Acute murine-betacoronavirus infection impairs testicular steroidogenesis and the quality of sperm production.

Although several testicular alterations promoted by coronavirus infection have been demonstrated, the extent, causes, and players of testicular pathogenesis are not totally understood. The present study aimed to investigate the short-term effects on male fertility of intranasally administered murine hepatitis virus strain 3 (MHV-3), a member of the genus Betacoronavirus, which causes a severe systemic acute infection. This mouse model might be used as a in vivo prototype for investigating the impact of betacoronavirus on the endocrine and exocrine testicular functions with the advantage to be performed in a biosafety level 2 condition. Herein, we performed virological, histopathological, and molecular studies regarding the testicular spermatogenesis and the spermatic quality analyses in an MHV-3-infected C57BL/6 mice. The main outcomes showed that MHV-3 infects mouse testis and induces a testicular inflammatory state, impairing the steroidogenic pathway. The infection led to several alterations in the testicular parenchyma, such as: seminiferous epithelium sloughing, retention of residual bodies, germ cell apoptosis, alterations in intercellular junction proteins, and worse spermatogenic parameters. Moreover, the levels of plasmatic testosterone as well as the quality of sperm production reduced. Therefore, the present data suggest that the viral/inflammatory impairment of the steroidogenic pathway and the consequent imbalance of androgen levels is critical in testicular pathology, disturbing the SC barrier function and the germ cell differentiation. Our study is important for comprehending the effects of beta coronavirus infections on testis function in order to develop treatments that could prevent virus-mediated male infertility.

The South American lungfish Lepidosiren paradoxa as a new host for Trichodina quelenii / O lungfish sul-americano Lepidosiren paradoxa como novo hospedeiro para Trichodina quelenii

Abstract Recently, the South American lungfish Lepidosiren paradoxa is being found inside aquaculture ponds, and even though there are a few studies on their parasite fauna, there is still much to be reported. Thus, the objective of this study is to report parasitism by trichodinids in L. paradoxa, as these ciliate protozoa are related to injuries and mortality in fish farming. The lungfish were collected from experimental tanks, had their tegument scraped and the resultant mucus was analyzed under an optical microscope for morphological and morphometrical analyses in Giemsa and silver nitrate stained slides. The species found was identified as Trichodina quelleni. This is the first report of this parasite in L. paradoxa, and the second report in cultivated fish in Brazil.

Resumo Recentemente, o peixe pulmonado sul-americano Lepidosiren paradoxa tem sido encontrado em tanques de cultivo da aquicultura e, embora existam alguns estudos sobre a fauna de parasitas neste hospedeiro, ainda há muito a ser relatado. Assim, o objetivo deste estudo é relatar o parasitismo por tricodinídeos em L. paradoxa, pois esses protozoários ciliados estão relacionados a lesões e mortalidade na piscicultura. Os peixes foram coletados de tanques experimentais, tiveram seu tegumento raspado e o muco resultante foi analisado sob um microscópio óptico para análises morfológicas e morfométricas em lâminas impregnadas com Nitrato de Prata e com Giemsa. Os espécimes encontrados foram identificados como Trichodina quelleni. Este é o primeiro registro deste parasita em L. paradoxa, e a segunda ocorrência de Trichodina quelenii em peixes cultivados no Brasil.

The South American lungfish Lepidosiren paradoxa as a new host for Trichodina quelenii.

Recently, the South American lungfish Lepidosiren paradoxa is being found inside aquaculture ponds, and even though there are a few studies on their parasite fauna, there is still much to be reported. Thus, the objective of this study is to report parasitism by trichodinids in L. paradoxa, as these ciliate protozoa are related to injuries and mortality in fish farming. The lungfish were collected from experimental tanks, had their tegument scraped and the resultant mucus was analyzed under an optical microscope for morphological and morphometrical analyses in Giemsa and silver nitrate stained slides. The species found was identified as Trichodina quelleni. This is the first report of this parasite in L. paradoxa, and the second report in cultivated fish in Brazil.

The South American lungfish Lepidosiren paradoxa as a new host for Trichodina quelenii

Abstract Recently, the South American lungfish Lepidosiren paradoxa is being found inside aquaculture ponds, and even though there are a few studies on their parasite fauna, there is still much to be reported. Thus, the objective of this study is to report parasitism by trichodinids in L. paradoxa, as these ciliate protozoa are related to injuries and mortality in fish farming. The lungfish were collected from experimental tanks, had their tegument scraped and the resultant mucus was analyzed under an optical microscope for morphological and morphometrical analyses in Giemsa and silver nitrate stained slides. The species found was identified as Trichodina quelleni. This is the first report of this parasite in L. paradoxa, and the second report in cultivated fish in Brazil.

Resumo Recentemente, o peixe pulmonado sul-americano Lepidosiren paradoxa tem sido encontrado em tanques de cultivo da aquicultura e, embora existam alguns estudos sobre a fauna de parasitas neste hospedeiro, ainda há muito a ser relatado. Assim, o objetivo deste estudo é relatar o parasitismo por tricodinídeos em L. paradoxa, pois esses protozoários ciliados estão relacionados a lesões e mortalidade na piscicultura. Os peixes foram coletados de tanques experimentais, tiveram seu tegumento raspado e o muco resultante foi analisado sob um microscópio óptico para análises morfológicas e morfométricas em lâminas impregnadas com Nitrato de Prata e com Giemsa. Os espécimes encontrados foram identificados como Trichodina quelleni. Este é o primeiro registro deste parasita em L. paradoxa, e a segunda ocorrência de Trichodina quelenii em peixes cultivados no Brasil.

Testis Morphometry and Stages of the Seminiferous Epithelium Cycle in an Epididymal Sperm-storing Neotropical Vespertilionid, Myotis levis (Chiroptera).

Yellowish myotis, Myotis levis, is a seasonal, epididymal sperm-storing Neotropical vespertilionid. In the dry season, males show simultaneous testis regression and sperm storage in cauda epididymis, enabling them to mate during this season. In this study, we investigated seasonal variations in body mass, diameter and height of seminiferous tubules and nuclei of Leydig cells in a population of southeastern Brazil. We also determined the frequencies of the stages of the seminiferous epithelium cycle (SEC) of mature individuals of this population. Body mass and diameter of Leydig cell nuclei showed no significant differences between dry and rainy seasons and stages of annual reproductive cycle; however, all other morphometric parameters varied significantly. The relative cumulative frequency of pre-meiotic stages of the SEC (1-3) was 51%, of meiotic stage (4) was 2% and of post-meiotic stages (5-8) was 47%. We confirmed that the yellowish myotis presents seasonal sperm production as revealed by testis regression and epididymal sperm storage during the dry season.

Functional polymorphisms of interleukin-18 gene and risk of breast cancer in a Brazilian population.

Interleukin-18 (IL-18) is a key cytokine responsible for immune response and involved in the process of cancer development. In this case-control study, we tested whether IL-18 promoter polymorphism contributes to breast cancer susceptibility in Brazilian patients. The two groups studied were 154 patients with breast cancer and 118 healthy individuals. The frequency of IL-18 promoter single nucleotide polymorphisms (SNPs) at positions -607 (C/A) (rs1946518) and -137 (G/C) (rs187238) was determined by polymerase chain reaction analyses. The polymorphisms genotyped in this study showed a significant association with breast cancer under different genetic models. Both SNPs showed a positive association. For the IL18-607 polymorphism the best model was the codominant genetic model [CC vs AA, P = 0.004, odds ratio (OR) = 2.782, 95% confidence interval (CI) 1.385-5.589]. For IL18-137 statistical significance was found using the recessive genetic model (P = 0.008, OR = 3.896, 95% CI 1.427-10.639). The association between the haplotypes of the IL18 gene and breast cancer was further confirmed. Our results suggest that IL18-607 and IL18-137 polymorphism contributes to increase the breast cancer risk. To our knowledge, this is the first report regarding Brazilian breast cancer patients and IL18 promoter polymorphisms.

Chronic glucocorticoid treatment enhances lipogenic activity in visceral adipocytes of male Wistar rats.

AIM: Glucocorticoid (GC) in excess promotes the redistribution of adipose tissue from peripheral to central sites of the body. In this study, we characterized an experimental condition of prolonged GC excess and investigated its effect on the lipogenic metabolism in white adipose tissue. METHODS: Twenty male Wistar rats were divided into control (CON) and dexamethasone-treated (DEX) groups. DEX group received dexamethasone (0.25 mg kg(-1) day(-1) ) during 4 weeks, while CON group received saline. Animals were killed and subcutaneous (SC), retroperitoneal (RP) and mesenteric (MS) fat pads were excised, weighed and processed for adipocyte isolation, morphometric cell analysis and incorporation of glucose into lipids. RESULTS: The treatment effectively blocked hypothalamic-pituitary-adrenal axis, as verified by a 58% decrease in plasma corticosterone levels and 19% atrophy in adrenal glands in DEX group. Animals from DEX group presented insulin resistance, glucose intolerance, dyslipidaemia and increased insulin and leptin plasma levels and hypertrophied adipocytes. They showed increased lipogenesis in RP and MS depots, with increased incorporation of glucose into fatty acids of triacylglycerol. Increased activity of lipogenic enzymes ATP-citrate lyase, fatty acid synthase, glucose-6-phosphate dehydrogenase and malic was only seen in the MS depot in DEX group, while gene expression of these enzymes was enhanced in SC and MS fat depots. CONCLUSION: The adaptations promoted by GC treatment in adipose metabolism seemed to be mainly due to the increased activity of enzymes that supply the NADPH required for lipogenesis than to the increase in enzymes that more directly deal with fatty acid synthesis itself.

CompCytogen of two of the smallest Amazonian fishes: Fluviphylax simplex Costa, 1996 and Fluviphylax zonatus Costa, 1996 (Cyprinodontiformes, Poeciliidae).

The genus Fluviphylax Whitley, 1965is comprized of five valid species (Fluviphylax pygmaeus Myers et Carvalho, 1955, Fluviphylax zonatus, Fluviphylax simplex, Fluviphylax obscurus Costa, 1996,and Fluviphylax palikur Costa et Le Bail, 1999), which are endemic to the Amazon region. These fishes are the smallest known South American vertebrates and among the smallest know vertebrates on Earth. All species but the type Fluviphylax pygmaeus have been described in late 1990's, and much remains unknown about the biology, taxonomy and systematics of this group of fishes. The aims of the present study were to establish the diploid and haploid number of Fluviphylax zonatus and Fluviphylax simplex, and to find species-specific markers for the discrimination of taxa. The diploid number for both species was 48 chromosomes, with no sex chromosome heteromorphism. Fluviphylax zonatus exhibited the karyotypic formula 4m+8sm+22st+14a and FN=82, and Fluviphylax simplex exhibited 4m+16sm+18st+10a and FN=86. The determination of the total mean length of the chromosomes and their grouping into five size classes demonstrated different chromosome composition of the two species. This difference was further supported by the distribution of constitutive heterochromatin. The meiotic analysis revealed 24 bivalents in both species, but Fluviphylax zonatus exhibited chromosomes with late pairing of the telomeric portions in the pachytene. These data reveal that cytogenetic characterization is useful and important for the discrimination of these species. Our study further indicates that this method could be employed in the analysis of other species of small fishes that are difficult to distinguish using traditional morphological traits or are morphologically cryptic.

Thermoluminescence dating of Brazilian indigenous ceramics.

Two indigenous ceramics fragments, one from Lagoa Queimada (LQ) and another from Barra dos Negros (BN), both sites located on Bahia state (Brazil), were dated by thermoluminescence (TL) method. Each fragment was physically prepared and divided into two fractions, one was used for TL measurement and the other for annual dose determination. The TL fraction was chemically treated, divided in sub samples and irradiated with several doses. The plot extrapolation from TL intensities as function of radiation dose enabled the determination of the accumulated dose (D(ac)), 3.99 Gy and 1.88 Gy for LQ and BN, respectively. The annual dose was obtained through the uranium, thorium and potassium determination by ICP-MS. The annual doses (D(an)) obtained were 2.86 and 2.26 mGy/year. The estimated ages were approximately 1375 and 709 y for BN and LQ ceramics, respectively. The ages agreed with the archaeologists' estimation for the Aratu and Tupi tradition periods, respectively.

Chemical process to separate iron oxides particles in pottery sample for EPR dating.

Ancient potteries usually are made of the local clay material, which contains relatively high concentration of iron. The powdered samples are usually quite black, due to magnetite, and, although they can be used for thermoluminescene (TL) dating, it is easiest to obtain better TL reading when clearest natural or pre-treated sample is used. For electron paramagnetic resonance (EPR) measurements, the huge signal due to iron spin-spin interaction, promotes an intense interference overlapping any other signal in this range. Sample dating is obtained by dividing the radiation dose, determined by the concentration of paramagnetic species generated by irradiation, by the natural dose so as a consequence, EPR dating cannot be used, since iron signal do not depend on radiation dose. In some cases, the density separation method using hydrated solution of sodium polytungstate [Na6(H2W12O40).H2O] becomes useful. However, the sodium polytungstate is very expensive in Brazil; hence an alternative method for eliminating this interference is proposed. A chemical process to eliminate about 90% of magnetite was developed. A sample of powdered ancient pottery was treated in a mixture (3:1:1) of HCl, HNO(3) and H(2)O(2) for 4h. After that, it was washed several times in distilled water to remove all acid matrixes. The original black sample becomes somewhat clearer. The resulting material was analyzed by plasma mass spectrometry (ICP-MS), with the result that the iron content is reduced by a factor of about 9. In EPR measurements a non-treated natural ceramic sample shows a broad spin-spin interaction signal, the chemically treated sample presents a narrow signal in g=2.00 region, possibly due to a radical of (SiO(3))(3-), mixed with signal of remaining iron [M. Ikeya, New Applications of Electron Spin Resonance, World Scientific, Singapore, 1993, p. 285]. This signal increases in intensity under gamma-irradiation. However, still due to iron influence, the additive method yielded too old age-value. Since annealing at 300 degrees C, Toyoda and Ikeya [S. Toyoda, M. Ikeya, Geochem. J. 25 (1991) 427-445] states that E1'-signal with maximum intensity is obtained, while annealing at 400 degrees C E1'-signal is completely eliminated, the subtraction of the second one from 300 degrees C heat-treated sample isolate E1'-like signal. Since this is radiation dose-dependent, we show that now EPR dating becomes possible.

Biochemical and thermostability features of acetyl esterase Aes from Escherichia coli.

Previously we characterized an acetyl-esterase from Escherichia coli, formally Aes, from a thermodynamic point of view in comparative studies with thermophilic homologs. Since the enzyme appeared unusually resistant to the thermal denaturation we analysed the kinetic behaviour with respect to the temperature. The enzyme displays a surprising optimal temperature at 65 degrees C, showing a specific activity of 250 U/mg using pNP-butanoate as substrate, but a low kinetic stability at the same temperature (t(1/2) of inactivation=5 min). By a random mutagenesis approach we searched for mutated versions of Aes with increased thermostability. We found the mutant T74A, which shows the same specific activity of wild type but a t(1/2) of inactivation of 30 min at 65 degrees C.

A tank-to-wheel analysis tool for energy and emissions studies in road vehicles.

Currently, oil based fuels are the primary energy source of road transport. The growing need for oil independence and CO(2) mitigation has lead to the increasing importance of alternative fuel usage. CO(2) is produced not only as the fuel is used in the vehicle (tank-to-wheel contribution), but also upstream, from the fuel extraction to the refueling station (well-to-tank contribution), and the life cycle of the fuel production (well-to-wheel contribution) must be considered in order to analyse the global impact of the fuel utilization. A road vehicle tank-to-wheel analysis tool that may be integrated with well-to-tank models was developed in the present study. The integration in a demonstration case study allowed to perform a life cycle assessment concerning the utilization of diesel and natural gas fuels in a specific network line of a bus transit company operating in the city of Porto, Portugal.

Elective neck dissection versus observation in stage I squamous cell carcinomas of the tongue and floor of the mouth.

A retrospective study was undertaken of patients with T1N0M0 squamous cell carcinoma of the oral tongue and floor of the mouth who underwent surgical treatment between 1985 and 1995. Evaluation of two groups of patients (neck dissection versus observation) was made according to the management of the neck. Results were obtained regarding the presence of occult metastases, recurrence in the neck, treatment failure, results of salvage treatment, and disease-free survival. Forty-nine patients underwent surgical treatment: 25 resection of primary and 24 resection plus neck dissection. Overall incidence of regional metastases was 24.5%. Eight patients (16%) developed recurrence of the disease. Seven (14%) had regional recurrences (including 1 with distant metastases) and 1(2%) had local recurrence. Twenty-four percent of patients from the resection of primary group developed neck recurrences in comparison with 4% of the resection plus neck dissection group (P = 0.05). Overall salvage rate was 37.5%. Second primary tumors developed in 16% of patients. Patients who underwent elective neck dissection had a 23% higher disease-free survival rate compared with those who underwent resection of the tumor alone (P = 0.03). The findings of this study stress the importance of control of the neck in early oral cancer. Elective neck dissection significantly improved regional control of the disease.

Supracricoid laryngectomy with CHEP: functional results and outcome.

OBJECTIVES: To assess whether supracricoid laryngectomy with cricohiodoepiglottopexy could successfully reach the cure and preserve the voice in glottic laryngeal cancer, we studied 27 patients with T2/T3 squamous cell carcinoma of the larynx treated in our institution with cricohiodoepiglottopexy. STUDY DESIGN: A retrospective analysis has been carried out between 1995 through 1997. We classified 11 patients as T2N0M0 and 16 patients as T3N0M0. Nineteen patients had bilateral selective lateral neck dissection, 3 patients had unilateral lateral neck dissection, and 5 patients had undissected neck. Survival was analyzed under the Kaplan-Meyer method. RESULTS: Five patients had postoperative complications, 2 were treated with a total laryngectomy. The remaining 25 patients kept the normal airway, swallowing, and speech. None of the patients in the neck dissection group had neck metastasis. Two patients had recurrences, 1 with local recurrence was treated with a total laryngectomy and is alive without disease; the other patient had neck recurrence, was treated with radical neck dissection plus radiotherapy, and is dead of the disease. One patient had a second tumor in oropharynx treated with palliative radiotherapy and is dead of the disease. Three years disease-free survival was 75% for T2 and 79% for T3. CONCLUSIONS: This technique is useful in the treatment of selected cases of T3/T2 glottic cancer regarding the extent of disease. The incidence of complications in need of a complete laryngectomy does not compromise the functionality of this technique. The survival is comparable to patients who submitted to total laryngectomy and near-total laryngectomy, regarding the extent of the lesion.

QBC for the diagnosis of human and canine american visceral leishmaniasis: preliminary data.

"Quantitative Buffy Coat" (QBC) is a direct and fast fluorescent method used for the identification of blood parasites. Since Leishmania chagasi circulates in blood, we decided to test it in American visceral leishmaniasis (AVL). Bone marrow (BM) and peripheral blood (PB) of 49 persons and PB of 31 dogs were analyzed. QBC was positive in BM of 11/11 patients with AVL and in 1/6 patients with other diseases. Amastigotes were identified in PB of 18/22 patients with AVL and in none without AVL. The test was positive in 30 out of the 31 seropositive dogs and in 28/28 dogs with Leishmania identified in other tissues. QBC is a promising method for diagnosis of human AVL, and possibly for the exam of PB of patients with AVL/AIDS, for the control of the cure and for the identification of asymptomatic carriers. Because it is fast and easy to collect and execute, QBC should be evaluated for programs of reservoir control.

Range of validity of the Rayleigh-Debye-Gans theory for optics of fractal aggregates.

The range of validity of the Rayleigh-Debye-Gans approximation for the optical cross sections of fractal aggregates (RDG-FA) that are formed by uniform small particles was evaluated in comparison with the integral equation formulation for scattering (IEFS), which accounts for the effects of multiple scattering and self-interaction. Numerical simulations were performed to create aggregates that exhibit mass fractallike characteristics with a wide range of particle and aggregate sizes and morphologies, including x(p) = 0.01-1.0, ‖m - 1‖ = 0.1-2.0, N = 16-256, and D(f) = 1.0-3.0. The percent differences between both scattering theories were presented as error contour charts in the ‖m - 1‖x(p) domains for various size aggregates, emphasizing fractal properties representative of diffusion-limited cluster-cluster aggregation. These charts conveniently identified the regions in which the differences were less than 10%, between 10% and 30%, and more than 30% for easy to use general guidelines for suitability of the RDG-FA theory in any scattering applications of interest, such as laser-based particulate diagnostics. Various types of aggregate geometry ranging from straight chains (D(f) ≈ 1.0) to compact clusters (D(f) ≈ 3.0) were also considered for generalization of the findings. For the present computational conditions, the RDG-FA theory yielded accurate predictions to within 10% for ‖m - 1‖ to approximately 1 or more as long as the primary particles in aggregates were within the Rayleigh scattering limit (x(p) ≤ 0.3). Additionally, the effect of fractal dimension on the performance of the RDG-FA was generally found to be insignificant. The results suggested that the RDG-FA theory is a reasonable approximation for optics of a wide range of fractal aggregates, considerably extending its domain of applicability.

Proteína C reactiva en síndrome diarreico agudo / C-reactive protein in acute diarrhea syndrome

La proteína C reactiva se ha usado en múltiples enfermedades infecciosas en la orientación de la etiología viral o bacteriana; su utilidad en relación al síndrome diarreico agudo (SDA) se desconoce. Con la hipótesis que las infecciones bacterianas enteroinvasoras muestran un valor mayor de PCR que las virales, se estudiaron 47 niños con SDA. Se realizaron: PCR, 2 coprocultivos, tinción violeta bicarbonato de Hucker y aislamiento de rotavoris por inmunoelectroforesis. Se detectaron 11 casos con bacteria enteroinvasoras en deposiciones, 9 con rotavirus , 1 mixto y 26 pacientes con estudio etiológico negativo. Para las infecciones virales el promedio, mediana y rango de PCR fueron (mg/lt): 13,55; 8; (0-75) y para las bacterias invasoras fueron: 116,7; 116; (1-318) respectivamente (p<0,01). Ocho de 11 niños con infección bacteriana mostraron PCR > 30 mg/lt versus 1 de 9 con infección viral (p=0,0091); 7 de los primeros tuvieron valores > 100 mg/lt. Los resultados obtenidos confirman nuestra hipótesis

Comparación de métodos de diagnóstico citológico, inmunofluorescencia y aislamiento viral en la detección de citomegalovirus (CMV) en orina / Comparision of diagnosis methods: cytology, immunofluorescence and viral isolation in the detection of urine cytomegalovirus

Las infecciones por citomegalovirus humano (CMV), ampliamente diseminadas, han sido señaladas como la principal causa de morbimortalidad en individuos inmunocomprometidos. Es característico en estas infecciones la aparición de grandes células con inclusiones intranucleares y/o intracitoplasmática. Con esta base se diseñó una investigación para evaluar el método de diagnóstico citológico y la detección de antígenos virales mediante la técnica de inmunofluorescencia indirecta (IFI) en relación al aislamiento viral en muestras de orina. Se estudiaron 70 casos con búsqueda de cuerpos de inclusión citomegálicos (CIC) y aislamiento viral. En 55 de estos se estudió además con IFI. Se encontro CIC en el 84% (59/70) de los casos, el aislamiento resultó positivo en un 29% (20/70) y la IFI alcanzó un 42% de positividad (23/55). En todos los casos con aislamiento positivo se encontró CIC en el sedimento de orina. La IFI que en relación al aislamiento viral parece carecer de especificidad, debe ser considerada ya que es un método que detecta específicamente antígenos virales. Finalmente, se recomienda trabajar estos métodos de diagnóstico con tres muestras de orina para optimizar su rendimiento