Optional: Please comment on any other aspects of the study that are not specifically covered by the previous questions

1. NO in-depth discussion of analytical methods (e.g., pyro-GC/MS) and limitation, QA/QC, spectrum, etc., 

2. pyro-GC/MS measures the concentration but it’s a destructive method. Therefore, before going for the pyro-GC/MS, it is imperative that obtaining data about the physical characteristics such as the shape and size are equally important. This study doesn't fully represent the MNPs contamination in the testicles from human and dogs. 

3. providing a more detailed roadmap or specific hypotheses for future studies would be helpful for researchers building on this work. 

As the investigators point out, it is beneficial to conduct detailed studies of dose-response with different micro- and nano-plastics. In these studies, special attention should be given to plastics like polycarbonate (PC), polyurethane (PU) and the like (PET) which have the potential to release estrogenic chemicals. 

For convenience, I would like to summarize some aspects that I identified
1 - Insufficient validation of the procedures for sampling, sample processing, and sample analysis. This is a key issue, especially since absolute concentrations of plastics are reported.
2 - Lack of statistical rigor: insufficient number of samples in view of the various co-variances and in view of the various additional factors that have not been considered.
3 - Lack of consideration of number of particles whilst not considering particle properties in the samples: size, morphology, charge, etc.
4 - Co-variance of different types of plastics in the biological samples and their relation with total concentrations of plastics. It is for instance not allowed to depict concentrations of individual plastics with endpoints of assessment, as done in Figure 4, whilst knowing that in each samples other plastics are present in differing concentrations, shpaes, sizes, etc.