NK cells not only in endometrium but also in ovulatory cervical mucus in patients with decreased fertility.

OBJECTIVE: An increased number of NK cells is associated with autoimmune disorder and is known to play a role in infertility. The aim of our research was to monitor the density of NK cells CD56+ and CD16+ in ovulatory cervical mucus (OCM) and in endometrium in infertile women as well as in connection with the actual status of antibodies against phospholipids, sperm and HHV-6 antibodies. TYPE OF STUDY: Original aticle. SETTING: Genetika - Plzeň. METHODS: Seventy-two randomly selected women aged 20-39 (mean age: 32.3) years old resulted in fifty-seven patients with repeated unexplained miscarriages, and fifteen fertile healthy women. The hormonal status was studied including ovulation, the humoral autoimmune responses to eight phospholipids, trombophilia, karyotyping, hysteroscopy, and endometrium immunohistology. Patients were without any clinical and laboratory symptoms of vaginitis at the time of OCM sampling and endometrium study. In one patient antiphospholipid syndrome was present, and in one woman diabetes mellitus was identified. Uterine NK cells CD56+ , CD16+ and NK cells in OCM were identified by immunocytochemistry, antiphospholipid antiboides by ELISA. We used indirect MAR-test for study of local spermagglutinating antibodies in OCM. Indirect immunofluorescent method was used for detection of serum and OCM IgM, IgG antibodies against HHV-6 levels at the time of ovulation. RESULTS: We found both high density of NK cells CD56+ and CD16+ in OCM and in endometrium in only two infertile women with repeated abortions. NK cells in OCM were missing in other samples of patients. The prevalence of high density of NK cells CD56+ in the endometrium was seen in twenty three (40%), NK cells CD16+ in eleven (19%), NK cells 56+ and NK cells 16+ together in eight (14%). Levels of serum and OCM IgG against HHV-6 in all examined patients were not elevated, no cervical sperm antibodies were found. CONCLUSION: We compared density of NK cells CD56+ and CD16+ in OCM and secretory endometrium in all infertile patients. Our results show that cell mucosal activity in the cervical area at the time of ovulation in two infertile patients was evident. We excluded the abnormal number of NK cells owing to local and general viral infection (HHV-6). But our question still remains - are cervical NK cells fixed or still migrating from endometrium into OCM? New research is planned.

[Results of the treatment in selected infertile patients with high density of endometrial NK cells CD56+ and CD16+ Second part]. / Výsledky lécby u vybraných infertilních pacientek s vysokou denzitou endometriálních NK bunek CD56+ a CD16+ Druhá cást.

INTRODUCTION: This statements follows the first part of our publication entitled Screening of endometrial NK cells in selected infertile patients (First part - Methods and current results), where we dealt with methodology and endometrial findings of the natural killer cells CD16+ and CD56+. These cells are among the most important in preimplantaion, and in implantation period, in early pregnancy too, but can also negatively influence above mentioned processes. OBJECTIVE: We focused on the immunomodulatory treatment with intramuscular immunoglobulins in 21 infertile patients with a high density of endometrial NK cells CD16+ and CD56+. DESIGN: Original work-prospective study. SETTING: Genetics-Pilsen. METHODS: From the originally investigated 57 women aged 25-41 (average 34) years with their history of 3-9 abortions, we target on a high selective subgroup of 21 infertile patients with pathological density of endometrial NK cells CD56+ and CD16+. RESULTS: Twelve patients underwent immunomodulatory treatment with intramuscular immunoglobulin in time of positive HCG and went on the therapeutic scheme, nine infertile women continued with IVF-ET but did not achieve HCG positivity despite of initial treatment with immunoglobulins. Of the 12 treated patients, seven of them gave a birth to healthy babies, as to the next five pregnancies - one was extrauterine pregnancy, one genetically defective fetus, three pregnancies go on successfully.Nine women from the IVF program did not get pregnat. CONCLUSION: Immunomodulatory treatment with immunoglobulins also influences endometrial NK cells CD56+ and CD16+ associating with an overproduction of embryocytoxic cytokines. Treatment of our patients with the pathological endometrial density of NK cells must be solved strictly individually.

[Screening of endometrial NK cells in selected infertile patients First part - Methods and current results]. / Screeningové vysetrení endometriálních NK bunek u vybraných infertilních pacientek 1. cást - metodika a prubezné výsledky.

INTRODUCTION: Generaly, natural killer cells (NK cells) are among the most important cells of our immune defense system. They are present in the blood, decidua but also in secretory endometrium. OBJECTIVE: We investigate an association between high density of NK cells CD56+ and NK cells CD16+ into blood and secretory endometrium in patients with up to now unexplained recurrent miscarriage. At the same time, we focused on diagnosis of antiphospholipid syndrome, congenital trombophilic factors, stress factor, and lenght of hormonal contraception before conception. DESIGN: Original work-prospective study. SETTING: Genetics-Pilsen. METHODS: We investigated 59 patients aged 25-41 (average 34) years with their history of 3-9 abortions. We monitored their blood cells by flow cytometry and endometrium obtained by hysteroscopy performed on the 22nd-24th day of the menstrual cycle. We concentrated on the cellular immunity focused on the identification of lymphocytes CD56+ and CD16+. ELISA method was used for identification of antiphospholipid antibodies. RESULTS: Our study goes on, we are publishing our preliminary results. We found a high density of endometrial NK cells CD56+ in 41/57, NK cells CD16+ in 40/57, both NK cells CD56+ and CD16+ in 36/57 patients. In 13 women, we identified only sporadically these cells in the secretory endometrium. NK cells CD56+ and NK cells CD16+ in the blood were all normal number in all our patients. Three of them have a homozygot form of Leiden mutation, and 18/57 primary antiphospholipid syndrome, 12 women have university education, four of them, medical doctors, have regular night services in the hospital. CONCLUSION: Overproduction of endometrial NK cells is associated with increased local activity of embryocytotoxic cytokines that may negatively affect pregnancy. Treatment of patients with pathological immunological findings must be solved individually, as we will show soon in part two of our study.

Proteomic identification of sperm antigens using serum samples from individuals with and without antisperm antibodies.

The aim of the study was to identify human sperm antigens reacting with polyclonal antisperm antibodies. Protein sperm extracts were subjected to electrofocusing, and next immune reactions (immunoblotting) were carried out with positive for antisperm antibodies and control (not containing antisperm antibodies) serum samples. Proteomic analysis of human sperm proteins resulted in identification of 80 sperm antigens that could be divided into three groups: antigens specific for patients with antisperm antibodies (32), antigens recognised by both infertile patients and control sera (35) and antigens detected by control serum samples only (13). Among antigens specific for infertile patients, there were 12 sperm entities known to be involved in fertilisation process. We have also characterised three protein entities identified only by sera of infertile women. Altogether, the proteomic analysis resulted in identification of 27 sperm entities not reported previously in human sperm proteome. Identified proteins are sperm antigens that could be potentially responsible for immunological infertility. The study also sheds new light on the sperm antigens in aspect of gender specificity. The investigation of human sperm proteome by the use of antisperm antibodies-containing sera of infertile individuals not only may indicate new proteins but also can draft their immunological nature.

Extremely hypotrophic newborn of mother with systemic lupus erythematosus and antiphospholipid syndrome.

The case presented describes a high-risk pregnancy of a woman with systemic lupus erythematosus (SLE) with multiple lesions of central nervous system (CNS), vasculitis, secondary epilepsy and antiphospholipid syndrome (APS). At gestational age 28 weeks and 3 days the pregnancy was urgently terminated via caesarean section and an extremely hypotrophic immature newborn with a birth weight of 580 g was born. The high disease activity in the mother at the time of conception and the histologically proven chronic placental insufficiency due to APS are presumably the causes for the extensive hypotrophy of the neonate. The significant comorbidity of the newborn, including respiratory distress syndrome, bronchopulmonary dysplasia, necrotizing enterocolitis, osteopathy of prematurity, transient hypothyroidism and hypocortisolism, vesicoureteral reflux, and hypertonic-hyperexcitation syndrome complicated his three-month stay in NICU. A positive titre of transplacentally transferred anticardiolipin and anti-ß2 glycoprotein antibody was detected in the child and persisted through the following 30 months. During the three-year follow-up, significantly delayed neuropsychological development with microcephaly (-4 SD) and short stature of the child was observed. Finally, the authors discuss possible causes of neuropsychological consequences in children of mothers with SLE and APS and emphasize the need for long-term monitoring and specialized care to improve development of these children.

Immunodominant semen proteins II: contribution of seminal proteins to female immune infertility.

Seminal fluid is a protective medium for sperm, but it also represents potential immunogenic structures for the female immune system. Anti-seminal antibodies may threaten early fertilization. The aim of our work is to detect and identify seminal proteins that are related to female isoimmunization. In this report, we quantified serum anti-seminal IgG antibodies. Seminal proteins were analysed by two-dimensional gel electrophoresis followed by immunoblotting. To identify IgG-binding proteins of interest, a proteomic approach was selected. The dominant seminal antigens were detected within the relative molecular mass ranging from 25 to 85 kDa and the isoelectric point from 5 to 7. The detected proteins were further identified as prostate-specific antigen, prostatic acid phosphatase, zinc-α-2-glycoprotein and zinc finger protein 778. Since these proteins were recognized by IgGs produced by infertile women and not by fertile women, we presume that major seminal antigens may play an important role in the pathogenesis of female immune infertility. Our study suggests the pattern of seminal proteins for further therapeutic attempts in the diagnosis of female immune infertility.

[The incidence of latent asymptomatic celiac disease in women with decreased fertility]. / Výskyt latentní asymptomatické celiakie u zen se snízenou plodností

OBJECTIVE: To determine whether patients with recurrent spontaneous abortions and IVF suffer from latent celiac disease. DESIGN: Perspective study. SETTINGS: Department of Gynecology and Obstetrics and Faculty Hospital, Charles University, Pilsen. MATERIAL AND METHODS: Sera from 442 infertile patients aged 23-47 years and 86 healthy fertile controls aged 22-38 years were examined by commercial ELISA sets for the presence (absence) of antibodies against gliadin IgG, IgA and tissue transglutaminase in IgG and IgA. RESULTS: We found a statistically significant difference between the control group and patients with decreased fertility. They have a positive values in anti-tissue transglutaminase IgA (p = 0.0096). Frequent, very low positive reaction of antigliadin IgG was also found in the control group of women. CONCLUSION: Patients with unexplained repeated miscarriages and repeated unsuccessful IVF with IgA positive tissue transglutaminase have new complement of other tests for autoimmunity and examination of a total IgA, and gastroenterological examination. Eleven patients with significant positive anti-tissue transglutaminase IgA were successfully conceived only under strict gluten-free diet.

Repeated miscarriages in patients with antiphospholipid syndrome and subjected to in vitro fertilization: the importance of preimplantation genetic diagnosis.

The aim of this study was to investigate two patients with antiphospholipid syndrome (APS) who suffered from multiple repeated pregnancy losses of probably genetically impaired embryos. Sera from these patients contained high levels of IgG antibodies against cardiolipin, IgG and IgM phosphatidyl inositol, IgG phosphatidyl L-serine, and IgG against anti-annexin V. The conventional treatment of APS was ineffective. Preimplantation genetic diagnosis (PGD) showed chromosomally impaired embryos. The course of pregnancies and deliveries were monitored due to the close collaboration of in vitro fertilization and PGD in early embryos. After the selection of normal embryos using PGD, and the treatment of APS, both patients became pregnant and delivered healthy babies. Without such selection, both women would probably have miscarried their embryos again.

Occurrence of selected antiphospholipid antibodies in offspring born from mothers with autoimmunity.

Autoimmune diseases can be accompanied by presence of various antiphospholipid antibodies (aPL). The laboratory criteria of antiphospholipid syndrome are based on detection of anticardiolipin, lupus anticoagulant or to antiß2- glycoprotein I but currently a significance of other multiple aPL is being discussed. Because of their vascular and neuroinflammatory effect aPL, if being transplacentally transferred, might inflict damage in developing organism. The aim of our study was to determine the occurrence of eight selected aPL in offspring of mothers with proven autoimmune disease with aPL positivity. The possible influence of aPL presence on clinical, ultrasound and laboratory outcome of children was observed as well. The prospective study included 38 women: 17 women with primary antiphospholipid syndrome and 21 women with other diagnosed autoimmune disease with detected aPL. Also included were 39 children born to the above mentioned mothers between January 2009 and April 2010 in Perinatology Centre in Pilsen, Czech Republic. The control group consisted of 30 mothers without AD and their 30 healthy singletons. Preliminary results of the study showed the presence of aPL in 42.1% neonates of aPL positive mothers with autoimmune disease, six month later aPL were present in only 37.5 % of these children. Observed occurrence of aPL positivity at 6 months of age in originally negative offspring could be attributed to vaccination or food exposure. Psychomotor development of children has proceeded without major deviations. The follow-up study continues and will evaluate both groups of children at two years of age.

Avidity of anti-ß2-glycoprotein I antibodies in patients with antiphospholipid syndrome.

Antibodies against ß(2)-glycoprotein I (anti-ß(2)GPI) are one of the hallmarks of the antiphospholipid syndrome (APS). However, they are heterogenic regarding their epitope specificity, pathogenic mechanisms and their avidity. In the current study we present some outstanding issues about avidity of anti-ß(2)GPI antibodies. Our results confirmed that high avidity anti-ß(2)GPI are associated with thrombosis and APS, while in low avidity anti-ß(2)GPI group non-APS (predominantly systemic lupus erythematosus) patients prevailed.

Female serum immunoglobulins G, A, E and their immunological reactions to seminal fluid antigens.

One in five couples of reproductive age has been diagnosed with infertility. Some diagnoses indicate an immunological basis for this disorder. Female immune infertility may be caused by iso-immunization by seminal components. We focused on the characterization of seminal proteins to illustrate the IgG, IgA and IgE immune responses of 31 infertile women. The biochemical characterization was performed by one-dimensional sodium dodecyl sulphate polyacrylamide gel electrophoresis and isoelectric focusing, both of which were followed by immunoblotting analyses. IgG mainly recognized the antigens with relative molecular masses (Mr) 95 and 183 kDa and isoelectric points ranging from 6.9 to 7.0. The immunodominant antigens recognized by IgA had the Mr of 35 kDa and isoelectric points ranging from 6.2 to 7.2. The reactivity of IgE was not confirmed within our group of patients. The seminal IgG- and IgA -binding patterns were analysed immunochemically to determine the characteristics of possible seminal proteins associated with female immune infertility.

[Comparison of selective oxidative stress parameters in the follicular fluid of infertile women and healthy fertile oocyte donors]. / Srovnání vybraných parametru oxidacního stresu ve folikulární tekutine u zen s poruchou plodnosti a u zdravých fertilních dárkyn oocytu

OBJECTIVE: Follicular fluid (FF) affects oocyte development and disruption of its homeostasis has a crucial effect on egg developmental potential. The aim of this study was to compare the levels of selected oxidative stress markers in the FF of women with impaired fertility and healthy fertile oocytes donors. DESIGN: A retrospective comparative study. SETTING: Faculty of Medicine in Pilsen, Charles University in Prague; Institute of Reproductive Medicine and Endocrinology, IVF Center Prof. Zech, Pilsen. METHODS: Levels of homocysteine (Hcy), malondialdehyde (MDA), glutathione peroxidase (GPx), total antioxidant capacity (AOK) and total protein (CB) were analyzed in the FF. We have analysed FF of 146 women - 74 infertile patients (mean age 31 years, SD = 4.65) and 72 healthy fertile oocyte donors (mean age 26 years, SD = 4.44). Only blood free samples were studied after pooling of all FF samples each patient. RESULTS: The study showed a statistically significantly higher Hcy levels (p < 0.0001) in the FF of healthy fertile women compared with impaired fertility group both - comparing the two groups regardless the age and in groups of the same age range (for the age group between 20 to 29 years isp = 0.0002, for the age group between 30 to 39 years is p < 0.0001). When divided into above age ranges we found statistically significantly higher levels of MDA in the control group aged 20 to 29 years compared to same age infertile patients (p = 0.0374) and statistically significantly higher AOK in infertile women between 30 to 39 years of age compared to same age control group (p = 0.0458). CONCLUSION: The presence or on the contrary the absence of prooxidant parameters in the FF has an important role in the ability of conception and subsequent embryo development.

The avidity of anti-beta2-glycoprotein I antibodies in patients with or without antiphospholipid syndrome: a collaborative study in the frame of the European forum on antiphospholipid antibodies.

OBJECTIVE: The objective of this study was to extend the findings of the preliminary study by measuring the avidity of IgG anti-ß2-glycoprotein I antibodies (anti-ß2-GPI) on a larger group of patients with primary or secondary antiphospholipid syndrome (APS) and anti-ß2-GPI positive patients without APS in the frame of the European Forum on antiphospholipid antibodies (aPL). METHODS: Serum from 137 patients with primary APS, APS associated with autoimmune diseases, and patients with autoimmune diseases other than APS from five EU rheumatology centres were tested for anti-ß2-GPI antibodies. The 109 patients who were sera positive for anti-ß2-GPI by the in-house anti-ß2-GPI enzyme-linked immunosorbent assay (ELISA) at the Immunology Laboratory, UMC Ljubljana were selected for further testing on avidity with chaotropic anti-ß2-GPI ELISA. RESULTS: High, low and heterogeneous avidity IgG anti-ß2-GPI was found in 32/109, 17/109 and 60/109 patients respectively. Significantly more patients with APS were in the high avidity than in the low avidity anti-ß2-GPI group, while the opposite was observed for non-APS (both p < 0.001). The most common clinical feature among patients with high avidity anti-ß2-GPI was thrombosis, mainly due to venous thrombosis (p < 0.01 and p < 0.001, versus low avidity anti-ß2-GPI group). CONCLUSION: Patients with or without APS had anti-ß2-GPI of high, low or heterogeneous avidity. High avidity anti-ß2-GPI was associated with thrombosis and APS, while in the low avidity anti-ß2-GPI group non-APS (predominantly SLE) patients prevailed. Determination of anti-ß2-GPI avidity should be considered in the analytical strategies for further differentiation of patients with anti-ß2-GPI antibodies.

[Decreased fertility and today's possibility of examination in reproductive immunology]. / Snízená plodnost a soucasné moznosti vysetrování v reprodukcní imunologii.

UNLABELLED: The aim of reproductive immunology is to look for a futher cause of decreased fertility, immunological reason, which depends on neuro-endocrine relationships. The article explains today's possibility of examination of cell and humoral immunity with regard to human fertility. SETTING: Department of Gynecology & Obsterics Charels University and Faculty Hospital, Pilsen.

[Local sperm antibodies and general levels of immunoglobulins in ovulatory cervical mucus]. / Lokální protilátky proti spermiím a celkové hladiny imunoglobulinu v ovulacním cervikálním hlenu.

OBJECTIVE: The aim of our preliminary study was to compare the levels of total local sIgA and IgG with activity of detected sperm antibodies in ovulatory cervical mucus (OCM). SETTING: Department of Gyneacology and Obstetrics, Medical School and Faculty Hospital, Charles University, Plzen. METHODS: We screened samples of OCM from 12 pacients aged 26-31 (29,6 years on average) by radial immunodifusion (RID) to determine s IgA and IgG. Indirect MAR test was used for detection of spermagglutinationg antibodies. RESULTS: We found out by RID the average concentration of sIgA in OCM 567,84 mg/l (0 -1250,47) and the average concentration of IgG in OCM 23,57 mg/l (8,74-47,99). Antibody activity against sperm cells dominates in IgA with 6 pacients, in IgA with 1 patient, in IgA and IgG together with 1 infertile woman and in IgA and IgM isotypes together with 1 patient. No local sperm antibodies were determined with 3 patients. CONCLUSION: We proved the hypothesis, that the levels of spermagglutinating antibodies do not correlate with findings of total sIgA and IgG in OCM with our patients.

Leukaemia inhibitory factor (LIF) gene mutations in women diagnosed with unexplained infertility and endometriosis have a negative impact on the IVF outcome. A pilot study.

The frequency of functionally relevant mutations of the leukaemia inhibitory factor (LIF) gene in infertile women is significantly enhanced in comparison with fertile controls. The objective of this retrospective cohort study was to evaluate the impact of LIF gene mutations on the outcome of the treatment in women with various causes of infertility. Fifteen infertile women with the G to A transition at position 3400 leading to the valine to methionine exchange at codon 64 were analysed. Group A was made up of women with diagnoses that are frequently accompanied by changes in humoral as well as cell-mediated immunity - idiopathic infertility and endometriosis (N = 7). Group B consisted of patients with polycystic ovary syndrome (PCOS), andrological factor, tubal factor and hyperprolactinaemia (N = 8). The control group comprised 136 infertile women with no LIF gene mutation diagnosed with idiopathic infertility and endometriosis (N = 37) (group C) and patients with PCOS, tubal and andrological factor (N = 99) (group D). Seven of the mutation-positive patients were successfully treated by in vitro fertilization (IVF), but nobody in this group was diagnosed with idiopathic infertility and only one with endometriosis, which means that there is a statistically significant difference in the pregnancy rates between groups A and B (P = 0.01, Fisher's 2 by 2 exact test) but no statistically significant difference when comparing patients with the LIF gene mutation (group A+B) to no LIF gene mutation (group C+D). The results suggest that in mutation-positive women the idiopathic infertility and endometriosis have a negative impact on the outcome of IVF treatment.

[IgG antibodies against laminin-1 in serum and in peritoneal fluid in patients with decreased fertility]. / IgG protilátky namírené proti lamininu-1 v séru a v peritoneální tekutine u pacientek se snízenou plodností.

OBJECTIVE: To study of IgG antibodies against laminin-1 (IgG-a-Ln-1) in patients with decreased fertility, especially with endometriosis and with diagnosis of polycystic ovaries. DESIGN: Prospective study. SETTING: Special Consultation for Immunology of Reproduction and Research Laboratories for Reproductive Immunology, Department of Gynaecology and Obstetrics, Medical School of Charles University and Faculty Hospital, Pilsen. METHODS: Commercial ELISA kits for detection of IgG antibodies against laminin-1 in serum (S) and peritoneal fluid (PF) in 120 patients after diagnostic laparoscopy, and in serum in 277 patients after 2-3 in vitro fertilizations and with diagnosed endometriosis. RESULTS: Only 16 positive results in IgG-a-Ln-1 antibodies (more than 15 IU/ml) were detected in sera and peritoneal fluids in the group of 120 women after laparoscopy. We have not detected any serum IgG-a-Ln-1 positivity in the group of 277 patients after 2-3 unsuccesfull IVF and with anamnestical date about endometriosis together. CONCLUSION: Increased levels of IgG-a-Ln are in particular found in endometriosis II-III and in diagnosis of polycystic ovaries, in female diseases also related to autoimmune process.

[Isolation and immunology identification of spermagglutinating antibodies from human serum]. / Izolace a imunologická identifikace spermaglutinacní protilátky z lidského séra.

OBJECTIVE: Isolation of spermagglutinating antibodies and their assesment. DESIGN: Retrospective study. SETTING: Special consulting for reptoduction immunology, Department of Obstetrics and Gynecology, Charles University and Faculty Hospital, Plzen. METHODS: Fractionation of serum samples by liquid exclusion chromatography, examination of full sera and their chromatographic fractions by Friberg teste (Tray Agglutination Test--TAT), indirect antiimmunoglobulin reaction test (i-MAR test) and by supplementar radial immunodiffusiona (RID). RESULTS: Isolation of spermagglutinating fractions of antisperm antibodies positive sera preserved spermagglutinating aktivity and confirmed great spermagglutinating potential of IgM. CONCLUSION: According to assesment of the presence of IgG and IgG we reported possible states of immunisation: actual immunisation with IgM activity, perpetual stimulation (IgG and IgM) and, finaly, anamnestic titres in IgG. These findings can help us to choose an optimal way of treatment. Excluding gel chromatography is suitable method for serum proteins fractionation, but not their identification--presence of antisperm antibodies does not affect the chromatographic spectrum, nor the RID patterns.

[The lifestyle of Pilsner University female students and its effect on vaginal mycosis]. / Zivotní styl plzenských vysokoskolacek a jeho vliv na mykotická onemocnení.

BACKGROUND: Up to now, it does not exit whole look at the life quality of the Pilsner female University students, especially in connection with gynecological history. It is reality that in last years the number of gynecological problems among young women increases such as the number of diseases with immunological background. METHOD: We prepared questionnaire with 36 questions directed to life style, gynecological history, partner's life, food and hygienic habits, and nicotinism. RESULTS: 28 female students from the total group of 141 suffered from vaginal mycosis--22 (78.6%) use pills, 6 (21.4%) no hormonal contraception. 17 (60.7%) prefer flyaway cloth and 9 (32.1%) tight cloth. 141 students mentioned various number (0-25) of their sexual partners: 1-3 (79 students) in 15 cases (19%) mycoses, more than 4 partners (49 students) in 13 (26.5%) mycoses appeared. The first menstruation appeared in o 13 years (10-17 years). High incidence of vaginal mycosis was proven in students using only tampons than hygienic towels. CONCLUSION: Vaginal mycosis is affected by many endogenous and exogenous factors. The main factors in pilsner university female students are hormonal contraception, promiscuity and hygienic habits.