powerful tool for studying sperm chemotactic behavior, analyzing the sperm accumulation in the direct vicinity of the stimuli. This module was validated by comparing fish sperm accumulation, with or without the influence of an attractant. The analysis clearly indicated cell accumulation in the treatment group, while the distribution of sperm was random in the control group. The second is the Sperm Functionality Module, based on the ability to recognize five sperm subpopulations according to their fluorescence patterns associated with the plasma membrane and acrosomal status. The last module is the Sperm Concentration Module, which expands the utilities of OpenCASA. These last two modules were validated, using bull sperm, by comparing them with visual counting by an observer. A high level of correlation was achieved in almost all the data, and a good agreement between both methods was obtained. With these newly developed modules, OpenCASA is consolidated as a powerful free and open-source tool that allows different aspects of sperm quality to be evaluated, with many potential applications for researchers.

Ability of the ISAS3Fun Method to Detect Sperm Acrosome Integrity and Its Potential to Discriminate between High and Low Field Fertility Bulls

“…In a recent work, we described a simple method that allows the simultaneous visualisation of sperm plasma and acrosomal membrane integrity in bulls by combining the fluorochromes Hoechst 33342, CFDA, and PI (ISAS3Fun) [ 8 ]. To facilitate the analysis using this test, a new module was added to the OpenCASA software, which allows for the automated analysis of these sperm quality parameters [ 12 ].…”

“…Sperm plasma membrane integrity (viability) was determined using an acridine orange and propidium iodide combination [ 11 ]. The number of total spermatozoa and the percentages of membrane-intact spermatozoa were determined using the OpenCASA software [ 12 ]. At least 300 sperm cells were analysed per sample.…”

“…The labelling mix included propidium iodide (PI), Hoechst 33342, and carboxyfluorescein diacetate (CFDA). Two hundred spermatozoa were examined on each slide, and the staining patterns were classified into four categories ( Figure 1 ), aided with v2 of the OpenCASA software [ 12 ]: (a) those with intact acrosomes and plasma membranes (IAIM); (b) those with intact acrosomes and damaged plasma membranes (IADM); (c) those with damaged acrosomes and intact plasma membranes (DAIM); and (d) those with damaged acrosomes and plasma membranes (DADM) [ 7 ].…”

Abnormality Determination of Spermatozoa Motility Using Gaussian Mixture Model and Matching-based Algorithm

“…With CASA [18], microscopic photographs of semen are taken, as well as analyzing the number, movement, and morphology of sperm in the semen [19]. Despite its capabilities, commercial CASA products are too expensive and rudimentary [20].…”

The Importance of Studying Factors That Affect the In Vitro Evaluation of Semen Quality to Predict Potential Fertility in Males

“…Of similar importance is a novel module that can automatically measure one technique for the evaluation of different sperm functional parameters. As a result of the introduction of these three new modules, Open CASA (hosted at Github) is now the most powerful CASA system [1].…”