Insights on the freshwater microbiomes metabolic changes associated with the world's largest mining disaster.

To evaluate the impacts of the Fundão tailings dam failure (Minas Gerais, Brazil) on water quality of the Doce River, we analyzed metagenomics and physicochemical parameters during the month of the disaster and again 6 and 10 months after the disaster. To compare dam conditions before and after the failure, we performed a meta-analysis of physicochemical data from a public database. Immediately after the failure, suspended particulate matter (SPM) in the Doce River was 225-1877 mg L-1. Turbidity and dissolved aluminum and iron concentrations were extremely high, whereas dissolved oxygen was below Brazilian legislation norm (<5 mg L-1) in several locations. Six months later, physicochemical values were below thresholds set by Brazilian guidelines (e.g., SPM = 8-166 mg L-1). Short-term impacts on microbial communities included an increase in Actinobacteria and Bacteroidetes and gene sequences related to microbial virulence, motility, respiration, membrane transport, iron and nitrogen metabolism, suggesting changes in microbial metabolic profiles. The 11 recovered partial genomes from metagenomes (MAGs) had genes related to Fe cycle and metal resistance.

Remote sensing, isotopic composition and metagenomics analyses revealed Doce River ore plume reached the southern Abrolhos Bank Reefs.

On November 5th, 2015, the Fundão dam rupture released >50 million m3 of ore tailings into the Doce River, Minas Gerais State, Brazil. The huge volume of mud spread along the river and reached the sea, 17 days after the disaster, in Regência, Espírito Santo State (ES). In 2018, after three years of the disaster, the impacts of the ore tailings in the marine environment are still unclear. This study aims to investigate possible short-term impacts in marine biodiversity caused by the ore tailings' mud over the reef ecosystems that are closest to the disaster area: i.e. recently discovered reefs in the southern Abrolhos Bank. A remote sensing surveillance including winds, sea surface temperature, total suspended material and watercolor (MODIS Aqua data) indicated that the iron tailings plume reached the southern portion of Abrolhos Bank on June 16th, 2016. Subsequently, to obtain further evidence of the presence of the tailings in the coral reefs, water samples were collected in a gradient spanning from the river estuary to the reefs in southern Abrolhos Bank, we also analyzed the isotopic and microbial composition of the samples, as well as the reef benthic composition. Despite no clues of negative impact on benthic (coral) communities, isotopic analysis confirmed the presence of the plume over the reefs area. This study serves as a baseline for future long-term impact assessments of the health of coral reefs in the Abrolhos Bank.

Brugada syndrome trafficking-defective Nav1.5 channels can trap cardiac Kir2.1/2.2 channels.

Cardiac Nav1.5 and Kir2.1-2.3 channels generate Na (INa) and inward rectifier K (IK1) currents, respectively. The functional INa and IK1 interplay is reinforced by the positive and reciprocal modulation between Nav15 and Kir2.1/2.2 channels to strengthen the control of ventricular excitability. Loss-of-function mutations in the SCN5A gene, which encodes Nav1.5 channels, underlie several inherited arrhythmogenic syndromes, including Brugada syndrome (BrS). We investigated whether the presence of BrS-associated mutations alters IK1 density concomitantly with INa density. Results obtained using mouse models of SCN5A haploinsufficiency, and the overexpression of native and mutated Nav1.5 channels in expression systems - rat ventricular cardiomyocytes and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) - demonstrated that endoplasmic reticulum (ER) trafficking-defective Nav1.5 channels significantly decreased IK1, since they did not positively modulate Kir2.1/2.2 channels. Moreover, Golgi trafficking-defective Nav1.5 mutants produced a dominant negative effect on Kir2.1/2.2 and thus an additional IK1 reduction. Moreover, ER trafficking-defective Nav1.5 channels can be partially rescued by Kir2.1/2.2 channels through an unconventional secretory route that involves Golgi reassembly stacking proteins (GRASPs). Therefore, cardiac excitability would be greatly affected in subjects harboring Nav1.5 mutations with Golgi trafficking defects, since these mutants can concomitantly trap Kir2.1/2.2 channels, thus unexpectedly decreasing IK1 in addition to INa.

Understanding patterns of land-cover change in the Brazilian Cerrado from 2000 to 2015.

Clearing tropical vegetation impacts biodiversity, the provision of ecosystem services, and thus ultimately human welfare. We quantified changes in land cover from 2000 to 2015 across the Cerrado biome of northern Minas Gerais state, Brazil. We assessed the potential biophysical and socio-economic drivers of the loss of Cerrado, natural regeneration and net cover change at the municipality level. Further, we evaluated correlations between these land change variables and indicators of human welfare. We detected extensive land-cover changes in the study area, with the conversion of 23 446 km(2) and the natural regeneration of 13 926 km(2), resulting in a net loss of 9520 km(2) The annual net loss (-1.2% per year) of the cover of Cerrado is higher than that reported for the whole biome in similar periods. We argue that environmental and economic variables interact to underpin rates of conversion of Cerrado, most severely affecting more humid Cerrado lowlands. While rates of Cerrado regeneration are important for conservation strategies of the remaining biome, their integrity must be investigated given the likelihood of encroachment. Given the high frequency of land abandonment in tropical regions, secondary vegetation is fundamental to maintain biodiversity and ecosystem services. Finally, the impacts of Cerrado conversion on human welfare likely vary from local to regional scales, making it difficult to elaborate land-use policies based solely on socio-economic indicators.This article is part of the themed issue 'Tropical grassy biomes: linking ecology, human use and conservation'.

An Integrative Approach Uncovers Biomarkers that Associate with Clinically Relevant Disease Outcomes in Vulvar Carcinoma.

UNLABELLED: Vulvar squamous cell carcinoma (VSCC) is a rare disease that has a high mortality rate (∼40%). However, little is known about its molecular signature. Therefore, an integrated genomics approach, based on comparative genome hybridization (aCGH) and genome-wide expression (GWE) array, was performed to identify driver genes in VSCC. To achieve that, DNA and RNA were extracted from frozen VSCC clinical specimens and examined by aCGH and GWE array, respectively. On the basis of the integration of data using the CONEXIC algorithm, PLXDC2 and GNB3 were validated by RT-qPCR. The expression of these genes was then analyzed by IHC in a large set of formalin-fixed paraffin-embedded specimens. These analyses identified 47 putative drivers, 46 of which were characterized by copy number gains that were concomitant with overexpression and one with a copy number loss and downregulation. Two of these genes, PLXDC2 and GNB3, were selected for further validation: PLXDC2 was downregulated and GNB3 was overexpressed compared with non-neoplastic tissue. By IHC, both proteins were ubiquitously expressed throughout vulvar tissue. High expression of GNB3 and low PLXDC2 immunostaining in the same sample was significantly associated with less lymph node metastasis and greater disease-free survival. On the basis of a robust methodology never used before for VSCC evaluation, two novel prognostic markers in vulvar cancer are identified: one with favorable prognosis (GNB3) and the other with unfavorable prognosis (PLXDC2). IMPLICATIONS: This genomics study reveals markers that associate with prognosis and may provide guidance for better treatment in vulvar cancer. Mol Cancer Res; 14(8); 720-9. ©2016 AACR.

Expression of stem cell-related genes in the endometrium and endometriotic lesions: a pilot study.

OBJECTIVE: To compare the expression of stem cell-related genes in the endometrium (END), superficial endometriosis (SE), and deep infiltrating endometriosis (DIE). STUDY DESIGN: We performed a prospective pilot study of six women suffering from SE and DIE who gave consent for laparoscopy surgery, endometrial biopsies, and participation in this study. Quantitative RT-PCR analysis of 84 stem cell-related genes was performed in 18 biopsy samples. RESULTS: A total of 40 of 84 genes were expressed in SE and DIE, but were different from END as follows. Seven genes were over-expressed in SE and 33 genes were under-expressed in DIE compared with END. Two genes were only over-expressed in SE and three genes were only over-expressed in DIE. Six under-expressed genes were exclusively located in SE and one was only located in DIE. The remaining 31 genes were not different among the groups. There was no significant difference in gene expression between SE and DIE samples. CONCLUSION: Tissue of DIE and SE appears to have similar stem cell-related genes. Nevertheless, there are differences in gene expression between SE and DIE.

Study of two strategies to induce follicular wave emergence for assisted reproductive treatments (ART)-a preliminary trial.

PURPOSE: This study aimed to induce follicular wave emergence (FWE) using pharmacological (recombinant hCG administration) or mechanical (aspiration of dominant follicle) interventions in infertile women. METHODS: Sixteen infertile women (≤35 years) with indications for in vitro fertilization due to tubal and/or male factor infertility were randomized into three groups: control (n = 6), pharmacological (n = 5) and mechanical (n = 5) groups. Women in both experimental groups underwent serial transvaginal sonograms (TVS) from menstrual cycle day 10 until identification of a dominant follicle ≥15 mm. Women in the pharmacological group received 250 µg of recombinant-hCG to induce ovulation, and resumed serial TVS 2 days later. In the mechanical group, dominant and subordinate follicles ≥10 mm were aspirated, and daily TVS was resumed on the following day. An increased pool of follicles ≥5 and ≤9 mm after interventions characterized FWE. Women in the control group underwent ovulation induction (OI) with 150 IU/day of recombinant follicle-stimulating hormone started on menstrual cycle day 3 (D3). OI was started on the day of FWE in the experimental groups. Endometrial asynchrony with development of the embryo was expected in the experimental groups. Therefore, all viable embryos were cryopreserved and transferred in an endometrial-stimulated cycle. RESULTS: The number of follicles ≥5 and ≤9 mm increased after the interventions in both experimental groups (p < .001), indicating induction of FWE. OI outcomes were similar among the groups. CONCLUSIONS: The pharmacological and mechanical interventions are efficient in inducing FWE; outcomes of OI synchronized with FWE should be further investigated.

Optimizing human semen cryopreservation by reducing test vial volume and repetitive test vial sampling.

OBJECTIVE: To investigate optimal test vial (TV) volume, utility and reliability of TVs, intermediate temperature exposure (-88°C to -93°C) before cryostorage, cryostorage in nitrogen vapor (VN2) and liquid nitrogen (LN2), and long-term stability of VN2 cryostorage of human semen. DESIGN: Prospective clinical laboratory study. SETTING: University assisted reproductive technology (ART) laboratory. PATIENT(S): A total of 594 patients undergoing semen analysis and cryopreservation. INTERVENTION(S): Semen analysis, cryopreservation with different intermediate steps and in different volumes (50-1,000 µL), and long-term storage in LN2 or VN2. MAIN OUTCOME MEASURE(S): Optimal TV volume, prediction of cryosurvival (CS) in ART procedure vials (ARTVs) with pre-freeze semen parameters and TV CS, post-thaw motility after two- or three-step semen cryopreservation and cryostorage in VN2 and LN2. RESULT(S): Test vial volume of 50 µL yielded lower CS than other volumes tested. Cryosurvival of 100 µL was similar to that of larger volumes tested. An intermediate temperature exposure (-88°C to -93°C for 20 minutes) during cryopreservation did not affect post-thaw motility. Cryosurvival of TVs and ARTVs from the same ejaculate were similar. Cryosurvival of the first TV in a series of cryopreserved ejaculates was similar to and correlated with that of TVs from different ejaculates within the same patient. Cryosurvival of the first TV was correlated with subsequent ARTVs. Long-term cryostorage in VN2 did not affect CS. CONCLUSION(S): This study provides experimental evidence for use of a single 100 µL TV per patient to predict CS when freezing multiple ejaculates over a short period of time (<10 days). Additionally, semen cryostorage in VN2 provides a stable and safe environment over time.

Matrix metalloproteinases 2 and 9 and e-cadherin expression in the endometrium during the implantation window of infertile women before in vitro fertilization treatment.

OBJECTIVE: To evaluate the expression of endometrial matrix metalloproteinases (MMPs) 2 and 9 and E-cadherin in peri-implantation phase of infertile women who have undergone in vitro fertilization (IVF) cycles. METHODS: This prospective study included 51 patients who underwent endometrial biopsy during the receptive phase in a menstrual cycle prior to IVF treatment. The samples were evaluated by tissue microarray for immunohistochemical study. RESULTS: The expression of MMP-2, MMP-9, and E-cadherin in the endometrium prior to IVF treatment was not associated with pregnancy. There was a decrease in E-cadherin immunodetection, the higher the age of the patients, a negative relationship between E-cadherin and MMP-2, and a positive association between MMP-9 and E-cadherin. CONCLUSIONS: The MMP-2, MMP-9, and E-cadherin are expressed in the endometrium of infertile patients during the receptive phase of the natural menstrual cycle. However, there is no correlation between the expression of these molecules and the clinical IVF outcomes.

ROCK1 as a novel prognostic marker in vulvar cancer.

BACKGROUND: Vulvar carcinoma is an infrequent tumour, accounting for fewer than 3% of all malignant tumours that affect women, but its incidence is rising in the past few decades. In young women, the manifestation of the vulvar carcinoma is often linked to risk factors such as smoking and HPV infection, but most cases develop in women aged over 50 years through poorly understood genetic mechanisms. Rho-associated coiled-coil-containing protein kinase 1 (ROCK1) has been implicated in many cellular processes, but its function in vulvar cancer has never been examined. In this study, we aimed to determine the prognostic value of ROCK1 gene and protein analysis in vulvar squamous cell carcinoma (VSCC). METHODS: ROCK1 expression levels were measured in 16 vulvar tumour samples and adjacent normal tissue by qRT-PCR. Further, 96 VSCC samples were examined by immunohistochemistry (IHC) to confirm the involvement of ROCK1 in the disease. The molecular and pathological results were correlated with the clinical data of the patients. Sixteen fresh VSCC samples were analyzed by array-based comparative genomic hybridization (aCGH). RESULTS: In each pair of samples, ROCK1 levels were higher by qRT-PCR in normal tissue compared with the tumour samples (p = 0.016). By IHC, 100% of invasive front areas of the tumour and 95.8% of central tumour areas were positive for ROCK1. Greater expression of ROCK1 was associated with the absence of lymph node metastasis (p = 0.022) and a lower depth of invasion (p = 0.002). In addition, higher ROCK1 levels correlated with greater recurrence-free survival (p = 0.001). Loss of ROCK1 was independently linked to worse cancer-specific survival (p = 0.0054) by multivariate analysis. This finding was validated by IHC, which demonstrated enhanced protein expression in normal versus tumour tissue (p < 0.001). By aCGH, 42.9% of samples showed a gain in copy number of the ROCK1 gene. CONCLUSIONS: ROCK1 is lower expressed in tumour tissue when compared with adjacent normal vulvar epithelia. In an independent sample set of VSCCs, lower expression levels of ROCK1 correlated with worse survival rates and a poor prognosis. These findings provide important information for the clinical management of vulvar cancer.

The role of the Hoxa10/HOXA10 gene in the etiology of endometriosis and its related infertility: a review.

PURPOSE: endometriosis and its associated infertility have been the object of continuous research for over a century. To understand the molecular mechanisms underlying the disease, it has become necessary to determine the aspects of its etiology that are not explained by the retrograde menstruation theory. This could in turn elucidate how various clinical and surgical treatments might affect the evolution and remission of the disease. METHODS: this review is focused on the most recent clinical and laboratory findings regarding the association of HOXA10 with endometriosis and infertility. RESULT: the homebox (Hox/HOX) proteins are highly conserved transcription factors that determine segmental body identities in multiple species, including humans. Hoxa10/HOXA10 is directly involved in the embryogenesis of the uterus and embryo implantation via regulation of downstream genes. Cyclical endometrial expression of Hoxa10/HOXA10, with a peak of expression occurring during the window of implantation, is observed in the adult in response to estrogen and progesterone. Women with endometriosis do not demonstrate the expected mid-luteal rise of HOXA10 expression, which might partially explain the infertility observed in many of these patients. Recent studies also demonstrated HOXA10 expression in endometriotic foci outside the Müllerian tract. CONCLUSIONS: multiple lines of evidence suggest that the actions of the homeobox A10 (Hoxa10/HOXA10) gene could account for some aspects of endometriosis.

Tests for ovarian reserve: reliability and utility.

PURPOSE OF REVIEW: This review discusses ovarian reserve tests for ovulation induction and their application in determining fertility capacity, and their current applications to assess risk of natural ovarian failure and to estimate ovarian function after cancer treatment. RECENT FINDINGS: The current arsenal of ovarian reserve tests comprises hormonal markers [basal follicle stimulating hormone, estradiol, inhibin-B, antimullerian hormone (AMH)] and ultrasonographic markers [ovarian volume, antral follicle counts (AFCs)]. These markers have limitations in terms of which test(s) should be used to reliably predict ovarian reserve with regard to accuracy, invasiveness, cost, convenience, and utility. Several studies have correlated sonographic AFCs with serum AMH levels for predicting the ovarian response to ovulation induction protocols during assisted reproduction treatments. SUMMARY: Serum AMH levels and AFC are reliable tests for predicting the ovarian response to ovulation induction. However, none of the currently employed tests of ovarian reserve can reliably predict pregnancy after assisted conception. Further, ovarian reserve tests cannot predict the onset of reproductive and hormonal menopause; thus, they should be used with caution for reproductive life-programming counseling. Moreover, there is no evidence to support the use of ovarian reserve tests to estimate the risk of ovarian sufficiency after cancer treatments.

Effects of semen storage and separation techniques on sperm DNA fragmentation.

OBJECTIVE: To determine the effect of semen storage and separation techniques on sperm DNA fragmentation. DESIGN: Controlled clinical study. SETTING: An assisted reproductive technology laboratory. PATIENT(S): Thirty normoozospermic semen samples obtained from patients undergoing infertility evaluation. INTERVENTION(S): One aliquot from each sample was immediately prepared (control) for the sperm chromatin dispersion assay (SCD). Aliquots used to assess storage techniques were treated in the following ways: snap frozen by liquid nitrogen immersion, slow frozen with Tris-yolk buffer and glycerol, kept on ice for 24 hours or maintained at room temperature for 4 and 24 hours. Aliquots used to assess separation techniques were processed by the following methods: washed and centrifuged in media, swim-up from washed sperm pellet, density gradient separation, density gradient followed by swim-up. DNA integrity was then measured by SCD. MAIN OUTCOME MEASURE(S): DNA fragmentation as measured by SCD. RESULT(S): There was no significant difference in fragmentation among the snap frozen, slow frozen, and wet-ice groups. Compared to other storage methods short-term storage at room temperature did not impact DNA fragmentation yet 24 hours storage significantly increased fragmentation. Swim-up, density gradient and density gradient/swim-up had significantly reduced DNA fragmentation levels compared with washed semen. Postincubation, density gradient/swim-up showed the lowest fragmentation levels. CONCLUSION(S): The effect of sperm processing methods on DNA fragmentation should be considered when selecting storage or separation techniques for clinical use.

Human choriogonadotropin prior to controlled ovarian stimulation and in vitro fertilization improves implantation, and pregnancy rates.

PURPOSE: Our purpose was to retrospectively compare controlled ovarian stimulation(COH) in IVF cycles with administration of hCG on the day of menses (D1-hCG) with women not receiving hCG at day 1 of menses (Control). METHODS: Data on maternal age, endocrine profile, amount of rFSH required, embryo characteristics, implantation and pregnancy rates were recorded for comparison between D1-hCG (n = 36) and Control (n = 64). RESULTS: Dose of rFSH required to accomplish COH was significantly lower in D1-hCG. Following ICSI, more top-quality embryos were available for transfer per patient in the D1-hCG and biochemical pregnancy rates per transfer were significantly higher in the D1-hCG. Significantly higher implantation and on-going pregnancy rates per embryo transfer were observed in D1-hCG (64%) compared to Control (41%). CONCLUSIONS: Administration of D1-hCG prior to COH reduces rFSH use and enhances oocyte developmental competence to obtain top quality embryos, and improves implantation and on-going pregnancy rates. At present it is not clear if the benefit is related to producing an embryo that more likely to implant or a more receptive uterus, or merely fortuitous and related to the relatively small power of the study.

Endometrial claudin-4 and leukemia inhibitory factor are associated with assisted reproduction outcome.

BACKGROUND: Claudin-4 (CLDN4) is one of several proteins that act as molecular mediators of embryo implantation. Recently, we examined immunolabeling of leukemia inhibitory factor (LIF) in the endometrial tissue of 52 IVF patients, and found that LIF staining intensity was strongly correlated with successful pregnancy initiation. In the same set of patients, we have now examined endometrial CLDN4 expression, to see how expression intensity may vary with LIF. We examined CLDN4 in the luteal phase of the menstrual cycle, immediately preceding IVF treatment. Our aim was to compare expression of LIF and CLDN4 in the luteal phase, and document these patterns as putative biomarkers for pregnancy. METHODS: Endometrial tissue was collected from women undergoing IVF. Endometrial biopsies were obtained during the luteal phase preceding IVF, and were then used for tissue microarray (TMA) immunolabeling of CLDN4. Previously published LIF expression data were then combined with CLDN4 expression data, to determine CLDN4/LIF expression patterns. Associations between successful pregnancy after IVF and combined CLDN4/LIF expression patterns were evaluated. RESULTS: Four patterns of immunolabeling were observed in the endometrial samples: 16% showed weak CLDN4 and strong LIF (CLDN4-/LIF+); 20% showed strong CLDN4 and strong LIF (LIF+/CLDN4+); 28% showed strong CLDN4 and weak LIF (CLDN4+/LIF-); and 36% showed weak CLDN4 and weak LIF (CLDN4-/LIF-). Successful implantation after IVF was associated with CLDN4-/LIF+(p = 0.003). Patients showing this endometrial CLDN4-/LIF+ immunolabeling were also 6 times more likely to achieve pregnancy than patients with endometrial CLDN4+/LIF- immunolabeling (p = 0.007). CONCLUSION: The combined immunolabeling expression of CLDN4-/LIF+ in endometrial tissue is a potential biomarker for predicting successful pregnancy in IVF candidates.

Fluoxetine treatment for anxiety in women undergoing in vitro fertilization.

OBJECTIVE: To compare anxiety levels experienced during 4 stressful periods of in vitro fertilization (IVF) and treatment outcomes between women taking fluoxetine and a placebo. METHODS: A prospective, randomized, double-blind, placebo-controlled trial of patients allocated to receive either fluoxetine (FLX) or folic acid (FA). Anxiety state was assessed at the beginning of ovarian stimulation (OS), ovum pick-up, embryo transfer, and on the day of the pregnancy test (DPT) using the State-Trait Anxiety Inventory (STAI). RESULTS: Baseline STAI-S and STAI-T were normal. From OS to DPT, STAI-S increased from 42.8+/-10.6 to 44+/-9.0 in the FLX group and from 40.9+/-8.1 to 45.3+/-8.3 in the FA group (P=0.03 and P=0.001, respectively). IVF outcome was not affected by the treatment in the two groups. CONCLUSIONS: Caution is needed in prescribing fluoxetine to alleviate anxiety in patients undergoing IVF. Studies are needed to determine whether other selective serotonin reuptake inhibitors or higher fluoxetine doses can relieve emotional distress without affecting IVF outcome.

Endometrial leukemia inhibitory factor as a predictor of pregnancy after in vitro fertilization.

OBJECTIVE: To determine whether there is an association between endometrial expression of leukemia inhibitory factor (LIF) in the luteal phase of the menstrual cycle preceding in vitro fertilization (IVF) and treatment outcome. METHODS: Biopsy specimens from the endometria of 52 women in the luteal phase were immunostained against LIF. Embryo culture and transfer were done according to standard procedures. RESULTS: Clinical pregnancy occurred in 39% of the women following IVF, and strong endometrial immunohistochemical staining for LIF was associated with pregnancy (P=0.01). The women with a strong LIF expression had a 6.4-fold higher chance of becoming pregnant than those with weaker intensities (P=0.005). CONCLUSION: Endometrial expression of LIF during the luteal phase can be used as a predictor of IVF success.