The analysis of semen microbiota is difficult due to the lack of established criteria for interpretation of microbiological tests. The aim of the study was to determine the stable clusters of semen microbiota analyzed by real-time PCR in samples with normozoospermia. Semen samples of 227 men with normal spermiograms were included in the study. The quantity of total bacterial DNA and at least one group of microorganisms was more than 103 GE/ml in 107 (41.7%) samples. Four stable microbiota clusters with the prevalence of a specific microorganism group were distinguished in these samples: obligate anaerobes (OA) cluster (proportion in the centroid — 81.1%); Lactobacillus spp. cluster (proportion in the centroid — 64.3%); gram-positive facultative anaerobes (GPFA) cluster (proportion in the centroid — 92.5%); Enterobacteriaceae/Enterococcoccus (EE) cluster (proportion in the centroid — 80.8%). The clusters were ranked by frequency of occurrence: OA cluster was the most prevalent (43 (40.2%) of 107), second-most frequent were GPFA-cluster (27 (25.2%)) and Lactobacillus-cluster (22 (20.6%)). EE-dominated cluster was found in 15 (14.0%) cases.

Одной из причин мужского бесплодия могут быть воспалительные заболевания урогенитального тракта, развитие которых в ряде случаев ассоциировано с условно-патогенными микроорганизмами (УПМ). В связи с этим актуальна проблема внедрения современных методов выявления и идентификации УПМ в урогенитальном тракте. Целью работы было провести сравнительный анализ результатов исследования микробиоты эякулята мужчин репродуктивного возраста с помощью количественной полимеразной цепной реакции (ПЦР) (тест Андрофлор) и культурального метода. Исследовали 86 образцов эякулята, собранных у мужчин в возрасте 18–57 лет после соблюдения полового воздержания в течение 3–5 суток. При культуральном исследовании в 50% образцов наблюдали рост грамположительных факультативно анаэробных бактерий в количестве менее 103 КОЕ/мл; в 16,3% образцов — роста бактерий отмечено не было. При использовании ПЦР в реальном времени (ПЦР-РВ) в каждом образце выявляли 8–15 групп микроорганизмов (в том числе определяли преобладающую) в количестве 102–106 ГЭ/мл. Во всех 86 образцах были обнаружены облигатные анаэробы, которые не культивируются in vitro. Преобладающие группы микроорганизмов, определяемые в ПЦР-РВ, были выявлены культуральным методом только в 24,4% случаев.

Study question Is there any correlation between the total bacterial load and the lactobacilli quantities in the vaginal and endometrial microbiomes in reproductive-age women? Summary answer There was no correlation between the vaginal and endometrial total bacterial loads and only a weak positive correlation between the quantities of lactobacilli. What is known already The Lactobacilli-dominated microbiota is considered to be the most favorable type of microbiota in the uterine cavity. It is associated with increased reproductive success in women undergoing in vitro fertilization. Whereas the non-Lactobacillus dominated microbial communities are more frequent in women with poor pregnancy outcomes. When analyzing endometrial microbiota, one of the challenges is sampling. Transvaginal sample intake involves the possibility of contaminating the samples with vaginal microbiota. Moreover, it is an invasive procedure leading to the development of infectious inflammatory diseases of the upper genital tract. Thus, researchers are currently searching for predictors of the state of endometrial microbiota. Study design, size, duration It is a cross-sectional study of the vaginal end endometrial micorbiomes from 64 reproductive-age women. Endometrial and vaginal samples were collected simultaneously on days 7–10 of the menstrual cycle. To avoid contamination by vaginal microbiota, Endobrush Standard for Endometrial Cytology (Laboratoire C.C.D.; France) was used for endometrial sampling. Participants/materials, setting, methods The study included women who came to the “Garmonia” Medical Center (Yekaterinburg, Russia) seeking infertility treatment. The average age of the patients was 32.2±5.0. DNA from vaginal and endometrial samples was extracted using PREP-NA-PLUS kit (DNA-Technology, Russia). Vaginal and endometrial microbiota was analyzed using Femoflor real-time PCR kit and DTprime 4M1 thermocycler (DNA-Technology, Russia). Main results and the role of chance Total bacterial load (TBL) in vaginal discharge was 3.8–7.9 lg (median — 7.1, interquartile range — 6.6–7.4). TBL in the endometrial samples was 0–5.1 lg (median — 3.9, interquartile range — 3.6–4.2). There was no correlation between TBL values in vaginal discharge and endometrial samples (Spearman’s rho — 0.247, p = 0.049). Lactobacilli quantities in vaginal discharge were 4.5–8.3 lg (median — 7.2, interquartile range — 6.4–7.6), in endometrial samples — 0–5.1 lg (median — 3.7, interquartile range — 3.1–4.2). There was a weak positive correlation between lactobacilli quantities in vaginal and endometrial samples (Spearman’s rho — 0.362, p = 0.003). The proportion of lactobacilli in vaginal discharge was 1–100% (median — 100%, interquartile range — 95–100%), in the endometrial samples — 0–100% (median — 96%, interquartile range — 25–100%). There was no correlation between lactobacilli proportions in vaginal and endometrial samples (Spearman’s rho — 0.225, p = 0.074). Furthermore, there was no correlation between lactobacilli quantity in the vagina and their proportion in the endometrial microbiota (Spearman’s rho — 0.294, p = 0.018). There was only a weak positive correlation between the quantities of lactobacilli in vaginal and endometrial samples. Vaginal TBL values and lactobacilli proportions did not correlate with lactobacilli quantities and proportions in the endometrial samples. Limitations, reasons for caution The study was conducted on a small sample. Moreover, it is notoriously difficult to interpret the analysis results for endometrial microbiota due to the high risk of contamination and its low microbial biomass. Wider implications of the findxings: Apparently, there is no obvious link between the vaginal and endometrial microbiomes. It is possible that, apart from vaginal microbiota, there are other predictors which could allow us to assume whether lactobacilli are present in the endometrial microbiota. Trial registration number Not applicable

Критерии нормы для микробиоценоза влагалища здоровой женщины остаются предметом для дальнейшего изучения. Субъективный характер формирования группы «здоровых женщин» при проведении исследований не позволяет счи-тать получаемые результаты однозначными. В данной работе мы сравнивали частоту встречаемости нормоценозов различных типов и дисбиоза влагалища у женщин, поделенных на группы в зависимости от характера их обследования (мнение женщины, осмотр врача, микроскопическое исследование). Были обследованы 234 женщины репродуктивно-го возраста из г. Екатеринбурга (средний возраст -30,3 ± 6,6 лет). Оценку микробиоценоза и определение видового состава лактобацилл (L. crispatus, L. iners, L. jensenii, L. gasseri, L. johnsonii, L. vaginalis) провели методом полимеразной цепной реакции с детекцией результатов в режиме «реального времени» с использованием тест-системы «Фемофлор» («НПО ДНК-Технология», Россия) и наборов реагентов для научного применения той же компании-производителя. У каждой пятой женщины репродуктивного возраста, считающей себя здоровой, был выявлен дисбиоз. Нормоце-ноз в этой группе чаще всего характеризовался преобладанием L. iners, а вариант нормоценоза с преобладанием L. crispatus определяли только у каждой третьей женщины. В то же время вариант нормоценоза с преобладанием L. crispatus был обнаружен у 46,2 % женщин, которые были отнесены к группе клинически здоровых на основании опроса, осмотра врача и микроскопического исследования. Это говорит о том, что клиническая оценка состояния нижних отделов гениталий без учета данных микроскопии носит субъективный характер и в ряде случаев не позволяет выявить у пациентки дисбиоз влагалища. NORMAL VAGINAL MICROBIOTA: PATIENT'S SUBJECTIVE EVALUATION, PHYSICAL EXAMINATION AND LABORATORY TESTSCriteria of normality for the vaginal microbiota of healthy women are still a subject of discussion. A decision to assign a study participant to a group of healthy individuals is quite subjective if based on the absence of complaints and physical examination only, which renders study results ambiguous. Below we compare occurrence of the normal vaginal flora and vaginal dysbiosis in women divided into 3 groups according to the examination type (patient's subjective evaluation of her condition, physical examination, and laboratory tests). We examined 234 women of reproductive age from Yekaterinburg (mean age was 30.3 ± 6.6 years). Microbiota composition and lactobacillus diversity (L. crispatus, L. iners, L. jensenii, L. gasseri, L. johnsonii, L. vaginalis) were evaluated by real-time polymerase chain reaction using the Femoflor assay and reagent kits by DNATechnology, Russia. One in 5 women of reproductive age who had no health complaints was found to have dysbiosis. The normal microbiota of those women was dominated mostly by L.iners, while dominant L. crispatus were observed in every third participant. Prevailing L. crispatus were also found in the normal microbiota of 46.2 % of women who were considered healthy based on the doctor's examination and laboratory tests. ...

Aim. To compare the species composition of lactobacilli in women of reproductive age using real time PCR on urogenital samples before and after culturing microorganisms in Blood-Serum-Y^ast extract-Agar (BSYA). Materials and methods. Using real time PCR, we have examined the species composition of vaginal lactobacilli in 25 healthy women of reproductive age. Samples of urogenital swabs (endocervical and vaginal swabs) were studied. Species identification has been carried out twice: in the native clinical material and in the samples received after using microorganism culture technique in BSYA. Results. After culturing vaginal microorganisms in BSYA, L. iners predominance was found only in 2 cases (8%). At the same time, when using native clinical material this species predominance was discovered in 10 samples (40%). Conclusion. When culturing lactobacilli in BSYA, the growth of L. iners is stunted compared to other species of vaginal lactobacilli. The use of cultural testing turned out to be ineffective in case of predominance of L. iners, the species associated with an increased risk of vaginal dysbiosis.