Stesura Seveso 519Archivio Italiano di Urologia e Andrologia 2022; 94, 4 LETTER TO EDITOR No conflict of interest declared. Submitted 18 July 2022; Accepted 24 July 2022 To the Editor, Currently, semen analysis is the unique test to evaluate men’s fertility potential. Semen analysis provides valuable infor- mation on sperm production and quality. In addition to the conventional assessment of the sperm characteristics in a basic semen analysis routine, performing a differential diagnosis of leukocytes and sperm precursors immature germ cells (IGC) is also pivotal (1-3), due to adding valuable and clinically suitable information to the semen report. While increased leukocyte count may indicate infections (4), increased exfoliation of IGC from seminiferous tubules suggests abnormal spermatogenesis (5, 6). Even the inflammation could induce exfoliation of IGC from seminiferous tubules (7). In semen analysis, a global and differential count of all seminal round cells can improve the diagnosis (8) and treatment options to achieve a successful pregnancy (9). A high count of IGC in semen would represent different seminal and reproductive alterations and would be an essential indicator for detecting testicular alterations (6). Therefore, an index could be developed comparing IGC count/mL vs. sperm count/mL. In some instances, this index would be paramount for estimating, for example, the negative impact of varicocele on the germinal epithelium in increased exfoliation of IGC in semen. The normal is to find one IGC for every 40 spermatozoa, values between 20/1 and 40/1 are indeterminate, and values lower than 20/1 indicate increased exfoliation of IGC, which suggests a loss of integrity of the germinal epitheli- um. Occasionally, ratios greater than 1/1000 are also found, mainly in ejaculate with high sperm count (Andrade-Rocha FT, Unpublished data). The lower this index, the worse is the integrity of the germinal epithelium. The cells are sloughed off before completing the spermatogenesis process and spermatozoa production. For example, the conven- tional semen parameters routinely evaluated in a 32 years old man in Lisa Andrology Lab (Petrópolis, RJ, Brazil) for investigating male infertility showed the ratio was six IGC for each sperm; it was even worse, showing an inversion in the index (Table 1 and Figure 1) show a representative aggregate of IGC in the semen specimen. Increased exfoliation of immature germ cells detected in semen analysis routine and its clinical significance Fernando Tadeu Andrade-Rocha 1, Walter D. Cardona Maya 2 1 Lisa Andrology Lab, Petrópolis, RJ, Brazil; 2 Reproduction Group, Department of Microbiology and Parasitology, Faculty of Medicine, Universidad de Antioquia UdeA, Medellín, Colombia. DOI: 10.4081/aiua.2022.4.519 Figure 1. Unequal aggregation of immature germ cells regarding to sperm. The semen smear used for the microphotograph shown in this editorial was made with concentration to show the proportionality between IGC and sperm in the analyzed semen specimen. Table 1. Patient semen characteristics and lower reference limit. Parameter Outcomes Lower reference limit – percentile 5th (11) Semen volume 6.1 mL 1.4 mL Sperm count/mL 317 000 16 x 106/mL Total sperm count 1 931 710 39 x 106 per ejaculate Vitality 20% 54% Total motility 10% 42% Progressive motility (a) 0 30% Slow/irregular motility (b) 9% - Non-progressive motility (c) 1% 1% Immotile (d) 90% 20% Normal morphology 0.53 4% Amorphous sperm 11% - Tapered sperm 7% - Sperm immature germ cells vs. sperm 1/6 1/40 Hypo-osmotic swelling 18% 58% pH 8.2 > 7.2 Archivio Italiano di Urologia e Andrologia 2022; 94, 4 F.T. Andrade-Rocha, W.D. Cardona Maya 520 This assessment measures the intensity of the famous seminal stress pattern that John MacLeod proposed in the 1960s (10). According to MacLeod, the stress pattern is characterized by an increase of amorphous and tapered and exfoliated IGC in semen and is usually diagnosed in some varicocele men. Unlike previous observations by MacLeod (10), it has been observed that varicocele men can cause structural changes in sperm, like amorphous and tapered sperms, increased exfoliation of sperm precursors and both. Therefore, further studies are needed to expand knowledge on this issue, which is practically unexplored in clinical and laboratory practice. REFERENCES 1. Jassim A, Festenstein H. Immunological and morphological characterisation of nucleated cells other than sperm in semen of oligospermic donors. J Reprod Immunol. 1987; 11:77-89. 2. Tomlinson MJ, Robert Barratt CL, Bolton AE, et al. Round cells and sperm fertilizing capacity: the presence of immature germ cells but not seminal leukocytes are associated with reduced success of in vitro fertilization. Fertil Steril. 1992; 58:1257-9. 3. Fedder J, Askjaer SA, Hjort T. Nonspermatozoal cells in semen: relationship to other semen parameters and fertility status of the couple. Arch Androl. 1993; 31:95-103. 4. Sharma R, Gupta S, Agarwal A, et al. Relevance of leukocytospermia and semen culture and its true place in diagnosing and treating male infertility. World J Mens Health. 2022; 40:191-207. 5. Andrade-Rocha FT. Semen analysis in laboratory practice: an overview of routine tests. J Clin Lab Anal. 2003; 17:247-58. 6. Palermo GD, Neri QV, Cozzubbo T, et al. Shedding light on the nature of seminal round cells. PLoS One. 2016; 11:e0151640. 7. Gandini L, Lenzi A, Lombardo F, et al. Immature germ cell separation using a modified discontinuous Percoll gradient technique in human semen. Hum Reprod. 1999; 14:1022-7. 8. Patil PS, Humbarwadi RS, Patil AD, Gune AR. Immature germ cells in semen - correlation with total sperm count and sperm motility. J Cytol. 2013; 30:185-9. 9. Ariagno J, Curi S, Mendeluk G, et al. Shedding of immature germ cells. Arch Androl. 2002; 48:127-31. 10. MacLeod J. Seminal cytology in the presence of varicocele. Fertil Steril. 1965; 16:735-57. 11. World Health Organization. WHO Laboratory Manual for the Examination and Processing of Human Semen. 6th ed. WHO Press; Geneva, Switzerland. 2021 (accessed on 15 July 2022). Correspondence Fernando Tadeu Andrade-Rocha, BS (Corresponding Author) lisalabrescenter@gmail.com Lisa Andrology Lab, Petrópolis, RJ (Brazil) Walter Darío Cardona Maya, PhD wdario.cardona@udea.edu.co Grupo Reproducción, Facultad de Medicina, Universidad de Antioquia, Medellin (Colombia)