HIV-1 Resistance to Islatravir/Tenofovir Combination Therapy in Wild-Type or NRTI-Resistant Strains of Diverse HIV-1 Subtypes.

Tenofovir disoproxil fumarate (TDF) and islatravir (ISL, 4'-ethynyl-2-fluoro-2'-deoxyadensine, or MK-8591) are highly potent nucleoside reverse transcriptase inhibitors. Resistance to TDF and ISL is conferred by K65R and M184V, respectively. Furthermore, K65R and M184V increase sensitivity to ISL and TDF, respectively. Therefore, these two nucleoside analogs have opposing resistance profiles and could present a high genetic barrier to resistance. To explore resistance to TDF and ISL in combination, we performed passaging experiments with HIV-1 WT, K65R, or M184V in the presence of ISL and TDF. We identified K65R, M184V, and S68G/N mutations. The mutant most resistant to ISL was S68N/M184V, yet it remained susceptible to TDF. To further confirm our cellular findings, we implemented an endogenous reverse transcriptase assay to verify in vitro potency. To better understand the impact of these resistance mutations in the context of global infection, we determined potency of ISL and TDF against HIV subtypes A, B, C, D, and circulating recombinant forms (CRF) 01_AE and 02_AG with and without resistance mutations. In all isolates studied, we found K65R imparted hypersensitivity to ISL whereas M184V conferred resistance. We demonstrated that the S68G polymorphism can enhance fitness of drug-resistant mutants in some genetic backgrounds. Collectively, the data suggest that the opposing resistance profiles of ISL and TDF suggest that a combination of the two drugs could be a promising drug regimen for the treatment of patients infected with any HIV-1 subtype, including those who have failed 3TC/FTC-based therapies.

An experimental evaluation of the efficacy of perinatal sulforaphane supplementation to decrease the incidence and severity of vinclozolin-induced hypospadias in the mouse model.

Determining the mechanisms of toxicity induced by pollutants has long been a research priority in lieu of considering the mechanisms of resilience that prevent deleterious impacts. Protective mechanisms in many taxa can be therapeutically targeted to enhance resilience to synthetic toxicants. For example, the environmental sensor, Nuclear factor (erythroid-derived 2)-like 2 (Nfe2l2 or Nrf2), a transcription factor, facilitates transcription of many protective genes. Hypospadias is a common malformation of the penis. The risk of being born with hypospadias increases with pollutant exposure. We use vinclozolin-induced hypospadias in the mouse as a model to test the hypothesis that pollutant-induced birth defects can be prevented and reduced in severity by augmenting natural mechanisms of resilience. Pregnant mice were exposed to the demasculinizing toxicant, vinclozolin, in combination with increasing doses of the NRF2 activator, sulforaphane. The sulforaphane dose that most effectively increased masculinization (anogenital distance) was identified and used to test the hypothesis that sulforaphane reduces the hypospadias-inducing potency of vinclozolin. Finally, a Nrf2 knockout study was conducted to test whether NRF2 was required for the sulforaphane-induced rescue effects. Sulforaphane supplementation to vinclozolin exposed embryos increased anogenital distance in a nonlinear fashion typical of Nrf2 activators. The most effective dose of sulforaphane (45 mg/kg) reduced the occurrence and severity of vinclozolin-induced hypospadias and corrected penis morphogenesis. The sulforaphane-induced rescue effect was dependent on the presence of Nrf2. Nrf2 plays a critical role in protecting the fetus from vinclozolin and reduces the incidence and severity of hypospadias, the most common birth defect in boys in many countries. This work lays a foundation for developing prenatal supplements that will protect the fetus from pollutant-induced hypospadias. Studying the protective mechanisms that drive resilience to toxicants will facilitate innovation of protective therapies.

Somatic cell fate maintenance in mouse fetal testes via autocrine/paracrine action of AMH and activin B.

Fate determination and maintenance of fetal testes in most mammals occur cell autonomously as a result of the action of key transcription factors in Sertoli cells. However, the cases of freemartin, where an XX twin develops testis structures under the influence of an XY twin, imply that hormonal factor(s) from the XY embryo contribute to sex reversal of the XX twin. Here we show that in mouse XY embryos, Sertoli cell-derived anti-Mullerian hormone (AMH) and activin B together maintain Sertoli cell identity. Sertoli cells in the gonadal poles of XY embryos lacking both AMH and activin B transdifferentiate into their female counterpart granulosa cells, leading to ovotestis formation. The ovotestes remain to adulthood and produce both sperm and oocytes, although there are few of the former and the latter fail to mature. Finally, the ability of XY mice to masculinize ovaries is lost in the absence of these two factors. These results provide insight into fate maintenance of fetal testes through the action of putative freemartin factors.

One Tool for Many Jobs: Divergent and Conserved Actions of Androgen Signaling in Male Internal Reproductive Tract and External Genitalia.

In the 1940s, Alfred Jost demonstrated the necessity of testicular secretions, particularly androgens, for male internal and external genitalia differentiation. Since then, our knowledge of androgen impacts on differentiation of the male internal (Wolffian duct) and external genitalia (penis) has been drastically expanded upon. Between these two morphologically and functionally distinct organs, divergent signals facilitate the establishment of tissue-specific identities. Conversely, conserved actions of androgen signaling are present in both tissues and are largely responsible for the growth and expansion of the organs. In this review we synthesize the existing knowledge of the cell type-specific, organ specific, and conserved signaling mechanisms of androgens. Mechanistic studies on androgen signaling in the Wolffian duct and male external genitalia have largely been conducted in mouse model organisms. Therefore, the majority of the review is focused on mouse model studies.

Thromboelastometry demonstrates endogenous coagulation activation in nonsevere and severe COVID-19 patients and has applicability as a decision algorithm for intervention.

In patients with severe forms of COVID-19, thromboelastometry has been reported to display a hypercoagulant pattern. However, an algorithm to differentiate severe COVID-19 patients from nonsevere patients and healthy controls based on thromboelastometry parameters has not been developed. Forty-one patients over 18 years of age with positive qRT-PCR for SARS-CoV-2 were classified according to the severity of the disease: nonsevere (NS, n = 20) or severe (S, n = 21). A healthy control (HC, n = 9) group was also examined. Blood samples from all participants were tested by extrinsic (EXTEM), intrinsic (INTEM), non-activated (NATEM) and functional assessment of fibrinogen (FIBTEM) assays of thromboelastometry. The thrombodynamic potential index (TPI) was also calculated. Severe COVID-19 patients exhibited a thromboelastometry profile with clear hypercoagulability, which was significantly different from the NS and HC groups. Nonsevere COVID-19 cases showed a trend to thrombotic pole. The NATEM test suggested that nonsevere and severe COVID-19 patients presented endogenous coagulation activation (reduced clotting time and clot formation time). TPI data were significantly different between the NS and S groups. The maximum clot firmness profile obtained by FIBTEM showed moderate/elevated accuracy to differentiate severe patients from NS and HC. A decision tree algorithm based on the FIBTEM-MCF profile was proposed to differentiate S from HC and NS. Thromboelastometric parameters are a useful tool to differentiate the coagulation profile of nonsevere and severe COVID-19 patients for therapeutic intervention purposes.

Characteristics and Outcomes of Heart Transplant Recipients With Coronavirus-19 Disease in a High-volume Transplant Center.

BACKGROUND: Heart transplant (HT) recipients may be at higher risk of acquiring SARS-CoV-2 infection and developing critical illness. The aim of this study is to describe characteristics and outcomes of HT recipients infected by SARS-COV-2, from a high-volume transplant center. METHODS: We have described data of all adult HT recipients with confirmed coronavirus disease 2019 by RT-PCR in nasopharyngeal samples from April 5, 2020, to January 5, 2021. Outcomes and follow-up were recorded until February 5, 2021. RESULTS: Forty patients were included. Twenty-four patients (60%) were men; the median age was 53 (40-60) y old; median HT time was 34 mo; and median follow-up time 162 d. The majority needed hospitalization (83%). Immunosuppressive therapy was reduced/withdrawn in the majority of patients, except from steroids, which were maintained. Seventeen patients (42.5%) were classified as having severe disease according to the ordinal scale developed by the World Health Organization Committee. They tended to have lower absolute lymphocyte count (P < 0.001) during follow-up when compared with patients with mild disease. Thirty-day mortality was 12.5%. However, a longer follow-up revealed increased later mortality (27.5%), with median time to death around 35 d. Bacterial nosocomial infections were a leading cause of death. Cardiac allograft rejection (10%) and ventricular dysfunction (12.5%) were also not negligible. CONCLUSIONS: Major findings of this study corroborate other cohorts' results, but it also reports significant rate of later events, suggesting that a strict midterm surveillance is advisable to HT recipients with coronavirus disease 2019.

Defective Allele of the Neuronal Nitric Oxide Synthase Gene Increases Insulin Resistance During Acute Phase of Myocardial Infarction.

BACKGROUND: Glycemic disorders are strong predictors of mortality in ST-elevation myocardial infarction (STEMI) patients, and disruption in nitric oxide (NO) production is associated with insulin-resistant states. We evaluated whether a defective allele of the neuronal nitric oxide synthase (nNOS) gene (NOS1) might influence insulin response and blood-glucose balance during the acute phase of STEMI and if post-infarction total plasma-NO levels and vasodilation scores varied across nNOS genotypes. METHODS: Consecutive patients with STEMI (n=354) underwent clinical evaluations and genotyping for the promoter variation rs41279104. In-hospital clinical and blood evaluations were performed at admission and five days after STEMI, with glycemic, insulinemic, and disposition indices assessed at the same times. Flow-mediated dilation (FMD) was assessed by reactive hyperemia on the 30th day. RESULTS: Homozygotes for the defective allele (A) showed lower glycemia and insulin sensitivity on day 1 while showing the highest ß-cell function and no changes in the circulating NO pool, which is compatible with hyperresponsive ß cells counteracting the inherent glucose-resistant state of AA patients. At day 5, glycemic scores had shifted to indicate greater insulin sensitivity among A homozygotes, paralleled by a significant yet poor increase in NO bioavailability compared to that among G carriers. All in all, defective homozygotes showed greater insulin resistance at admission that had reversed by 5 days after STEMI. Even so, A carriers developed lower FMD scores compared to G homozygotes after the acute phase. CONCLUSION: A defective nNOS allele (and due decline in NO production) seemed to elicit a hyperinsulinemia response to compensate for an insulin-resistant state during the acute phase of STEMI and to be associated with poor endothelial function after the acute phase.

Toxicant exposure during pregnancy increases protective proteins in the dam and a sexually dimorphic response in the fetus.

Endocrine disrupting compounds (EDCs) are ubiquitous environmental pollutants that alter endocrine system function, induce birth defects, and a myriad of other negative health outcomes. Although the mechanism of toxicity of many EDCs have been studied in detail, little work has focused on understanding the mechanisms through which pregnant dams and fetuses protect themselves from EDCs, or if those protective mechanisms are sexually dimorphic in fetuses. In this study, we examined proteomic alterations in the livers of mouse dams and their male and female fetuses induced by vinclozolin, a model antiandrogenic EDC. Dam livers upregulated nine phase I and phase II detoxification pathways and pathway analysis revealed that more pathways are significantly enriched in dam livers than in fetal livers. Phase I and II detoxification proteins are also involved in steroid and steroid hormone biosynthesis and vinclozolin likely alters steroid levels in both the dam and the fetus. The response of the fetal liver proteome to vinclozolin exposure is sexually dimorphic. Female fetal livers upregulated proteins in xenobiotic metabolism pathways, whereas male fetal livers upregulated proteins in oxidative phosphorylation pathways. These results suggest that female fetuses increase protective mechanisms, whereas male fetuses increase ATP production and several disease pathways that are indicative of oxidative damage. Females fetuses upregulate proteins and protective pathways that were similar to the dams whereas males did not. If this sexually dimorphic pattern is typical, then males might generally be more sensitive to EDCs.

Impacto socioeconómico del manejo actual de la pancreatitis aguda de origen biliar severa: estudio comparativo / Socioeconomic impact of the current management of severe biliary acute pancreatitis: comparative study

RESUMEN Objetivo La pancreatitis aguda de origen biliar es una patología gastrointestinal común, en donde el tratamiento oportuno es el pilar más importante a pesar de sus discrepancias. El objetivo del estudio es establecer el impacto socioeconómico en el manejo actual de esta patología, comparando dos hospitales de tercer nivel de estrato socioeconómico alto y bajo de la ciudad de Bogotá, Colombia. Materiales y Métodos Se realizó un estudio retrospectivo, comparativo de corte transversal entre enero de 2012 y diciembre de 2017, en dos hospitales de Bogotá D. C. Se evaluaron sus características socioeconómicas, género, tiempo de evolución al momento de la consulta, Marshall score, estancia en UCI, estancia hospitalaria, complicaciones, manejo quirúrgico y mortalidad. Resultados Se analizaron 101 pacientes de dos estratos socioeconómicos diferentes (alto y bajo). Se encontró que los pacientes de estrato bajo tienen un riesgo diez veces mayor de requerir un procedimiento quirúrgico. Asimismo, registraron una mayor mortalidad en comparación con pacientes de estrato alto (11,3% vs. 4,2%). También se evidenciaron más complicaciones en el grupo de nivel socioeconómico bajo con respecto al alto, como en la falla exocrina (81,1% vs. 31,3%) y el síndrome compartimental (35,8% vs. 4,2%). Conclusión Se encuentra mayor morbimortalidad en los pacientes de bajo nivel socioeconómico en el contexto de esta patología. Este estudio puede guiar a nuevas investigaciones acerca del impacto socioeconómico en los desenlaces de pancreatitis aguda severa.(AU)

ABSTRACT Objetive Acute pancreatitis of biliary origin is a common gastrointestinal pathology, in which timely management still is the most important. The aims of this research is establish the socioeconomic impact in the current management of severe acute pancreatitis of biliary origin comparing two centers of the third level, one of high socioeconomic population and another of low in Bogotá, Colombia. Materials and Methods A retrospective, cross-sectional comparative study was conducted between January 2012 and December 2017, in two hospitals of Bogotá DC. We evaluated their socioeconomic characteristics, gender, time of evolution at the time of consultation, Marshall score, ICU stay, hospital stay, complications, surgical management and mortality. Results 101 patients from two different socioeconomic strata (high and low) were analyzed, where a 10 times higher risk of requiring a surgical procedure in the group of patients with low stratum was found, as well as a higher mortality compared with those of high stratum. (11.3% Vs 4.2%). There were also more complications in the low socioeconomic group with respect to the high, as in the exocrine failure (81.1% vs 31.3%) and the compartment syndrome (35.8% vs 4.2%). Conclusion There is greater morbidity and mortality in patients of low socioeconomic status in the context of this pathology. This study can guide new research that increases the clarity of the socioeconomic impact on the outcomes of severe acute pancreatitis.(AU)

Equine chorionic gonadotropin increases estradiol levels in the bovine oviduct and drives the transcription of genes related to fertilization in superstimulated cows.

In the bovine oviduct, estradiol (E2) stimulates secretion and cell proliferation, whereas progesterone (P4) suppresses them. In this study, we have evaluated the effect of two superstimulatory protocols (follicle-stimulating hormone [FSH] or FSH combined with equine chorionic gonadotropin [eCG]) on the oviductal levels of E2 and P4 and its outcome on oviductal cells. Compared with the control group (a single pre-ovulatory follicle), we have observed that the cows submitted to FSH/eCG treatment showed a higher concentration of E2 in the oviduct tissue, together with a higher abundance of messenger RNA encoding steroid receptors (ESR1 and progesterone receptor), and genes linked to gamete interactions and regulation of polyspermy (oviduct-specific glycoprotein 1, heat-shock protein family A member 5, α-l-fucosidase 1 [FUCA1], and FUCA2) in the infundibulum and ampulla segments of the oviduct. However, we did not observe any modulation of gene expression in the isthmus segment. Even though the FSH protocol upregulated some of the genes analyzed, we may infer that the steady effect of FSH combined with eCG on oviduct regulation might benefit fertilization and may potentially increase pregnancy rates.

Inflammasome gene expression is associated with immunopathology in human localized cutaneous leishmaniasis.

Localized cutaneous leishmaniasis (LCL) can ultimately progress to chronic ulcerated lesions with strong local inflammatory reactions. The functional role of certain inflammasomes in mediating inflammation caused by Leishmania braziliensis needs to be addressed. By combining PCR-array, quantitative real-time PCR and immunohistochemical analysis, we identified inflammasome genes, such as IL-1ß, NLRP3, NLRP1, NLRC5, AIM2 and P2RX7, that were upregulated in LCL patients. Temporal gene expression studies showed that the early phase of LCL displayed increased NLRP3 and reduced AIM2 and NLRP1 expression, while the late stages showed increased AIM2 and NLRP1 and lower NLRP3 expression. Our findings also showed that AIM2, NLRP1, and P2RX7 promoted susceptibility to experimental L. braziliensis infection. These results highlight the importance of inflammasome machinery in human LCL and suggest that inflammasome machinery plays a role in the acute and chronic phases of the disease.

[Socioeconomic impact of the current management of severe biliary acute pancreatitis: comparative study]. / Impacto socioeconómico del manejo actual de la pancreatitis aguda de origen biliar severa: estudio comparativo.

OBJETIVE: Acute pancreatitis of biliary origin is a common gastrointestinal pathology, in which timely management still is the most important. The aims of this research is establish the socioeconomic impact in the current management of severe acute pancreatitis of biliary origin comparing two centers of the third level, one of high socioeconomic population and another of low in Bogotá, Colombia. MATERIALS AND METHODS: A retrospective, cross-sectional comparative study was conducted between January 2012 and December 2017, in two hospitals of Bogotá DC. We evaluated their socioeconomic characteristics, gender, time of evolution at the time of consultation, Marshall score, ICU stay, hospital stay, complications, surgical management and mortality. RESULTS: 101 patients from two different socioeconomic strata (high and low) were analyzed, where a 10 times higher risk of requiring a surgical procedure in the group of patients with low stratum was found, as well as a higher mortality compared with those of high stratum. (11.3% Vs 4.2%). There were also more complications in the low socioeconomic group with respect to the high, as in the exocrine failure (81.1% vs 31.3%) and the compartment syndrome (35.8% vs 4.2%). CONCLUSION: There is greater morbidity and mortality in patients of low socioeconomic status in the context of this pathology. This study can guide new research that increases the clarity of the socioeconomic impact on the outcomes of severe acute pancreatitis.

OBJETIVO: La pancreatitis aguda de origen biliar es una patología gastrointestinal común, en donde el tratamiento oportuno es el pilar más importante a pesar de sus discrepancias. El objetivo del estudio es establecer el impacto socioeconómico en el manejo actual de esta patología, comparando dos hospitales de tercer nivel de estrato socioeconómico alto y bajo de la ciudad de Bogotá, Colombia. MATERIALES Y MÉTODOS: Se realizó un estudio retrospectivo, comparativo de corte transversal entre enero de 2012 y diciembre de 2017, en dos hospitales de Bogotá D. C. Se evaluaron sus características socioeconómicas, género, tiempo de evolución al momento de la consulta, Marshall score, estancia en UCI, estancia hospitalaria, complicaciones, manejo quirúrgico y mortalidad. RESULTADOS: Se analizaron 101 pacientes de dos estratos socioeconómicos diferentes (alto y bajo). Se encontró que los pacientes de estrato bajo tienen un riesgo diez veces mayor de requerir un procedimiento quirúrgico. Asimismo, registraron una mayor mortalidad en comparación con pacientes de estrato alto (11,3% vs. 4,2%). También se evidenciaron más complicaciones en el grupo de nivel socioeconómico bajo con respecto al alto, como en la falla exocrina (81,1% vs. 31,3%) y el síndrome compartimental (35,8% vs. 4,2%). CONCLUSIÓN: Se encuentra mayor morbimortalidad en los pacientes de bajo nivel socioeconómico en el contexto de esta patología. Este estudio puede guiar a nuevas investigaciones acerca del impacto socioeconómico en los desenlaces de pancreatitis aguda severa.

Organizational effects of the antiandrogen, vinclozolin, on penis development in the mouse.

Endocrine disrupting chemicals (EDCs) are pollutants found throughout the environment that disrupt normal endocrine processes. In mice, penis development is thought to be most susceptible to EDCs during a critical developmental window occurring on embryonic days (E) 15.5-17.5. However, androgen signaling begins on E13.5 when androgen receptor (AR) protein is found in the genitalia and testosterone is circulating. We hypothesize that disrupting androgen signaling prior to the established critical window sensitizes the penis to future androgen disruption. To test this hypothesis, CD1 dams were exposed to vinclozolin or a corn oil solvent control on E13.5 and E14.5 and AR levels were measured with immunohistochemistry on E14.5. Early antiandrogen exposure reduced AR within nuclei and decreased intensity of AR expression within E14.5 genitalia. To evaluate the influence of antiandrogen exposure before the known critical window of penis development, two groups of pregnant dams (n = 3) were exposed to vinclozolin starting at either E13.5 or E14.5 and continued exposure through E16.5. Histology and M.O.U.S.E. scoring were used to quantify penis abnormalities. To account for differences in total doses mice experienced due to differences in length of dosing time, we compared animals that received the same total doses. Exposure to antiandrogens on E13.5 exacerbated malformations when exposure was continued through sexually dimorphic development. Both exposure time and vinclozolin dose are important for severity of vinclozolin-induced penis abnormalities in mice. This work shows that antiandrogen exposure prior to sensitive periods can exacerbate the effects of later antiandrogen exposure on reproductive development.

Emerging Role for the PERK/eIF2α/ATF4 in Human Cutaneous Leishmaniasis.

Leishmania parasites utilize adaptive evasion mechanisms in infected macrophages to overcome host defenses and proliferate. We report here that the PERK/eIF2α/ATF4 signaling branch of the integrated endoplasmic reticulum stress response (IERSR) is activated by Leishmania and this pathway is important for Leishmania amazonensis infection. Knocking down PERK or ATF4 expression or inhibiting PERK kinase activity diminished L. amazonensis infection. Knocking down ATF4 decreased NRF2 expression and its nuclear translocation, reduced HO-1 expression and increased nitric oxide production. Meanwhile, the increased expression of ATF4 and HO-1 mRNAs were observed in lesions derived from patients infected with the prevalent related species L.(V.) braziliensis. Our data demonstrates that Leishmania parasites activate the PERK/eIF2α/ATF-4 pathway in cultured macrophages and infected human tissue and that this pathway is important for parasite survival and progression of the infection.

Giant toads (Rhinella marina) living in agricultural areas have altered spermatogenesis.

Across diverse taxa, germ cell development is controlled by an intricate cascade of processes that are tightly controlled by the hypothalamic-pituitary-gonadal axis. Endocrine disturbances, such as those induced by endocrine disrupting chemicals (EDCs) can negatively affect spermatogenesis. Here, we investigate whether spermatogenesis is altered in the giant toad, Rhinella marina, living in agricultural areas where EDCs are used relative to suburban areas. We also ask if reductions in spermatogenesis were associated with developmental gonadal abnormalities (intersex) found in the same frogs. We found that toads in agricultural areas exhibited reduced spermatogenesis relative to non-agricultural animals, and that those reductions were not associated with gross gonadal abnormalities. All toads living in agricultural areas had reduced spermatogenesis relative to those living in non-agricultural areas regardless of whether they had gonadal abnormalities originating during development. Similarities in reproductive dysfunction among diverse taxa living in agricultural areas, including humans, suggest that many vertebrate taxa living in agricultural areas around the globe are likely experiencing some level of reproductive dysfunction.

Unexpectedly high leprosy seroprevalence detected using a random surveillance strategy in midwestern Brazil: A comparison of ELISA and a rapid diagnostic test.

BACKGROUND: Leprosy diagnosis is mainly based on clinical evaluation, although this approach is difficult, especially for untrained physicians. We conducted a temporary campaign to detect previously unknown leprosy cases in midwestern Brazil and to compare the performance of different serological tests. METHODS: A mobile clinic was stationed at the main bus terminal in Brasília, Brazil. Volunteers were quizzed and given a clinical exam to allow categorization as either patients, known contacts of patients or non-contacts, and blood was collected to determine anti-PGL-I and anti-LID-1 antibody titers by ELISA and by the NDO-LID rapid test. New cases of leprosy and the impact of performing this broad random surveillance strategy were evaluated. Accuracy values and concordance between the test results were evaluated among all groups. RESULTS: Four hundred thirty-four individuals were evaluated, and 44 (10.1%) were diagnosed with leprosy. Borderline forms were the most frequent presentation. Both tests presented higher positivity in those individuals with multibacillary disease. Serological tests demonstrated specificities arround 70% for anti-PGL-1 and anti-LID ELISA; and arround 40% for NDO-LID. Sensitivities ranged from 48 to 62%. A substantial agreement between NDO-LID and ELISA with concomitant positive results was found within leprosy patients (Kappa index = 0.79 CI95% 0.36-1.22). CONCLUSIONS: The unexpectedly high leprosy prevalence in this population indicates ongoing community-based exposure to Mycobacterium leprae antigens and high rates of subclinical infection. All tests showed low specificity and sensitivity values and therefore cannot be considered for use as stand-alone diagnostics. Rather, considering their positivity among MB patients and non-patients, these tests can be considered effective tools for screening and identifying individuals at high risk who might benefit from regular monitoring.

A validated protocol to quantify severity of male urogenital feminization using the MOUSE (Mouse objective urethral severity evaluation).

BACKGROUND: Congenital abnormalities vary in presentation, yet studies using model organisms tend to focus on occurrence rather than severity of the defect. Scoring severity of abnormalities in model systems allows explicit hypothesis testing during basic, translational, and reverse translational studies. We developed and validated a protocol to quantify severity of male urogenital feminization (hypospadias) in the mouse model. Hypospadias is one of the most common birth defects in the world. METHODS: To induce genital feminization, pregnant mice were exposed to different concentrations of the antiandrogen vinclozolin. Genitalia were photographed at gestational age 18.5. A dichotomous scoring system to evaluate genital feminization was developed, and validated against histological measurements of urethral length. A training protocol was developed for novice scorers, and criteria were defined to evaluate precision and accuracy of scores. RESULTS: Vinclozolin induced variation in hypospadias severity. Severity scores were tightly correlated with histologically determined urethral length and both techniques showed similar dose-response relationships. Novice observers were trained to precisely and accurately score hypospadias severity. CONCLUSION: This standardized scoring system advances the mouse as a model to study urogenital development, and will facilitate research on the mechanisms driving genital feminization in males, and aid translational hypospadias research.

Screening of biotechnical parameters for production of bovine inter-subspecies embryonic chimeras by the aggregation of tetraploid Bos indicus and diploid crossbred Bos taurus embryos.

The aggregation of a tetraploid zebu embryo (Bos indicus, a thermotolerant breed) with a diploid taurine embryo (Bos taurus, a thermosensitive breed) should create a complete taurine fetus, whose extra-embryonic components, e.g., the chorion, is derived mainly from the zebu embryo. These zebu-derived extra-embryonic components may interact positively with the taurine embryo/fetus during pregnancy in a tropical environment. We tested different parameters for the production of tetraploid Nelore (Bos indicus) embryos to be combined via aggregation with crossbred Bos taurus (diploid) embryos in order to produce viable chimeric blastocysts. Bovine (Bos indicus or crossbred Bos taurus) embryos were produced in vitro according to standard procedures. Two-cell Bos indicus embryos were submitted to electrofusion with varying numbers of pulses (1 or 2), voltages (0.4, 0.5, 0.75, 1.0, 1.4 and 5.0 kV/cm) and time (20, 25, 50 and 60 µs) to produce tetraploid embryos. Electrofused embryos were cultured with crossbred non-fused embryos to form chimeras that developed until the blastocyst stage. The best fusion parameter was 0.75 kV/cm for 60 µs. Four chimeric blastocysts (tetraploid Nelore with diploid crossbred Holstein) were formed after 31 attempts in 4 replicates (13%). We established an optimal procedure for the production of tetraploid Bos indicus (4n) embryos and embryonic chimeras by aggregation of crossbred Bos taurus (2n) with Bos indicus (4n) embryos. This technique would be valid in applied research, by producing exclusively taurine calves, but with placental elements from the Bos indicus breed, following transfer of these chimeras into recipient cows.