Andrological effects of SARS-Cov-2 infection: a systematic review and meta-analysis.

PURPOSE: The short- and long-term andrological effects of coronavirus disease 2019 (COVID-19) have not been clarified. Our aim is to evaluate the available evidence regarding possible andrological consequences of COVID-19 either on seminal or hormonal parameters. The safety of the COVID-19 vaccines in terms of sperm quality was also investigated. METHODS: All prospective and retrospective observational studies reporting information on severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) mRNA semen and male genitalia tract detection (n = 19), as well as those reporting data on semen analysis (n = 5) and hormonal parameters (n = 11) in infected/recovered patients without any arbitrary restriction were included. RESULTS: Out of 204 retrieved articles, 35 were considered, including 2092 patients and 1138 controls with a mean age of 44.1 ± 12.6 years, and mean follow-up 24.3 ± 18.9 days. SARS-CoV-2 mRNA can be localized in male genitalia tracts during the acute phase of the disease. COVID-19 can result in short-term impaired sperm and T production. Available data cannot clarify long-term andrological effects. Low T observed in the acute phase of the disease is associated with an increased risk of being admitted to the Intensive Care Unit or death. The two available studies showed that the use of mRNA COVID-19 vaccines does not affect sperm quality. CONCLUSIONS: The results of our analysis clearly suggest that each patient recovering from COVID-19 should be monitored to rule out sperm and T abnormalities. The specific contribution of reduced T levels during the acute phase of the infection needs to be better clarified.

Safety issues in semen banks during the COVID-19 pandemic: data from a European survey.

PURPOSE: We performed a survey among European semen banks enquiring safety protocols during the COVID-19 pandemic. We report the experience from a center searching SARS-CoV-2 mRNA in semen of patients undergoing cryopreservation from May 2020 to January 2021. METHODS: A questionnaire was submitted to accredited semen banks of the European Academy of Andrology (EAA) and the Italian Society of Andrology and Sexual Medicine (SIAMS). A total of 22 centers answered to the survey. SARS-CoV-2 mRNA in semen was evaluated by RT-PCR in 111 subjects banking in the Semen Bank of Careggi University Hospital (Florence, Italy). RESULTS: No particularly drastic safety measures were adopted by the majority of the centers to prevent the risk of contamination or transmission of the virus. The most common strategy (77.3%) was the administration of an anamnestic questionnaire. About half of the centers request a negative nasopharyngeal swab (NPS) before cryopreservation. Few centers use a quarantine tank, in case of late response of NPS, and only 4 store in a dedicated tank in case of infection. SARS-CoV-2 mRNA was not found in 111 semen samples cryopreserved in the Florentine bank. CONCLUSIONS: European semen banks use different measures to handle semen samples for cryopreservation during COVID-19 pandemic. The request of NPS is advised to better manage couples undergoing ART and to protect the personnel operating in the bank/ART center. Finally, due to the areas of uncertainties of an almost unknown virus, it is absolutely recommended the use of safe devices for sample handling and storage.

Semen impairment and occurrence of SARS-CoV-2 virus in semen after recovery from COVID-19.

STUDY QUESTION: How is the semen quality of sexually active men following recovery from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection? SUMMARY ANSWER: Twenty-five percent of the men with recent SARS-Cov-2 infections and proven healing were oligo-crypto-azoospermic, despite the absence of virus RNA in semen. WHAT IS KNOWN ALREADY: The presence of SARS-CoV-2 in human semen and its role in virus contagion and semen quality after recovery from coronavirus disease 2019 (COVID-19) is still unclear. So far, studies evaluating semen quality and the occurrence of SARS-CoV-2 in semen of infected or proven recovered men are scarce and included a limited number of participants. STUDY DESIGN, SIZE, DURATION: A prospective cross-sectional study on 43 sexually active men who were known to have recovered from SARS-CoV2 was performed. Four biological fluid samples, namely saliva, pre-ejaculation urine, semen, and post-ejaculation urine, were tested for the SARS-CoV-2 genome. Female partners were retested if any specimen was found to be SARS-CoV-2 positive. Routine semen analysis and quantification of semen leukocytes and interleukin-8 (IL-8) levels were performed. PARTICIPANTS/MATERIALS, SETTING, METHODS: Questionnaires including International Index of Erectile Function and Male Sexual Health Questionnaire Short Form were administered to all subjects. The occurrence of virus RNA was evaluated in all the biological fluids collected by RT-PCR. Semen parameters were evaluated according to the World Health Organization manual edition V. Semen IL-8 levels were evaluated by a two-step ELISA method. MAIN RESULTS AND THE ROLE OF CHANCE: After recovery from COVID-19, 25% of the men studied were oligo-crypto-azoospermic. Of the 11 men with semen impairment, 8 were azoospermic and 3 were oligospermic. A total of 33 patients (76.7%) showed pathological levels of IL-8 in semen. Oligo-crypto-azoospermia was significantly related to COVID-19 severity (P < 0.001). Three patients (7%) tested positive for at least one sample (one saliva; one pre-ejaculation urine; one semen and one post-ejaculation urine), so the next day new nasopharyngeal swabs were collected. The results from these three patients and their partners were all negative for SARS-CoV-2. LIMITATIONS, REASONS FOR CAUTION: Although crypto-azoospermia was found in a high percentage of men who had recovered from COVID-19, clearly exceeding the percentage found in the general population, the previous semen quality of these men was unknown nor is it known whether a recovery of testicular function was occurring. The low number of enrolled patients may limit the statistical power of study. WIDER IMPLICATIONS OF THE FINDINGS: SARS-CoV-2 can be detected in saliva, urine, and semen in a small percentage of men who recovered from COVID-19. One-quarter of men who recovered from COVID-19 demonstrated oligo-crypto-azoospermia indicating that an assessment of semen quality should be recommended for men of reproductive age who are affected by COVID-19. STUDY FUNDING/COMPETING INTEREST(S): None. TRIAL REGISTRATION NUMBER: N/A.

General practitioners' management of symptomatic uncomplicated diverticular disease of the colon by using rifaximin, a non-adsorbable antibiotic.

OBJECTIVE: Symptomatic uncomplicated diverticular disease of the colon (SUDD) is generally managed by gastroenterologists rather than General Practitioners (GPs). The aim of this study was to assess the efficacy of the treatment of SUDD with rifaximin, a non-absorbable antibiotic, in a primary care setting by GPs. PATIENTS AND METHODS: This retrospective, observational study investigated the use of rifaximin at a dose of 400 mg b.i.d. for 5, 7 or 10 days monthly, up to 3 months. The symptoms were reported by the patients using a visual analogic scale (VAS) of 0-10. RESULTS: 286 SUDD patients were enrolled (44.4% of men, average age 70.92±10.98). Respectively, 15 (5.2%) patients received the treatment for 5 days, 205 (71.7%) for 7 days and 66 (23.1%) for 10 days. After three months, a significant reduction of VAS score was observed in almost all symptoms assessed: 135 (47.2%) patients reported no abdominal pain (p<0.001) and 23 (8.1%) reported no symptom. Adverse events related to the treatment were recorded in 3 (1.04%) patients, all of them mild and not requiring interruption of the treatment. Acute diverticulitis occurred in 9 (3.1%) patients, but only 2 of them [0.7% (n=2)] underwent surgery due to complicated diverticulitis. Analysis within the different treatment groups (5, 7 and 10 days) shows that rifaximin treatment is effective in reducing the severity of symptoms in almost all groups except for the constipation in the 5-day group. CONCLUSIONS: Rifaximin can be effectively used by GPs in real-life for the management of SUDD.

Dimethyl fumarate-induced lymphocyte count drop is related to clinical effectiveness in relapsing-remitting multiple sclerosis.

BACKGROUND AND PURPOSE: Dimethyl fumarate (DMF) causes a mean lymphocyte count drop of approximately 30% in relapsing-remitting multiple sclerosis (RRMS) patients. The relationship between this reduction and DMF effectiveness is controversial. The objective was to investigate if the decrease in absolute lymphocyte count (ALC) from baseline during DMF treatment is associated with clinical and magnetic resonance imaging (MRI) disease activity. A secondary aim was to evaluate ALC variations over time in a real-life cohort of DMF-treated patients. METHODS: Demographic, laboratory, clinical and MRI data were collected in this observational multicentre study, conducted on RRMS patients treated with DMF for at least 6 months. Multivariate Cox models were performed to evaluate the impact of 6-month ALC drop on time to no evidence of disease activity (NEDA-3) status loss. NEDA-3 is defined as absence of clinical relapses, MRI disease activity and confirmed disability progression. RESULTS: In all, 476 patients (312 females, age at DMF start 38.4 ± 9.97 years) were analysed up to 5-year follow-up. A greater lymphocyte decrease was associated with a lower risk of NEDA-3 status loss (hazard ratio 0.87, P = 0.01). A worse outcome in patients with lower ALC drop (<11.5%), compared with higher tertiles (11.5%-40.5% and >40.5%), was observed (P = 0.008). The nadir of ALC drop (-33.6%) and 35% of grade III lymphopaenia cases occurred after 12 months of treatment. CONCLUSION: A higher lymphocyte count drop at 6 months is related to better outcomes in DMF-treated patients. A careful ALC monitoring should be pursued up to 24 months of treatment.

The European Academy of Andrology (EAA) ultrasound study on healthy, fertile men: Scrotal ultrasound reference ranges and associations with clinical, seminal, and biochemical characteristics.

BACKGROUND: Scrotal color Doppler ultrasound (CDUS) still suffers from lack of standardization. Hence, the European Academy of Andrology (EAA) has promoted a multicenter study to assess the CDUS characteristics of healthy fertile men (HFM) to obtain normative parameters. OBJECTIVES: To report and discuss the scrotal organs CDUS reference ranges and characteristics in HFM and their associations with clinical, seminal, and biochemical parameters. METHODS: A cohort of 248 HFM (35.3 ± 5.9years) was studied, evaluating, on the same day, clinical, biochemical, seminal, and scrotal CDUS following Standard Operating Procedures. RESULTS: The CDUS reference range and characteristics of the scrotal organs of HFM are reported here. CDUS showed a higher accuracy than physical examination in detecting scrotal abnormalities. Prader orchidometer (PO)- and US-measured testicular volume (TV) were closely related. The US-assessed TV with the ellipsoid formula showed the best correlation with the PO-TV. The mean TV of HFM was ~ 17 ml. The lowest reference limit for right and left testis was 12 and 11 ml, thresholds defining testicular hypotrophy. The highest reference limit for epididymal head, tail, and vas deferens was 12, 6, and 4.5 mm, respectively. Mean TV was associated positively with sperm concentration and total count and negatively with gonadotropins levels and pulse pressure. Subjects with testicular inhomogeneity or calcifications showed lower sperm vitality and concentration, respectively, than the rest of the sample. Sperm normal morphology and progressive motility were positively associated with epididymal head size/vascularization and vas deferens size, respectively. Increased epididymis and vas deferens sizes were associated with MAR test positivity. Decreased epididymal tail homogeneity/vascularization were positively associated with waistline, which was negatively associated with intratesticular vascularization. CDUS varicocele was detected in 37.2% of men and was not associated with seminal or hormonal parameters. Scrotal CDUS parameters were not associated with time to pregnancy, number of children, history of miscarriage. CONCLUSIONS: The present findings will help in better understanding male infertility pathophysiology, improving its management.

LH supplementation of ovarian stimulation protocols influences follicular fluid steroid composition contributing to the improvement of ovarian response in poor responder women.

In this prospective study, we evaluated the steroid levels in 111 follicular fluids (FF) collected from 13 women stimulated with FSH monotherapy and 205 FF collected from 28 women stimulated with FSH + LH because of a previous history of hypo-responsiveness to FSH. Steroid levels were measured by HPLC/MS-MS and related to ovarian stimulation protocol, oocyte maturity, fertilization and quality of blastocysts, after individually tracking the fate of all retrieved oocytes. 17-Hydroxy-Progesterone, Androstenedione, Estradiol and Estrone were significantly higher in the FSH + LH protocol. Progesterone, 17-Hydroxy-Progesterone and Estradiol were more expressed in FF yielding a mature oocyte (p < 0.01) in the FSH + LH protocol. FF Progesterone concentration was correlated with the rate of normal fertilization in the FSH protocol. None of the FF steroids measured were associated with blastocyst quality and achievement of pregnancy. Our results indicate that LH supplementation in hypo-responsive women modifies ovarian steroid production, mimicking physiological production better and likely contributing to an improved ovarian response. Employing a correct methodological procedure to evaluate the relationship between FF steroid hormones and assisted reproduction outcomes, our study reveals that some steroids in single follicles may be helpful in predicting oocyte maturity and fertilization.

SARS-CoV-2 infection, male fertility and sperm cryopreservation: a position statement of the Italian Society of Andrology and Sexual Medicine (SIAMS) (Società Italiana di Andrologia e Medicina della Sessualità).

PURPOSE: The recent pandemic of severe acute respiratory syndrome (SARS) due to coronavirus (CoV) 2 (SARS-CoV-2) has raised several concerns in reproductive medicine. The aim of this review is to summarize available evidence providing an official position statement of the Italian Society of Andrology and Sexual Medicine (SIAMS) METHODS: A comprehensive Pubmed, Web of Science, Embase, Medline and Cochrane library search was performed. Due to the limited evidence and the lack of studies, it was not possible to formulate recommendations according to the Oxford 2011 Levels of Evidence criteria. RESULTS: Several molecular characteristics of the SARS-CoV-2 can justify the presence of virus within the testis and possible alterations of spermatogenesis and endocrine function. Orchitis has been reported as a possible complication of SARS-CoV infection, but similar findings have not been reported for SARS-CoV-2. Alternatively, the orchitis could be the result of a vasculitis as COVID-19 has been associated with abnormalities in coagulation and the segmental vascularization of the testis could account for an orchitis-like syndrome. Finally, available data do not support the presence of SARS-CoV-2 in plasma seminal fluid of infected subjects. CONCLUSION: Data derived from other SARS-CoV infections suggest that in patients recovered from COVID-19, especially for those in reproductive age, andrological consultation and evaluation of gonadal function including semen analysis should be suggested. Studies in larger cohorts of currently infected subjects are warranted to confirm (or exclude) the presence of risks for male gametes that are destined either for cryopreservation in liquid nitrogen or for assisted reproduction techniques.

The exploration of a new environment leads to the modulation of gene expression for prolonged times in the rat.

In the present study we performed a transcriptional analysis in order to evaluate changes in gene expression induced by exploration in prolonged times. The analysis was carried out 3, 10 and 20 days after exploration. We analyzed the modulation of the expression levels of Pfn2, Casp3, Pdrg1, Pea15, Ywhaz genes which previously were found not modulated 2 days after exploration. Our data show that the expression of Pfn2, Casp3, Pdrg1, Pea15, Ywhaz genes was modulated at 10 or 20 days. The transcript, whose expression had been evaluated with the qRT-PCR, code for proteins which belong to the following functional categories: synaptic modulation, apoptosis, signal transduction. It is interesting to note that the modulation of the expression of these genes was evident some days after environmental exploration, and not previously at 2 days after conditioning as occurred after contextual fear conditioning (CFC). Hence it is possible to hypothesize that the spatial memory processes require a longer period of elaboration than the emotional ones, fundamental for the survival of the species.

Acute effects on human sperm exposed in vitro to cadmium chloride and diisobutyl phthalate.

Epidemiological studies reported a negative relationship between concentrations of heavy metals and phthalates in seminal fluid and semen quality, likely compromising male fertility potential. The aim of this study was to investigate the in vitro effects of cadmium chloride (CdCl2), a common heavy metal, and diisobutyl phthalate (DIBP), a common phthalate ester, on human sperm functions necessary for fertilization. After in vitro incubation of spermatozoa with 10 µM CdCl2 or 100 and 200 µM DIBP for 24 h, a significant decrease of sperm progressive and hyperactivated motility was observed. The exposure to each of the two toxic agents also induced spontaneous sperm acrosome reaction and blunted the physiological response to progesterone. Both agents induced an increase of caspase activity suggesting triggering of an apoptotic pathway. Our results suggest that acute exposure of spermatozoa to these pollutants may impair sperm ability to reach and fertilize the oocyte.

Sperm selection with density gradient centrifugation and swim up: effect on DNA fragmentation in viable spermatozoa.

Subjects increasing sperm DNA fragmentation (sDF) during Density Gradient Centrifugation (DGC), a common sperm selection procedure in Assisted Reproduction Techniques (ARTs), experience a 50% lower probability of pregnancy. Hence, identification of these subjects is of clinical importance. Here, we investigated whether such subjects are identified with higher accuracy detecting DNA fragmentation in viable (viable sDF) instead of total spermatozoa (total sDF) and whether swim up, an alternative procedure to DGC, does not increase sDF. With DGC, we identified 10/20 subjects increasing total sDF, and 2 more subjects using viable sDF. With swim up, we identified 8/40 subjects increasing total sDF, and 8 more subjects using viable sDF. In addition, viable sDF reveals more accurately the increase of the damage when it occurs. Finally, a multivariate analysis demonstrated that the proportional increase of sDF was higher after DGC respect to swim up. In conclusion, viable sDF is a more accurate parameter to reveal the increase of the damage by selection both with swim up and DGC. Swim up increases sDF in some samples, although at a lesser extent than DGC, suggesting that it should be used to select spermatozoa for ARTs when possible.

Vitamin D receptor expression and acid sphingomyelinase activity in prefrontal region of a learning animal model.

Contextual fear conditioning (CFC) paradigm is routinely used to study fear-based learning in animals and it provides a useful model for understanding fear and anxiety in human. In the present study, such model was used following the previously established CFC protocol, and immunohistochemistry, enzymatic activity and western blotting analysis approaches were used to identify the expression of acid sphingomyelinase (aSMase) and vitamin D receptor (VDR) in prefrontal region brain of rat. Results revealed an increase of aSMase activity in conditioned rats, suggesting an apoptotic condition in such animals. In addition, an increase of density and organization of axonal neurofilaments and of VDR expression has been observed in brain of conditioned rats, supporting an induction of growth and organization of new neurons in prefrontal regions, whose contribution to various aspects of contextual fear learning is still largely unknown.

The prevalence of midline prostatic cysts and the relationship between cyst size and semen parameters among infertile and fertile men.

STUDY QUESTION: When is the investigation and treatment of midline prostatic cysts (MPC) of clinical value in the work-up of males of infertile couples? SUMMARY ANSWER: With a prevalence of 10.2% in infertile men, MPC should be investigated according to a seminal algorithm detecting a MPC volume >0.117 ml, which may impair semen parameters, and could be treated to improve sperm count and achieve natural pregnancy. WHAT IS KNOWN ALREADY: MPC are frequent and are considered a correctable cause of male infertility. However, they have been poorly investigated in an infertility setting. In addition, no study has investigated clinical and ultrasound (US) characteristics of men with MPC. STUDY DESIGN, SIZE, DURATION: A cross-sectional analysis was carried out of 693 consecutive subjects consulting for couple infertility from September 2012 to March 2017. As a control group, 103 age-matched healthy, fertile men were studied. Furthermore, a longitudinal evaluation of 11 infertile men undergoing trans-rectal ultrasonically-guided cyst aspiration (TRUCA), semen analyses 1 and 3 months after TRUCA and a follow-up 1 year after TRUCA to assess natural pregnancy were performed. PARTICIPANTS/MATERIALS, SETTING, METHODS: All subjects underwent, in our outpatient clinic, clinical, hormonal, scrotal and transrectal US evaluation and semen analysis within the same day. Of 693 males of infertile couples, 648 (37.1 ± 7.9 years, mean+SD) without genetic abnormalities were studied, along with 103 fertile men (36.6 ± 5.0 years). Eleven infertile men underwent TRUCA and were followed-up as reported above. MAIN RESULTS AND THE ROLE OF CHANCE: A MPC was present in 66/648 (10.2%) males of infertile couples and in 6/103 (5.8%) fertile men. MPC occurrence and volume were higher in patients with severe oligo- or azoospermia than in fertile men (all P < 0.05). Infertile men with a MPC showed a lower seminal volume and sperm count and a higher prevalence of azoospermia than the rest of the infertile sample or fertile men, and a higher frequency of US signs suggestive of ejaculatory duct obstruction. MPC volume was negatively associated with total sperm count (r = -0.452, P < 0.0001). In fertile men, the highest MPC volume was 0.117 ml, suggesting it as a biological threshold not compromising semen quality. In infertile men, using receiver operating characteristic curve analyses, a MPC volume >0.117 ml identified subjects with severe oligo- or azoospermia with an overall accuracy of ~75% (both P < 0.005). Eleven men with infertility, semen abnormalities and large MPC (>0.250 ml) underwent TRUCA, which led to sperm count improvement in all patients 1 month after surgery. Three months after TRUCA a lower sperm count and a higher MPC volume than 2 months before were observed (P < 0.005 and P < 0.05, respectively), although improved when compared to baseline. After TRUCA a natural pregnancy occurred in four couples. Finally, we propose an algorithm, based on semen parameters, useful in identifying a MPC in males of infertile couples. LIMITATIONS, REASONS FOR CAUTION: Although in line with the sample size of previous studies (n = 7-20), the number of infertile men with MPC evaluated longitudinally after treatment is limited (n = 11). In addition, although a MPC volume >0.117 ml can negatively affect the sperm count, only MPC > 0.250 ml have been treated in this study. WIDER IMPLICATIONS OF THE FINDINGS: First, the algorithm proposed is easy to use and useful for selecting patients who can benefit from a prostate US in the infertility work-up. Second, a MPC volume ≤0.117 ml may not impair semen quality, while a larger volume can lead to severe oligo- or azoospermia and could be treated. Third, TRUCA is effective, and simpler and less invasive than other surgical techniques for MPC treatment. Finally, since the MPC can increase in size and sperm count decrease over time after TRUCA, semen cryopreservation should be considered 1 month after TRUCA. STUDY FUNDING/COMPETING INTEREST(S): Grants from the Ministry of University and Scientific Research (SIR project to F.L., protocol number: RBSI14LFMQ). No conflicts of interest.

Epididymal more than testicular abnormalities are associated with the occurrence of antisperm antibodies as evaluated by the MAR test.

STUDY QUESTION: Is there any association between mixed antiglobulin reaction (MAR) test positivity and clinical features or genital tract ultrasound (US) parameter values in males of infertile and fertile couples? STUDY ANSWER: In males of infertile and fertile couples MAR test positivity was associated with clinical and US features suggestive of chronic epididymal inflammation. WHAT IS KNOWN ALREADY: MAR test positivity has been more often reported in males of infertile couples than in fertile men. A positive MAR test has been detected in men with a history of testicular or post-testicular damage. No previous study has reported US alterations related to MAR test positivity. This is the first study that has systematically evaluated associations between a positive MAR test and clinical, seminal and US characteristics of the entire male genital tract. STUDY DESIGN, SIZE, DURATION: This cross-sectional analysis included 109 fertile men and 699 consecutive subjects seeking medical care for couple infertility from September 2012 to September 2017. PARTICIPANTS/MATERIALS, SETTING, METHODS: All subjects underwent, in our outpatient clinic, a complete physical, endocrine, scrotal and transrectal US evaluation and semen analysis (including sIL-8) on the same day. Of the 699 males of infertile couples, 181 (age 38.6 ± 6.6 years) had an assessable MAR test, whereas the test was assessable in all 109 fertile men (age 36.6 ± 5.2 years). The associations among MAR test positivity and the other studied parameters were investigated on a caseload of 290 men (patients + fertile men) and in the two cohorts of males of infertile and fertile couples. MAIN RESULTS AND THE ROLE OF CHANCE: Of the 181 men of infertile couples studied, 20 (11%) had a positive MAR test, including 12 (6.6%) who had a MAR test ≥ 50%, which is considered as a pathological threshold according to the WHO. Of the 109 fertile men, four (3.7%) had a positive MAR test, of which one (0.9%) had a MAR test ≥ 50%. MAR test positivity was therefore found more often in men of infertile couples (P < 0.05). In the entire caseload (n = 290) of males of both infertile and fertile couples, no correlations between MAR test positivity and seminal characteristics were observed. A positive MAR test was associated with epididymal US abnormalities, particularly with the mean size of the epididymal body and tail (both P < 0.0001), and in infertile men, a positive MAR test was also associated with an abnormal epididymal echotexture. In addition, subjects with a positive MAR test more frequently showed a history of epididymitis and high sIL-8 levels. Considering endocrine parameters, only a positive correlation between MAR test positivity and LH levels was observed, even after adjusting for age and life-style factors (adj. r = 0.232, P < 0.0001), while no associations with testosterone and FSH levels were found. LIMITATIONS, REASONS FOR CAUTION: Antisperm antibodies (ASA) were detected in this study by using the SpermMAR test IgG, but other tests are available. In addition, for technical reasons, the MAR test is not assessable in subjects with severe oligo-astheno-zoospermia and, therefore, this test may lead to an intrinsic selection bias. Finally, owing to the cross-sectional nature of the study, neither a causality hypothesis nor mechanistic models can be inferred. WIDER IMPLICATIONS OF THE FINDINGS: First, our results indicate that MAR test positivity is associated with clinical and US signs suggestive of chronic epididymal inflammation and not testicular damage. Hence, when investigating a subject with a positive MAR test, the epididymis and not just the testis should be evaluated. Furthermore, MAR test positivity was more often detected in males of infertile couples than in fertile men, but it was not associated with conventional semen parameter values. Our data support a role of ASA in couple infertility, regardless of the conventional sperm analysis. How ASA affects couple fertility needs to be addressed by further studies. STUDY FUNDING/COMPETING INTEREST(S): Grants were received from the Ministry of University and Scientific Research (SIR project to F.L., protocol number: RBSI14LFMQ). There are no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

Contextual fear conditioning modulates the gene expression over time.

Contextual fear conditioning (CFC) is a quick cognitive test based on the association context-aversive stimulus in which a single training leads to a long-term memory. Previously, we showed that 2 days after conditioning the expression of the genes Napa, Pnf2, Casp3, Pdrg1, Ywhaz, Stmn1, Bpgm, were positively modulated in CFC rats respect to naïve rats, explor rats which had freely explored the experimental apparatus and SO rats to which the same number of aversive shocks used in CFC paradigm had been administered in the same CFC apparatus in less time to prevent the association between painful stimuli and apparatus, whereas the genes Actr3, Pea15 and Tiprl were more expressed in SO rats and Cplx1, Trim32 and Ran genes were more expressed in explor rats. At 2 days, Tomm20 gene expression resulted positively modulated in both CFC and explor rats. Herein, we have tested the expression of these genes for a period longer than 2 days, by monitoring the modulation of transcripts within 20 days after conditioning. The expression of the transcripts was assessed by qRT-PCR.We found that three days after CFC only the genes Tiprl and Trim32 were positively modulated in CFC rats whereas the gene Tomm20 was negatively modulated in CFC rats as well as in SO and explor rats. Ten days after CFC, the expression of Trim32 was still positively modulated whereas the genes Tiprl and Tomm20 returned to the constitutive level, and the gene Ran was significantly more expressed in CFC rats than in naïve, SO and explor rats. Interestingly, 20 days after CFC, the genes Stmn1 and Tiprl again became significantly more expressed in CFC rats compared with naïve, SO and explor rats.

Effect of compost application on the dynamics of carbon in a nectarine orchard ecosystem.

The aim of the present study was to compare the quantity and the type of carbon (C) stored during the 14-year lifetime of a commercial nectarine orchard ecosystem fertilized with mineral or organic fertilizers. The study was carried out in the Po valley, Italy, in a nectarine orchard of the variety Stark RedGold, grafted on GF677 hybrid peach × almond. Since orchard planting in August 2001, the following treatments were applied in a randomized complete block design with four replicates per block and compared: 1. unfertilized control; 2. mineral fertilization (including P and K at planting and N applied as NO3NH4 yearly at the rate of 70-130 kg ha-1); 3. compost application at a rate of 5 Mg DW ha-1 yr-1; 4. compost application at a rate of 10 Mg DW ha-1 yr-1. Compost was obtained from domestic organic wastes mixed with pruning material from urban ornamental trees and garden management after a 3-month stabilization period. Application of compost at the highest rate increased C in the soil; the amount of C sequestered was approximately 60% from amendment source and 40% from the net primary production of trees and grasses with a net increase of C compared to mineral fertilization. Compost application was found to be a win-win strategy to increase C storage in soil and, at the same time, to promote plant growth and yield to levels similar to those obtained with mineral fertilization. The rate of C application is crucial, indicated by the fact that compost supply at the rate of 10 Mg ha-1 yr-1 was the only fertilization strategy of the ones tested that resulted in higher C sequestration. This shows that compost amendment may stimulate an increase in the net primary production of plants.

Chromatin Protamination and Catsper Expression in Spermatozoa Predict Clinical Outcomes after Assisted Reproduction Programs.

Identification of parameters predicting assisted reproductive technologies (ARTs) success is a major goal of research in reproduction. Quality of gametes is essential to achieve good quality embryos and increase the success of ARTs. We evaluated two sperm parameters, chromatin maturity and expression of the sperm specific calcium channel CATSPER, in relation to ART outcomes in 206 couples undergoing ARTs. Chromatin maturity was evaluated by Chromomycin A3 (CMA3) for protamination and Aniline Blue (AB) for histone persistence and CATSPER expression by a flow cytometric method. CMA3 positivity and CATSPER expression significantly predicted the attainment of good quality embryos with an OR of 6.6 and 14.3 respectively, whereas AB staining was correlated with fertilization rate. In the subgroup of couples with women ≤35 years, CATSPER also predicted achievement of clinical pregnancy (OR = 4.4). Including CMA3, CATSPER and other parameters affecting ART outcomes (female age, female factor and number of MII oocytes), a model that resulted able to predict good embryo quality with high accuracy was developed. CMA3 staining and CATSPER expression may be considered two applicable tools to predict ART success and useful for couple counseling. This is the first study demonstrating a role of CATSPER expression in embryo development after ARTs programs.

Tumor necrosis factor-alpha levels in blood cord is directly correlated with the body weight of mothers.

BACKGROUND: Obesity has emerged as major public health problem leading to increased morbidity and mortality. Epidemiological studies indicate that in many regions of the world, children and teenagers are increasingly affected by obesity, which contributes for a pessimistic projection for the near future. Maternal obesity has been implicated in metabolic disorders of the offspring, but there are no biological markers that can be detected early on life that predict the development of obesity in the offspring. OBJECTIVE: To evaluate the expression of inflammatory markers in the umbilical cord blood of babies of mothers with obesity/overweight, and correlate these markers with the body weight at age 9 months. METHODS: Anthropometric data of mothers and babies were obtained during prenatal evaluation, at birth and 9 months after birth. Cord blood was collected during delivery of 54 babies from mothers with obesity/overweight and of 50 babies from lean mothers. Tumour necrosis factor-alpha (TNF-α), transforming growth factor 1 beta, monocyte chemoattractant protein-1 and 2 (MCP-1/MCP-2) were determined in serum samples using enzyme-linked immunosorbent assay methods. Correlations were evaluated using the Spearman correlation coefficient, and comparisons were evaluated using the non-parametric Mann-Whitney U-test. RESULTS: Cord blood TNF-α was positively correlated with maternal body mass index. There was an inverse correlation between cord blood transforming growth factor 1 beta and baby body weight at birth. There was no biological marker that predicted body weight at age 9 months. CONCLUSION: Although we have not found a biological marker to predict increased body weight at 9 months of age, the study shows that maternal obesity exposes the baby to higher TNF-α level in the early stages of life, and this can affect metabolic and inflammatory parameters during adulthood.

Is thyroid hormones evaluation of clinical value in the work-up of males of infertile couples?

STUDY QUESTION: Is thyroid hormones (TH) evaluation of clinical value in the work-up of males of infertile couples? STUDY ANSWER: Our results suggest that TH evaluation is not mandatory in the work-up of male infertility. WHAT IS KNOWN ALREADY: A few previous studies performed on a limited series of subjects reported a negative impact of hyper- and hypo-thyroidism on semen volume, sperm concentration, progressive motility and normal morphology. No previous study has systematically evaluated associations between TH variation, semen parameters and ultrasound characteristics of the male genital tract. STUDY DESIGN, SIZE AND DURATION: Cross-sectional analysis of a consecutive series of 172 subjects seeking medical care for couple infertility from September 2010 to November 2014. PARTICIPANTS/MATERIALS, SETTING, METHODS: Of the entire cohort, 163 men (age 38.9 ± 8.0 years) free of genetic abnormalities were studied. All subjects underwent a complete andrological and physical examination, biochemical and hormonal assessment, scrotal and transrectal colour-Doppler ultrasound (CDUS) and semen analysis (including seminal interleukin 8 levels, sIL-8) evaluation within the same day. MAIN RESULTS AND THE ROLE OF CHANCE: Among the patients studied, 145 (88.9%) showed euthyroidism, 6 (3.7%) subclinical hyper- and 12 (7.4%) subclinical hypo-thyroidism. No subjects showed overt hyper- or hypo-thyroidism. At univariate analysis, no associations among thyroid-stimulating hormone (TSH) or TH levels and sperm parameters were observed. Conversely, we observed positive associations among free triiodothyronine (fT3) and free thyroxine (fT4) levels, ejaculate volume and seminal fructose levels. In a multivariate model, after adjusting for confounders such as age, body mass index, smoking habit, sexual abstinence, calculated free testosterone, prolactin and sIL-8 levels, only the associations found for fT3 levels were confirmed. When CDUS features were investigated, using the same multivariate model, we found positive associations between fT3 levels and seminal vesicles (SV) volume, both before and after ejaculation (adj. r = 0.354 and adj. r = 0.318, both P < 0.0001), as well as with SV emptying (ΔSV volume; adj. r = 0.346, P < 0.0001) and echo-texture inhomogeneity. In addition, after adjusting for confounders, negative associations between fT4 levels and epididymal body and tail diameters were found. No significant associations between TSH or TH levels and CDUS features of other organs of the male genital tract, including testis and prostate, were found. Finally, when the features of subjects with euthyroidism, subclinical hypo- and hyper-thyroidism were compared, no significant differences in seminal or hormonal parameters were found. Conversely, evaluating CDUS parameters, subjects with subclinical hyperthyroidism showed a higher difference between the SV longitudinal diameters measured before and after ejaculation when compared with that of subclinical hypothyroid men, even after adjusting for confounders (P < 0.007). All the other male genital tract CDUS characteristics did not differ among groups. LIMITATIONS, REASONS FOR CAUTION: First, the number of patients investigated is relatively small and those with (subclinical) thyroid dysfunctions are an even smaller number; hence, it is therefore difficult to draw firm conclusions. Moreover, the present results are derived from patients consulting an Italian Andrology Clinic for couple infertility, and could have different characteristics from the male general population or from those males consulting general practitioners for reasons other than couple infertility. Finally, due to the cross-sectional nature of the study, neither a causality hypothesis nor mechanistic models can be inferred. WIDER IMPLICATIONS OF THE FINDINGS: Although no associations between TH and sperm parameters were observed, present data support a positive effect of TH on SV size and a permissive role on the ejaculatory machinery, likely through an action on SV and epididymal contractility. This is the first study reporting such evidence. However, in contrast with the view that TH assessment is important for female fertility, our results do not support a systematic evaluation of thyroid function in males of infertile couples. How TH abnormalities impact male fertility needs to be addressed by further studies. STUDY FUNDING/COMPETING INTERESTS: No funding was received for the study. None of the authors have any conflict of interest to declare.