Measurement method of optical properties of ex vivo biological tissues of rats in the near-infrared range

An optical fiber-based supercontinuum setup and a custom-made spectrophotometer that can measure spectra from 1100 to 2300 nm, are used to describe attenuation properties from different ex vivo rat tissues. Our method is able to differentiate between scattering and absorption coefficients in biological tissues. Theoretical assumptions combined with experimental measurements demonstrate that, in this infrared range, tissue attenuation and absorption can be accurately measured, and scattering can be described as the difference between both magnitudes. Attenuation, absorption, and scattering spectral coefficients of heart, brain, spleen, retina, and kidney are given by applying these theoretical and experimental methods. Light through these tissues is affected by high scattering, resulting in multiple absorption events, and longer wavelengths should be used to obtain lower attenuation values. It can be observed that the absorption coefficient has a similar behavior in the samples under study, with two main zones of absorption due to the water absorption bands at 1450 and 1950 nm, and with different absolute absorption values depending on the constituents of each tissue. The scattering coefficient can be determined, showing slight differences between retina and brain samples, and among heart, spleen and kidney tissues

Phylogeography of Hypostomus strigaticeps (Siluriformes: Loricariidae) inferred by mitochondrial DNA reveals its distribution in the upper Paraná River basin.

In this study, phylogenetic and phylogeographic analyses of populations identified as Hypostomus strigaticeps from the upper Paraná River basin were conducted in order to test whether these different populations comprises cryptic species or structured populations and to assess their genetic variability. The sequences of the mitochondrial DNA ATP sintetase (subunits 6/8) of 27 specimens from 10 populations (one from Mogi-Guaçu River, five from Paranapanema River, three from Tietê River and one from Peixe River) were analyzed. The phylogeographic analysis showed the existence of eight haplotypes (A-H), and despite the ancestral haplotype includes only individuals from the Tietê River basin, the distribution of H. strigaticeps was not restricted to this basin. Haplotypes A, B and F were the most frequent. Haplotypes D, E, F, G, and H were present in the sub-basin of Paranapanema, two (A and B) were present in the sub-basin of the Tietê River, one (C) was exclusively distributed in the sub-basin of the Peixe River, and one (B) was also present in the sub-basin of the Grande River. The phylogenetic analysis showed that the populations of H. strigaticeps indeed form a monophyletic unit comprising two lineages: TG, with representatives from the Tietê, Mogi-Guaçu and Peixe Rivers; and PP, with specimens from the Paranapanema River. The observed degree of genetic divergence within the TG and PP lineages was 0.1% and 0.2%, respectively, whereas the genetic divergence between the two lineages themselves was approximately 1%. The results of the phylogenetic analysis do not support the hypothesis of existence of crypt species and the phylogeographic analysis confirm the presence of H. strigaticeps in other sub-basins of the upper Paraná River: Grande, Peixe, and Paranapanema sub-basins

Microperimetry and Optical Coherence Tomography Changes in Type-1 Diabetes Mellitus without Retinopathy

Background: We aimed to measure and correlate inner retinal layer (IRL) thickness and macular sensitivity by optical coherence tomography (OCT) and by microperimetry, respectively, in type 1 diabetes mellitus patients (DM1) without diabetic retinopathy (DR). Methods: Fifty-one DM1 patients and 81 age-matched healthy subjects underwent measurement of the axial length (AL), retinal thickness in the macular ETDRS areas by swept source (SS)-OCT and macular sensitivity by microperimeter. Results: The total retinal and IRL thicknesses were thicker in the DM1 group (p < 0.05) in practically all ETDRS areas, and they had a generalized decrease in sensitivity (p < 0.05) in 9 areas between both groups. There was a significant negative correlation between retinal sensitivity and age in all areas and in visual acuity (VA) in 5 out of the 9 areas for DM1 patients. Only a mild negative correlation was observed between retinal sensitivity in the 5 degrees nasal inner (5NI) area and in IRL thickness in the temporal inner (TI) area (-0.309 with p = 0.029) in the DM1 group. Conclusion: Aging and disease evolution in DM1 patients without DR signs generate a decrease in retinal sensitivity. There was a direct relationship between retinal sensitivity and macular thickness in the DM1 group

Origin and history of Phoxinus (Cyprinidae) introductions in the Douro basin (Iberian Peninsula): an update inferred from genetic data

The number of non-native freshwater fishes in the Iberian Peninsula has been greatly increasing. In this study, individuals of the genus Phoxinus were detected in 18 out of 138 stream sites sampled across the Douro Basin in 2017 and 2018. A total of 26 individuals were barcoded using partial cytochrome c oxidase subunit I (COI) and cytochrome b (cytb) genes for species identification and determination of geographical origin. Molecular data provided the first record of a second Phoxinus species in western Douro (Portugal, Iberian Peninsula), with haplotypes closely matching those found in the Charente River (southern France). This species is suspected to be a recent introduction associated with the use of minnows as live bait by freshwater anglers, which was facilitated by human movements between France and Portugal. Individuals from watercourses in eastern Douro (Spain) were genetically assigned to Phoxinus bigerri, an introduced species previously known for that region, which confirms reports of introduction events from Ebro to Douro Basin probably also related to freshwater angling and facilitated by geographic proximity. The potential ecological impacts of this genus in the region are unknown and need further investigation.We acknowledge Fernando Teixeira, Fernando Miranda, Mario Ferreira, Sara Carona, Jose Pedro RamiAo and Francisco Carvalho for the valuable assistance during fieldwork. We specially thank Maria Filomena MagalhAes for previous fruitful discussions and logistic support. We are grateful to Matthias F. Geiger and Andrea Corral Lou for facilitating genetic data and coordinates of sampling sites. Finally, we appreciate the comments of the three anonymous reviewers that improved the quality of the manuscript. AFF and AGR were supported by the project FRESHING founded by the Portuguese Foundation for Science and Technology (FCT) and COMPETE (PTDC/AAGMAA/2261/2014 - POCI-01-0145-FEDER-356016824). FMSM was supported by the FCT PhD grant SFRH/BD/104703/2014. This study was conducted as part of the projects FRESHING and FRESHCO. The latter is also supported by FCT and COMPETE (PTDC/AGR-FOR/1627/2014 - 04/SAICT/2015) and UID/AGR/04033/2019. Logistic support was also facilitated by the ENVMETAGEN - Capacity Building at InBIO for Research and Innovation Using Environmental Metagenomics project at CIBIO laboratories (668981; EUH2020-WIDESPREAD-2014-2)

Deregulation of miR-324/KISS1/kisspeptin in early ectopic pregnancy: mechanistic findings with clinical and diagnostic implications

[Abstract] BACKGROUND: Ectopic pregnancy is a life-threatening condition for which novel screening tools that would enable early accurate diagnosis would improve clinical outcomes. Kisspeptins, encoded by KISS1, play an essential role in human reproduction, at least partially by regulating placental function and possibly embryo implantation. Kisspeptin levels are elevated massively in normal pregnancy and reportedly altered in various gestational pathologic diseases. Yet, the pathophysiologic role of KISS1/kisspeptin in ectopic pregnancy has not been investigated previously. OBJECTIVE: The purpose of this study was to evaluate changes of KISS1/kisspeptin levels in ectopic pregnancy and their underlaying molecular mechanisms and to ascertain the diagnostic implications of these changes. STUDY DESIGN: A total of 122 women with normal pregnancy who underwent voluntary termination of pregnancy and 84 patients who experienced tubal ectopic pregnancy were recruited. Measurements of plasma kisspeptins and KISS1 expression analyses in human embryonic/placental tissue were conducted in ectopic pregnancy and voluntary termination of pregnancy control subjects during the early gestational window (<12 weeks). Putative microRNA regulators of KISS1 were predicted in silico, followed by expression analyses of selected microRNAs and validation of repressive interactions in vitro. Circulating levels of these microRNAs were also assayed in ectopic pregnancy vs voluntary termination of pregnancy. RESULTS: Circulating kisspeptins gradually increased during the first trimester of normal pregnancy but were reduced markedly in ectopic pregnancy. This profile correlated with the expression levels of KISS1 in human embryonic/placental tissue, which increased in voluntary termination of pregnancy but remained suppressed in ectopic pregnancy. Bioinformatic predictions and expression analyses identified miR-27b-3p and miR-324-3p as putative repressors of KISS1 in human embryonic/placental tissue at <12 weeks gestation, when expression of microRNAs was low in voluntary termination of pregnancy control subjects but significantly increased in ectopic pregnancy. Yet, a significant repressive interaction was documented only for miR-324-3p, occurring at the predicted 3'-UTR of KISS1. Interestingly, circulating levels of miR-324-3p, but not of miR-27b-3p, were suppressed distinctly in ectopic pregnancy, despite elevated tissue expression of the pre-microRNA. A decision-tree model that used kisspeptin and miR-324-3p levels was successful in discriminating ectopic pregnancy vs voluntary termination of pregnancy, with a receiver-operating characteristic area under the curve of 0.95±0.02 (95% confidence interval). CONCLUSION: Our results document a significant down-regulation of KISS1/kisspeptins in early stages of ectopic pregnancy via, at least partially, a repressive interaction with miR-324-3p. Our data identify circulating kisspeptins and miR-324-3p as putative biomarkers for accurate screening of ectopic pregnancy at early gestational ages.Ministerio de E$conomía y Competitividad (España); BFU2014-57581-PMinisterio de Economía y Competitividad ; BFU2017-83934-PInstituto de Salud Carlos III; PIE-00005Junta de Andalucía; P08-CVI-03788Junta de Andalucía; P12-FQM-0194

Hybridization and invasive species in a threatened freshwater fish community under environmental pressures: Morphometric and molecular evidence

Mediterranean freshwater systems are under threat owing to increased drought driven by climate change, intensive human land uses and non-native species. This is causing increased fish hybridization in isolated watercourses. The genetic and morphological characteristics of hybrids of sympatric native and non-native fish species were studied in four streams of the Mediterranean Guadalquivir basin (south-west Spain). Fish morphology was analysed using geometric morphometrics, and molecular determination of parenthood was inferred through one mitochondrial gene (cytb) and one nuclear gene (Beta-actin) for all hybrids and a subset of pure parental specimens. Molecular analyses confirmed hybrids between the native Squalius alburnoides and non-native Alburnus alburnus in a stream with continuous flow. Haplotype analyses suggested that they originated from backcrossing of hybrid offspring. Intergeneric crosses between native species S. alburnoides and Pseudochondrostoma willkommii, and S. alburnoides and Iberochondrostoma lemmingii were detected in streams under reduced connectivity scenarios. Morphometrics revealed that hybrid phenotypes were similar to S. alburnoides. In some cases, molecular markers uncovered hybridization events that were neither detected in the field nor by morphometric analyses, potentially supporting a backcrossing/introgression scenario. Hybridization is likely to be increasing in Mediterranean rivers where S. alburnoides are present owing to increased fragmentation caused by summer drought exacerbated by climate change and human land uses and pressures. This can become a problem for these endemic vulnerable species if genetic diversity is lost, morphological homogenization occurs and hybrids cannot be easily detected in the field. The potential risks could be addressed by monitoring and eradication of non-native species and segregation from natives. To avoid native–native crosses, habitat quality and desiccation risk could be tackled by improved water quality and riparian reforestation to provide shade and reduce evapotranspiration. This would need increased coordination and intervention between the institutions that share conservation responsibilities in the area

Selective loss of kisspeptin signaling in oocytes causes progressive premature ovulatory failure

Study question: Does direct kisspeptin signaling in the oocyte have a role in the control of follicular dynamics and ovulation?Summary answer: Kisspeptin signaling in the oocyte plays a relevant physiological role in the direct control of ovulation; oocyte-specific ablation of kisspeptin receptor, Gpr54, induces a state of premature ovulatory failure in mice that recapitulates some features of premature ovarian insufficiency (POI).What is known already: Kisspeptins, encoded by the Kiss1 gene, are essential for the control of ovulation and fertility, acting primarily on hypothalamic GnRH neurons to stimulate gonadotropin secretion. However, kisspeptins and their receptor, Gpr54, are also expressed in the ovary of different mammalian species, including humans, where their physiological roles remain contentious and poorly characterized.Study design, size, duration: A novel mouse line with conditional ablation of Gpr54 in oocytes, named OoGpr54-/-, was generated and studied in terms of follicular and ovulatory dynamics at different age-points of postnatal maturation. A total of 59 OoGpr54-/- mice and 47 corresponding controls were analyzed. In addition, direct RNA sequencing was applied to ovarian samples from 8 OoGpr54-/- and 7 control mice at 6 months of age, and gonadotropin priming for ovulatory induction was conducted in mice (N = 7) from both genotypes.Participants/materials, setting, methods: Oocyte-selective ablation of Gpr54 in the oocyte was achieved in vivo by crossing a Gdf9-driven Cre-expressing transgenic mouse line with a Gpr54 LoxP mouse line. The resulting OoGpr54-/- mouse line was subjected to phenotypic, histological, hormonal and molecular analyses at different age-points of postnatal maturation (Day 45, and 2, 4, 6 and 10-11 months of age), in order to characterize the timing of puberty, ovarian follicular dynamics and ovulation, with particular attention to identification of features reminiscent of POI. The molecular signature of ovaries from OoGpr54-/- mice was defined by direct RNA sequencing. Ovulatory responses to gonadotropin priming were also assessed in OoGpr54-/- mice.Main results and the role of chance: Oocyte-specific ablation of Gpr54 caused premature ovulatory failure, with some POI-like features. OoGpr54-/- mice had preserved puberty onset, without signs of hypogonadism. However, already at 2 months of age, 40% of OoGpr54-/- females showed histological features reminiscent of ovarian failure and anovulation. Penetrance of the phenotype progressed with age, with >80% and 100% of OoGpr54-/- females displaying complete ovulatory failure by 6- and 10 months, respectively. This occurred despite unaltered hypothalamic Gpr54 expression and gonadotropin levels. Yet, OoGpr54-/- mice had decreased sex steroid levels. While the RNA signature of OoGpr54-/- ovaries was dominated by the anovulatory state, oocyte-specific ablation of Gpr54 significantly up- or downregulated of a set of 21 genes, including those encoding pituitary adenylate cyclase-activating polypeptide, Wnt-10B, matrix-metalloprotease-12, vitamin A-related factors and calcium-activated chloride channel-2, which might contribute to the POI-like state. Notably, the anovulatory state of young OoGpr54-/- mice could be rescued by gonadotropin priming.Large scale data: N/A. .Limitations, reasons for caution: Conditional ablation of Gpr54 in oocytes unambiguously caused premature ovulatory failure in mice; yet, the ultimate molecular mechanisms for such state of POI can be only inferred on the basis of RNAseq data and need further elucidation, since some of the molecular changes observed in OoGpr54-/- ovaries were secondary to the anovulatory state. Direct translation of mouse findings to human disease should be made with caution since, despite the conserved expression of Kiss1/kisspeptin and Gpr54 in rodents and humans, our mouse model does not recapitulate all features of common forms of POI.Wider implications of the findings: Deregulation of kisspeptin signaling in the oocyte might be an underlying, and previously unnoticed, cause for some forms of POI in women.Study funding/competing interest(s): This work was primarily supported by a grant to M.P. and M.T.-S. from the FiDiPro (Finnish Distinguished Professor) Program of the Academy of Finland. Additional financial support came from grant BFU2017-83934-P (M.T.-S.; Ministerio de Economía y Competitividad, Spain; co-funded with EU funds/FEDER Program), research funds from the IVIRMA International Award in Reproductive Medicine (M.T.-S.), and EFSD Albert Renold Fellowship Programme (S.T.R.). The authors have no conflicts of interest to declare in relation to the contents of this work.</p

Guide to the alien and invasive species of rivers, lakes and estuaries in the Iberian Peninsula.

Guide to the alien and invasive species of rivers, lakes and estuaries in the Iberian Peninsula