Lactobacillus crispatus-dominated vaginal microbiome and Acinetobacter-dominated seminal microbiome support beneficial ART outcome.

INTRODUCTION: Despite the considerable progress made in assisted reproductive technologies (ART), the implantation rate of transferred embryos remains low and in many cases, the reasons for failure remain unclear. We aimed to determine the potential impact of female and male partners' reproductive tract microbiome composition on ART outcome. MATERIAL AND METHODS: The ART couples (n = 97) and healthy couples (n = 12) were recruited into the study. The smaller healthy group underwent a careful selection according to their reproductive and general health criteria. Both vaginal and semen samples were subjected to 16S rDNA sequencing to reveal the bacterial diversity and identify distinct microbial community types. Ethics statement The study was approved by the Ethics Review Committee on Human Research of Tartu University, Tartu, Estonia (protocol no. 193/T-16) on 31 May 2010. Participation in the research was voluntary. Written informed consent was obtained from all study participants. RESULTS: The men with Acinetobacter-associated community who had children in the past, had the highest ART success rate (P < 0.05). The women with bacterial vaginosis vaginal microbiome community and with L. iners-predominant and L. gasseri-predominant microbiome had a lower ART success rate than women with the L. crispatus-predominant or the mixed lactic-acid-bacteria-predominant type (P < 0.05). The 15 couples where both partners had beneficial microbiome types had a superior ART success rate of 53%, when compared with the rest of the couples (25%; P = 0.023). CONCLUSIONS: Microbiome disturbances in the genital tract of both partners tend to be associated with couple's infertility as well as lower ART success levels and may thus need attention before the ART procedure. The incorporation of genitourinary microbial screening as a part of the diagnostic evaluation process may become routine for ART patients if our results are confirmed by other studies.

Seminal microbiome in men with and without prostatitis.

OBJECTIVES: To profile the seminal microbiome applying next generation sequencing. METHODS: Semen samples of 67 men were involved in the study (21 men with and 46 men without prostatitis). Seminal microbiomes were profiled applying the method that uses combinatorial sequence tags attached to polymerase chain reaction primers that amplify the ribosomal ribonucleic acid V6 region. Amplified polymerase chain reaction products were sequenced using an Illumina paired-end protocol on HiSeq2000 platform. RESULTS: The most abundant phylum in semen was Firmicutes, comprising nearly half of the sequences found (median 41.7%, quartiles 28.5-47.2%) followed by Bacteroidetes, Proteobacteria and Actinobacteria. The counts of lactobacilli were higher in healthy men than prostatitis patients (27% [20.2-34.6%] vs 20.2% [4.9-25.0%]; P = 0.05), especially for Lactobacillus iners. Proteobacteria comprised higher proportions in prostatitis patients than healthy men. The species richness was higher in prostatitis patients than healthy men (inverted Simpson index 13.5 ± 5.8 vs 10.3 ± 4.0). CONCLUSIONS: The semen of chronic prostatitis patients contains fewer health-supporting lactobacilli, and has higher species diversity than that of healthy men. Firmicutes (especially lactobacilli), Bacteroidetes, Proteobacteria and Actinobacteria comprise the highest proportion of seminal microbiome.

T Cell Distribution in Relation to HIV/HBV/HCV Coinfections and Intravenous Drug Use.

Intravenous drug use (IDU) is one of the most important transmission routes for blood borne viruses, including human immunodeficiency virus (HIV), hepatitis B virus (HBV), and hepatitis C virus (HCV). These infections alter the subset distributions of T cells; however, knowledge of such effects during HIV, HBV, and or HCV coinfection is limited. Therefore, we aimed to evaluate any associations between T cell distribution and the presence of HIV, HBV, and HCV coinfections among persons who inject drugs (PWID). Blood samples from 88 Caucasian PWID (mean age 30; 82% male) and 47 age-matched subjects negative for all three infections (mean age of 29; 83% male) were analyzed. The T cell markers CD3, CD4, CD8, CD45RA, CCR7, HLA-DR, and CCR5 were assessed using flow cytometry. Of the PWID, 40% were HIV+HBV+HCV+, 20% HBV+HCV+, 19% HCV+, and 13% negative for all three infections. The HIV+HBV+HCV+ PWID had lower percentages of CD4+ and higher percentages of CD8+ cells compared to triple negative PWID (p < 0.001 in all cases). The only difference between HBV+HCV+ with triple negative PWID was the lower CD4+ cell percentages among the former (52.1% and 58.6%, p = 0.021). Triple negative PWID had higher immune activation and number of CCR5+ cells compared to the controls. We suggest that the altered T cell subset distribution among PWID is mainly triggered by HIV infection and or IDU, while HBV and or HCV seropositivity has minimal additional effects on CD4+ cell distribution.

Differences in T cell distribution and CCR5 expression in HIV-positive and HIV-exposed seronegative persons who inject drugs.

Some individuals remain uninfected despite repeated exposure to HIV. This protection against HIV has been partly associated with altered T cell subset distributions and CCR5 expression levels. However, the majority of studies have been conducted in sexually exposed subjects. We aimed to assess whether HIV infection and intravenous drug use were associated with differences in CCR5 expression, immune activation on the CD4+ and CD8+ T cells and T cell distribution among Caucasian persons who inject drugs (PWIDs). Analyses of the data from 41 HIV-positive PWIDs, 47 HIV-exposed seronegative PWIDs (ESNs) and 47 age- and gender-matched HIV-negative non-drug users are presented. Of all of the study subjects, 111 (82 %) were male, and the median age was 29 years. T cell phenotyping was performed in peripheral blood mononuclear cells with multicolour flow cytometry using anti-CD3, CD4, CD8, CD45RA, CD45RO, HLA-DR and CCR5 antibodies. The ESNs exhibited greater levels of immune activation and higher percentages of CD4+ CD45RA+RO+ and CD8+ CD45RA+RO+ cells compared to the controls but not the HIV-positive people. The CCR5 expression on the CD4+ T cell subsets in the ESNs was lower than that in the controls but similar to that the HIV positives. The percentages of CCR5+ T cells were similar in all study groups and in most of the studied cell populations. Intravenous drug use was similarly associated with differences in T cell subset distributions and CCR5 expression among both the HIV-positive and HIV-negative PWIDs compared with the controls.

Coryneform bacteria in human semen: inter-assay variability in species composition detection and biofilm production ability.

BACKGROUND: Coryneform bacteria constitute an important segment of male urogenital microbiota. They have been generally considered as saprophytes, although some species have been associated with prostatitis as well. At the same time, biofilm infections have been suspected as a cause of prostatitis. OBJECTIVE: To identify a set of coryneform bacteria isolated from semen of either healthy men or prostatitis patients applying different methods to reveal inter-assay variability and to determine their ability of adhesion and biofilm production. DESIGN: Coryneform bacteria were identified by API Coryne 2.0 biochemical identification system and 16S rDNA sequencing using different primer sets. Quantitative assessment of biofilm production was performed using crystal violet binding assay method. RESULTS: The most common species were Corynebacterium seminale, C. minutissimum, and Dermabacter hominis. Altogether 14 species and related genera were found. We observed the best inter-assay agreement when identifying C. seminale. Biofilm was observed in 7 out of 24 strains. The biofilm-producing strains belonged to Arthrobacter cumminsii, Dermabacter hominis, C. minutissimum, and Actinomyces neuii. No differences were found between the strains originating from prostatitis patients and healthy men. Dermabacter hominis strains were more potent biofilm producers than C. seminale strains (p=0.048). CONCLUSIONS: We can conclude that a wide variety of coryneform bacteria can be found from the male genital tract, although their exact identification is problematic due to insufficient representation in databases. Nearly one third of the strains are able to form biofilm that may give them an advantage for surviving several host- and treatment-related conditions.

Male infertility: decreased levels of selenium, zinc and antioxidants.

In this study, we aimed to compare the level of zinc, selenium, glutathione peroxidase activity and antioxidant status in following populations of men: severe inflammation in prostate (>10(6) white blood cells in prostate secretion; n=29), severe leukocytospermia, (>10(6) white blood cells in semen; n=31), mild inflammation, (0.2-1M white blood cells in semen or prostate secretion; n=24), non-inflammatory oligozoospermia (n=32) and healthy controls (n=27). Male partners of infertile couples had reduced level of antioxidative activity, selenium and zinc in their seminal plasma. Most importantly, reduced selenium levels were evident in all patient groups regardless of inflammation status. Therefore, these patients might gain some benefit from selenium supplementation.

Oxidative stress--cause or consequence of male genital tract disorders?

BACKGROUND: Inflammatory prostatitis patients are characterized by oxidative stress (OxS) at local and systemic levels. Less is known about the occurrence of OxS in the case of other frequent male genital tract disorders. METHODS: The study included 196 men: controls (n = 28), asymptomatic inflammatory (NIH category IV) prostatitis patients (n = 21), non-inflammatory (NIH category IIIb) chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) patients (n = 48), inflammatory (NIH category IIIa) CP/CPPS patients (n = 44), benign prostate hyperplasia (BPH) patients (n = 33), and patients with BPH and NIH IV category prostatitis (n = 22). In all subjects, 8-isoprostanes (8-EPI) in urine were determined by competitive enzyme-linked immunoassay. RESULTS: The levels of 8-EPI were substantially higher in all diseased groups-inflammatory CP/CPPS (P < 0.001), non-inflammatory CP/CPPS (P = 0.03), asymptomatic inflammatory prostatitis (AIP; P = 0.02), BPH (P = 0.007), and BPH + AIP (P = 0.014) in comparison with controls. Importantly, our study showed that OxS is also present in the case of NIH IIIb category prostatitis when the patients have just chronic pelvic pain but no inflammation in prostate-specific materials, as well as in the patients with just lower urinary tract symptoms without pain or overt inflammation. CONCLUSIONS: This study revealed that several male genital tract disorders-BPH and different forms of prostatitis (NIH categories IIIa, IIIb, and IV)-are tightly interconnected via OxS-mediated pathways. Acknowledging OxS as an important pathogenesis mechanism of these diseases helps to open up new horizons for their treatment.

Are prostatitis symptoms associated with an isoprostane-mediated vicious circle?

Prostatitis is a disease that seriously affects the quality of patients' life. In the majority of cases, chronic prostatitis (chronic pelvic pain syndrome--CPPS) has an unclear pathogenesis. Anti-inflammatory and anti-infectious treatments have remained controversial. According to the latest research, prostatitis has been associated with oxidative stress (OxS) and/or OxS-related genetic polymorphisms. We have observed that prostatitis patients have systemic OxS in case of inflammation and pain. We propose a new explanation for the role of OxS in the pathogenesis of prostatitis and describe the putative OxS-related pathways in detail. The neural vicious circle starts by activation of primary sensory afferents. Glutamate mediates the signal to the neurons in the dorsal horn of the spinal cord, and facilitates calcium influx into their mitochondria. The latter causes an increased production of superoxide radicals. If the superoxide production is not effectively controlled by mitochondrial superoxide dismutase (Mn-SOD), then superoxide leads to OxS and lipid peroxidation. Consequent release of electrophilic lipid peroxidation products (LPP) from dorsal horn of the spinal cord causes pain by activating the primary sensory afferents, again. Additional LPP-mediated causes of pain include glutathione depletion and neuron sensitisation by isoprostanes. Excretion of LPP into urine may exert positive feedback as well. Currently, different information exists about chronic prostatitis (inflammation, pain, oxidative stress, neural sensitisation, lower urinary tract symptoms). The clear links between these data are actually absent. We propose that vicious circle based on LPP, especially isoprostanes, is the linking mechanism.

Influence of sexual intercourse on genital tract microbiota in infertile couples.

Several studies have suggested the association of disturbed genital tract microbiota with infertility. Our aim was to clarify the influence of sexual intercourse on partner's genital tract microbiota in infertile couples. Seventeen couples were studied, and in 5 men inflammatory prostatitis (IP) was diagnosed. Semen samples were collected during menstruation of the female counterpart, two self-collected vaginal samples were taken 3-5 days later - before intercourse and 8-12 h after intercourse. Ureaplasma parvum was found in 59% of women, its prevalence was higher in women whose partner had IP, as well as in half of their male partners. Sexual intercourse caused significant shifts in vaginal microbiota - increase of Nugent score and shifts in cultured microbiota (emergence and disappearance of several species). These changes were less expressed in the presence of normal vaginal microbiota but more prominent in the partners of IP men. These changes may interfere with fertilization.

Coryneform bacteria in semen of chronic prostatitis patients.

Our aim was to compare the presence and species composition of coryneform bacteria in chronic prostatitis patients and controls. Semen of 50 men with inflammatory prostatitis and 59 controls (without pelvic pain/discomfort complaints and leukocytospermia) was investigated. First-catch urine was additionally investigated in 36 men (30 with and 6 without prostatitis). Coryneform bacteria were found in semen of 76% men with inflammatory prostatitis and 83% controls. More than half of the isolates were identified as Corynebacterium seminale. Prostatitis patients with severe leukocytospermia (>1 million white blood cells per ml) harboured significantly more Corynebacterium group G (33% vs. 2%, p = 0.0003) and Arthrobacter sp. (17% vs. 2%, p = 0.03) in comparison with controls. Nine species of coryneforms with high concentration (>or=10,000 CFU per ml) were found in prostatitis patients as against only four species in controls. Half of the men harboured corynebacteria in semen as well as in urine, 22% of men in semen only, and 3% in urine only. The total concentration of coryneforms was greater in semen than in urine (median 5000 vs. 100 CFU per ml, p = 0.053). We suggest that although coryneforms are generally considered as saprophytes, they are not uniform and some species (Corynebacterium group G and Arthrobacter sp.) may be associated with inflammatory prostatitis.

Seminal microflora in asymptomatic inflammatory (NIH IV category) prostatitis.

OBJECTIVES: To prove the need for the quantitative full-microflora semen analysis for determining the role of microorganisms in the etiology of asymptomatic inflammatory prostatitis, and to correlate the seminal white blood cell (WBC) counts with interleukin 6 (IL-6) levels. METHODS: Thirty-seven men with asymptomatic inflammatory (National Institutes of Health [NIH] IV category) prostatitis and 32 controls were investigated by using routine semen analysis, IL-6 levels of seminal plasma, and quantitative microbiological analysis of semen. RESULTS: The IL-6 concentration in seminal plasma was significantly higher in NIH IV category prostatitis patients than in the controls, and was in good correlation with the WBC count in semen (r=0.74, p<0.001). In most of the specimens, the counts of anaerobic microorganisms were equal to or outnumbered the aerobic ones. One to eight different microorganisms could be found in any particular semen sample, and the total concentration of microorganisms ranged from 2.0 to 7.5log(10)CFU/ml. Both parameters were significantly higher in NIH IV category prostatitis patients than in controls (median: 4.8 vs. 3.9log(10)CFU for total concentration, p<0.001; median: 5 vs. 3 for number of different species, p=0.004). CONCLUSIONS: Unlike the controls the NIH IV category prostatitis patients harbour abundant polymicrobial microbiocenosis in their semen, containing anaerobic, microaerophilic and aerobic bacteria. Detection of IL-6 in seminal plasma serves as an additional tool for diagnosing NIH IV category prostatitis.

Mycoplasmas in semen of chronic prostatitis patients.

OBJECTIVE: To evaluate the occurrence of mycoplasmas in the semen of chronic prostatitis patients. MATERIAL AND METHODS: Genital mycoplasmas (Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma urealyticum, Ureaplasma parvum) were sought in the semen of 121 chronic prostatitis patients [38 National Institutes of Health (NIH) category IIIa, 59 NIH category IIIb and 24 NIH category IV] and 40 controls. The commercially available kit Mycoplasma IST was applied to the semen samples of all 161 men, and polymerase chain reaction (PCR) to those of 60 randomly selected men. RESULTS: Ureaplasmas were found in all study groups (at frequencies ranging from 12% to 25%) using the Mycoplasma IST test, but M. hominis was found only in one NIH category IIIb patient. Using PCR, most of the ureaplasmas appeared to be U. parvum, which was found in all prostatitis groups (18% of NIH category IIIa, 15% of NIH category IIIb and 25% of NIH category IV patients) but not in the controls. M. genitalium was found in 18% of the NIH category IIIa patients. All of the mycoplasmas occurred significantly more frequently in prostatitis patients than in controls and in NIH category IIIa patients than in controls. CONCLUSION: Mycoplasmas occur more frequently in the semen of prostatitis patients than in that of healthy controls, with U. parvum being the most frequently occurring species.