Analysis of microbial communities of ocular prostheses and anophthalmic sockets using 16S rRNA gene sequencing.

Ocular prostheses are liable to host microbial adhesion, which can favor conjunctival inflammation. Knowing the microbiota of the ocular prosthesis and anophthalmic socket is important for predicting infection-related pathogens. This study evaluated the microbiota of the prostheses and anophthalmic sockets of six individuals by 16S rRNA sequencing. The microbial abundance and diversity were analyzed using the Operational Taxonomic Units at the genus level. Forty-seven phyla, capturing 1,258 named genera, were recorded in the sample set. In both sites, the most frequent genera were Fusobacterium, Staphylococcus, Prevotella, and Streptococcus. The microbial abundance was higher for the anophthalmic socket than for the prosthesis. Alpha diversity showed no significant differences in bacterial richness or diversity between sites. Although the microbial abundance in the anophthalmic socket was higher, both sites had similar microbiota with high diversity and low dominance among the genera.

The CODATwins Project: The Current Status and Recent Findings of COllaborative Project of Development of Anthropometrical Measures in Twins.

The COllaborative project of Development of Anthropometrical measures in Twins (CODATwins) project is a large international collaborative effort to analyze individual-level phenotype data from twins in multiple cohorts from different environments. The main objective is to study factors that modify genetic and environmental variation of height, body mass index (BMI, kg/m2) and size at birth, and additionally to address other research questions such as long-term consequences of birth size. The project started in 2013 and is open to all twin projects in the world having height and weight measures on twins with information on zygosity. Thus far, 54 twin projects from 24 countries have provided individual-level data. The CODATwins database includes 489,981 twin individuals (228,635 complete twin pairs). Since many twin cohorts have collected longitudinal data, there is a total of 1,049,785 height and weight observations. For many cohorts, we also have information on birth weight and length, own smoking behavior and own or parental education. We found that the heritability estimates of height and BMI systematically changed from infancy to old age. Remarkably, only minor differences in the heritability estimates were found across cultural-geographic regions, measurement time and birth cohort for height and BMI. In addition to genetic epidemiological studies, we looked at associations of height and BMI with education, birth weight and smoking status. Within-family analyses examined differences within same-sex and opposite-sex dizygotic twins in birth size and later development. The CODATwins project demonstrates the feasibility and value of international collaboration to address gene-by-exposure interactions that require large sample sizes and address the effects of different exposures across time, geographical regions and socioeconomic status.

Characterization of teratogenic potential and gene expression in canine and feline amniotic membrane-derived stem cells.

The biosafety of innovative procedures that utilize stem cells in regenerative medicine has been addressed in several studies. Previous work has showed no tumour formation following the use of feline and human amniotic membrane-derived stem cells (AMSCs). In contrast, tumour formation was observed when canine AMSCs were utilized. These findings suggested that feline and human, but not canine, AMSCs are suitable for cell transplantation trials. This study aimed to further evaluate the feasibility of utilizing canine AMSCs for transplantation purposes as well as for felines. We tested teratoma formation following cell injection into BALB/c nude mice and then assessed expression of haematopoietic, mesenchymal, tumorigenic, pluripotency and cellular regulation markers using flow cytometry and qPCR. The use of canine AMSCs did not result in macroscopic tumour formation as determined 60 days after transplantation. The immunophenotypic characterization by flow cytometry revealed expression of mesenchymal markers (CD73 and CD90) and expression of the pluripotent marker OCT4 and SOX2. Quantitative PCR analysis revealed that there were no differences in the patterns of gene expression (CD34, CD73, OCT4, CD30 and P53) between canine and feline AMSCs, with the exception of the expression of SOX2 and CD90.

Controversial results of therapy with mesenchymal stem cells in the acute phase of canine distemper disease.

Distemper disease is an infectious disease reported in several species of domestic and wild carnivores. The high mortality rate of animals infected with canine distemper virus (CDV) treated with currently available therapies has driven the study of new efficacious treatments. Mesenchymal stem cell (MSC)-based therapy is a promising therapeutic option for many degenerative, hereditary, and inflammatory diseases. Therefore, the aim of this study was to characterize stem cells derived from the canine fetal olfactory epithelium and to assess the systemic response of animals infected with CDV to symptomatic therapy and treatment with MSCs. Eight domestic mongrel dogs (N = 8) were divided into two groups: support group (SG) (N = 5) and support group + cell therapy (SGCT) (N = 3), which were monitored over 15 days. Blood samples were collected on days 0, 6, 9, 12, and 15 to assess blood count and serum biochemistry (urea, creatinine, alanine transferase, alkaline phosphatase, gamma-glutamyl transferase, total protein, albumin, and globulin), and urine samples were obtained on days 0 and 15 for urinary evaluation (urine I). The results showed a high mortality rate (SG = 4 and SGCT = 2), providing inadequate data on the clinical course of CDV infection. MSC therapy resulted in no significant improvement when administered during the acute phase of canine distemper disease, and a prevalence of animals with high mortality rate was found in both groups due to the severity of symptoms.

Denture Adhesives Associated with Silver Vanadate: Antimicrobial Approach Against Multi- Species Biofilms on Acrylic Resin Surfaces.

Alternatives have been sought to add an antimicrobial property to denture adhesives. This study evaluated the antimicrobial potential of adhesives associated with nanostructured silver vanadate decorated with silver nanoparticles (ß-AgVO3). Specimens in acrylic resin were treated with the adhesives associated with ß-AgVO3 (1%, 2.5%, 5% and 10%). As control, specimens treated only with Ultra Corega Cream (UCC) or Ultra Corega Powder (UCP) adhesive were used. Multispecies biofilm of Candida albicans, Candida glabrata, Streptococcus mutans and Staphylococcus aureus was evaluated by counting colony forming units per milliliter (CFU/mL), colorimetric assay and fluorescence microscopy. The data were analyzed using the two-way analysis of variance (ANOVA) and Bonferroni multiple comparisons test (α=0.05). For both adhesives, a small amount of ß-AgVO3 (1%) completely inhibited S. mutans (P⟨0.05). For the other microorganisms, there was a reduction in metabolic activity and complete inhibition in the groups with intermediate or greater amounts of nanomaterial (P⟨0.05), except for C. albicans, which was reduced (P⟨0.05) but not completely inhibited in UCP. Microscopy that showed less biofilm in the groups with ß-AgVO3 and in the UCC than UCP. Denture adhesives in powder and cream form with ß-AgVO3 showed potential antimicrobial activity against multispecies biofilm. Powder adhesive showed higher biofilm formation.

A distinct tymovirus infecting Cassia hoffmannseggii in Brazil.

Leaves of Cassia hoffmannseggii, a wild fabaceous species found in the Atlantic Forest, with a severe mosaic symptom were collected in Pernambuco State, Brazil. By transmission electron microscopy, two types of virus particles were found: the first was recognized as particles of a potyvirus, which was later identified as Cowpea aphid-borne mosaic virus; and the second was isometric and present in high concentration. The observation of vesicles at the periphery of chloroplasts suggested a tymovirus infection, which was confirmed by subsequent assays. A serological assay against several tymovirus antisera resulted in positive reaction of this tymo-like virus with an antiserum of Passion fruit yellow mosaic virus. By means of RT-PCR and using degenerated primers for the conserved region of RNA-dependent RNA polymerase (RdRp) gene of tymoviruses, a specific DNA fragment was amplified and sequenced. Based on this sequence, a specific forward primer was synthesized and successfully used to amplify the 3' terminal genome region, containing the partial RdRp gene and the complete coat protein (CP) sequences. The CP was 188 amino acids (aa) long, and the highest CP aa identity was observed with Kennedya yellow mosaic virus (61 %). Based on the current ICTV demarcation criterion, this isolate was considered as a distinct tymovirus and tentatively named as Cassia yellow mosaic-associated virus.

Characterization of yolk sac proteins of Bos indicus cattle embryos.

The yolk sac is an embryonic membrane that is essential for the embryo's initial survival in many mammals. It also plays an important role in the production of proteins necessary for development. We studied proteins of the yolk sac in bovine embryos at up to 40 days of gestation. We examined the yolk sac of 17 bovine embryos at different gestational periods, measuring α-fetoprotein, α-1-antitrypsin, and transferrin. This experiment was carried out by Western blot technique, associated with electrophoresis on a 6% sodium dodecyl sulfate polyacrylamide gel. Mouse monoclonal antibody anti-human-α-fetoprotein, mouse antibody anti-human-transferrin and rabbit polyclonal anti-human-α-1-antitrypsin were used as primary antibodies, and conjugated peroxidase as a secondary antibody. We detected the three proteins in some of the yolk sac samples; however, the bands in some specimens (samples) were weak, maybe a result of poor antigen-antibody reaction, since the antibodies used in this study were not specific to bovine proteins. The fact that weak bands appeared might be due to a weak cross-reaction.

Frequency of the S65C mutation in the hemochromatosis gene in Brazil.

Development of hereditary hemochromatosis is associated with the C282Y, H63D or S65C mutations in the hemochromatosis gene. Though there is extensive knowledge about the former two, there is little information on the mechanism of action and the allelic frequency of the S65C mutation. We examined the prevalence of the S65C mutation of the hemochromatosis gene in Brazilians with clinical suspicion of hereditary hemochromatosis. Genotyping for this mutation was carried out in 633 individuals with clinical suspicion of hereditary hemochromatosis, using the polymerase chain reaction, followed by enzymatic digestion. The sample comprised 77.1% men and 22.9% women, giving a ratio of approximately 3:1; the mean age was 48.8 +/- 13.8 years. More than half (57.3%) of the individuals in the sample were 41 to 60 years old. The frequency of heterozygotes for this mutation was 0.016; no homozygous mutant patients were found. This is the first analysis of the S65C mutation in individuals suspected of having hereditary hemochromatosis in Brazil.

A985G mutation incidence in the medium-chain acyl-CoA dehydrogenase (MCAD) gene in Brazil.

In view of the serious consequences of medium-chain acyl-CoA dehydrogenase (MCAD) deficiency and the absence of information about its incidence in the Brazilian population, we examined the frequency of the A985G mutation in the MCAD gene. A retrospective analysis was made of data on 1722 individuals (844 females) genotyped for the A985G mutation in the MCAD gene, using genomic DNA extracted from peripheral blood leukocytes and genotyping with PCR-RFLP; 0.41% of these individuals were heterozygous for the A985G mutation. The mutant homozygous genotype was not found. The 985G mutant and 985A normal alleles had allelic frequencies of 0.0020 and 0.9980, respectively. Given the A985G allele frequency, genotyping would be recommended in cases of family history of MCAD deficiency and sudden infant death syndrome, and when there is suspicion of medium-chain fatty acid metabolic alterations; genetic counseling should be offered in cases involving 985GG and A985G individuals and consanguineous marriages.

Bone marrow-derived cells are recruited by the melanoma tumor with endothelial cells contributing to tumor vasculature.

PURPOSE: Tumor expansion is dependent on neovascularization, a process that requires sustained new vessel formation. Although the critical role of angiogenesis by endothelial sprouting in this process, controversy still prevails on whether angiogenesis involving bone marrow-derived endothelial cells, does contribute to this process. This study aims to evaluate the recruitment of bone marrow-derived cells by the melanoma tumor, including endothelial cells, and if they contribute to angiogenesis. METHODS: A chimeric mouse model of GFP bone marrow was used to induce melanoma tumors derived from murine B16-F10 cell line. These tumors were evaluated for the presence of myeloid cells (CD11b), T lymphocytes (CD3, CD4 and CD8) and endothelial cells (VEGFR2 and CD31) derived from bone marrow. RESULTS: Mice transplanted with GFP+ cells showed significant bone marrow chimerism (90.9 ± 0.87 %) when compared to the GFP transgenic mice (90.66 ± 2.1 %, p = 0.83) demonstrating successful engraftment of donor bone marrow stem/progenitor cells. Analysis of the murine melanoma tumor showed the presence of donor cells in the tumors (3.5 ± 1.7 %) and interestingly, these cells represent endothelial cells (CD31+ cells; 11.5 ± 6.85 %) and myeloid cells (CD11b+ cells; 80 ± 21 %), but also tumor-infiltrating lymphocytes (CD8+ T cells, 13.31 ± 0.2 %; CD4+ T-cells, 2.1 ± 1.2 %). Examination of the tumor endothelium by confocal microscopy suggests the presence of donor CD31+/GFP+ cells in the wall of some blood vessels. CONCLUSION: This study demonstrates that bone marrow-derived cells are recruited by the murine melanoma tumor, with myeloid cells and CD4 and CD8 T lymphocytes migrating as antitumor immune response, and endothelial cells participating of the tumor blood vessels formation.

Patients' perceived level of social isolation affects the prognosis of low back pain.

BACKGROUND: Perceived social isolation is prevalent among patients with low back pain (LBP) and could be a potential prognostic factor for clinical outcomes following an episode of LBP. METHODS: A secondary analysis of an original prospective cohort study, which investigated the validity of the Danish version of the STarT Back Screening Tool (STarT), investigated whether social isolation predicts the clinical outcomes of disability, anxiety, depression and pain catastrophizing in people with LBP. Patients with LBP of any duration (N = 204) from Middelfart, Denmark, were included. Social isolation was measured at baseline using the friendship scale (score ranges from 0 to 24, with lower values meaning higher perceived social isolation), and outcomes were measured at baseline and at 6-month follow-up. Regression models investigated whether social isolation at baseline predicted the outcomes at 6-month follow-up. RESULTS: Some level of social isolation was reported by 39.2% of the participants (n = 80) with 5.9% (n = 12) being very socially isolated. One-point difference on social isolation predicted one point on a 100-point disability scale (adjusted unstandardized coefficient: -0.91; 95% confidence interval (CI): -1.56 to -0.26). Social isolation predicted anxiety; however, a change of one point on the social isolation scale represents a difference of only 0.08 points on a 22-point scale in anxiety (95% CI: 0.01-0.15) and is unlikely to denote clinical importance. Social isolation did not predict pain catastrophizing or depression. CONCLUSIONS: Patients' perceived social isolation predicts disability related to LBP. Further understanding of the role of social isolation in LBP is warranted.

Isolation and characterization of mesenchymal stem cells from the yolk sacs of bovine embryos.

The yolk sac (YS) represents a promising source of stem cells for research because of the hematopoietic and mesenchymal cell niches that are present in this structure during the development of the embryo. In this study, we report on the isolation and characterization of YS tissue and mesenchymal stem cells (MSCs) derived from bovine YSs. Our results show that the YS is macroscopically located in the exocoelomic cavity in the ventral portion of the embryo and consists of a transparent membrane formed by a central sac-like portion and two ventrally elongated projections. Immunohistochemistry analyses were positive for OCT4, CD90, CD105, and CD44 markers in the YS of both gestational age groups. The MSCs of bovine YS were isolated using enzymatic digestion and were grown in vitro for at least 11 passages to verify their capacity to proliferate. These cells were also subjected to immunophenotypic characterization that revealed the presence of CD90, CD105, and CD79 and the absence of CD45, CD44, and CD79, which are positive and negative markers of MSCs, respectively. To prove their multipotency, the cells were induced to differentiate into three cell types, chondrocytes, osteoblasts, and adipocytes, which were stained with tissue-specific dyes (chondrogenic: Alcian Blue, osteogenic: Alizarin Red, and adipogenic: Oil Red O) to confirm differentiation. Gene expression analyses showed no differences in the patterns of gene expression between the groups or passages tested, with the exception of the expression of SOX2, which was slightly different in the G1P3 group compared to the other groups. Our results suggest that YS tissue from bovines can be used as a source of MSCs, which makes YS tissue-derived cells an interesting option for cell therapy and regenerative medicine.

Occurrence of Smut Caused by a Ustilago sp. on Dry-Beans.

In September 1997, after the tropical winter season and following a long warm (28 to 33°C) and rainy (27 mm) period, smut symptoms were observed on two dry-bean (Phaseolus vulgaris) fields in Brazil. Brownish to blackish powdery masses of spores were observed on stems, pods, and roof green and senescent plants. No galls were observed. Hyphae in infected tissue were intercellular; no clamp connections were observed. Typical globose teliospores were observed that measured 7 to 11 µm. Spores were uninucleate or binucleate; predominantly brownish with flattened poles and equatorial band; the exospore wall was thick and surface ornamented (echinulate-type). Teliospores incubated in water drops generated transversely septate promycelia, unbranched, producing terminal and lateral basidiospores. The fungus did not grow well on potato dextrose agar (PDA) medium, generating yeastlike colonies. To confirm pathogenicity of a Ustilago sp., spores collected from field-infected plants were inoculated on 8- and 20-day old seedlings of dry-bean cultivars Rosinha and Perola. Spore suspensions of 105 teliospores were sprayed all over the plants. After inoculation, plants were kept in a moist chamber (relative humidity [RH] 100%) for 2 days. At the third day, inoculated plants were covered with clear polyethylene bags and transferred to the greenhouse (RH 50%, temperature 25 ± 2°C). Noninoculated controls were included for comparisons. Within 2 weeks, all inoculated plants reproduced field symptoms of darkened tissues covered by profuse sori containing teliospores and basidiospores. Eight-day-old plants died 2 weeks after inoculation. Noninoculated plants did not develop any symptoms. Teliospores obtained from dead plants were identical to those used for inoculation, thus completing Koch's postulates. Fischer (1), and later Fischer and Shaw (2), proposed a species concept for Ustilago based on morphology (primarily of the teliospore), symptomatology, and host specialization at the host-family level. Although the teliospores found on dry-beans, in Brazil, are much like those of Ustilago maydis, the fungus did not cause galls on plants, was able to infect roots and kill seedlings. and is very well adapted to a legume plant. Cross inoculation tests indicated that spores from dry-beans cannot cause disease symptoms on maize. We therefore regard the bean smut as a new species. We are not aware of any other Ustilago sp. being reported on a legume host. References: (1) G. W. Fischer. 1953. Manual of the North America Smut Fungi. (2) G. W. Fischer and C. G. Shaw. Phytopathology 43:181, 1953.

[Linear accelerator radiosurgery]. / Radiocirurgia por acelerador linear.

Radiosurgery is the precise radiation of a known intracranial target with a high dose of energy, sparing the adjacent nervous tissue. Technological advances in the construction of linear accelerators, stereotactic instruments and in computer sciences made this technique easier to perform and affordable. The main indications for radiosurgery are inoperable cerebral vascular malformations, vestibular and other cranial schwannomas, skull base meningiomas, deep seated gliomas and cerebral metastases. More recently, the development of fraccionated stereotactic radiotherapy increased the spectrum of indications to bigger lesions and to those adjacent to critical nervous structures. We present our initial experience in the treatment of 31 patients. An adequate control of the neoplastic lesions was obtained and the adequate time of observation is still needed to evaluate the results in arteriovenous malformations.

Population dynamics of calyptrate Diptera (Muscidae and Sarcophagidae) at the Rio-Zoo Foundation, Rio de Janeiro, RJ, Brazil.

Twenty-seven species of calyptrate muscoids (Muscidae and Sarcophagidae) were collected from December 1993 to November 1994 with wind oriented traps (W.O.T.) baited with decomposing beef liver at the Rio de Janeiro Zoo. The most abundant species found were Musca domestica (57.84%), Peckia chrysostoma (28.16%), Ophyra aenescens (17.11%), Oxysarcodexia thornax (17.82%), Synthesiomyia nudiseta (13.05%), and Oxysarcodexia diana (14.52%).

[Influence of dose regimen on gentamycin nephrotoxicity in rats]. / Interferência do intervalo de administração da droga sobre a nefrotoxicidade da gentamicina em ratos.

The acute renal failure (ARF), that still presents a right mortality rate (50%) can be defined as an abrupt decline of the glomerular filtration, resultant of ischemic or toxicity event. The drugs nephrotoxicity is one of the most frequent cause (27%) of ARF and it is suggested that the interval of administration of the drug can interfere in this side effect, however the best administration regimen is not very well established. This study evaluated the renal function of rats that received gentamicin (100 mg/kg) in one dose or in two doses (2 x 50 mg/kg), by intraperitoneal infusion. The results obtained in this research, indicated that the single infusion of gentamicin determined smaller nephrotoxicity by the reduction of serum concentration of this drug in 24 hours, decreasing the intracellular accumulation of this gentamicin, which is one of the main cellular mechanisms of this renal injury. The single dose treatment regime, otherwise, shows advantages not only related to the nephrotoxicity effect, but also it is relevant to the cost and safety, which can be rationable factors in the administration of this drug.

Colonisation of leaf litter by aquatic invertebrates in an Atlantic Forest stream.

Riparian vegetation along streams in the Atlantic Forest in Brazil contributes to the formation of a highly heterogeneous leaf litter in streambeds. To investigate the structure and composition of the aquatic invertebrate community during the process of leaf decomposition of two plant species present along the banks of the stream studied, 21 plastic mesh bags containing 2.5g (dry weight) of leaf matter from each species (Alchornea glandulosa (Vell) and Cabralea canjerana End. and Poeppig), for a total of 5.0g, were placed in the streambed. Three bags were removed after 3, 6, 9, 12, 24, 48 and 96 days. The taxonomic density was negatively correlated with the remaining weight. The high density of collector organisms, such as Chironomidae, Oligochaeta and Amphipoda, on the last day of incubation, probably occurred due to the increased amount of fine organic matter in the more advanced decomposition stages. The highest α diversity (Shannon-Wiener) values were observed for the 3rd and 96th days of the experiment, while the ß diversity values showed that these days presented the highest variation in the taxonomic composition, thus presenting a different faunistic composition. This study showed that the trophic structure and composition of aquatic invertebrates changes during the decomposition of leaf litter. The faunistic abundance and diversity observed in this study indicate that the entrance of material from plants growing along streams provides favorable conditions for the colonisation and establishment of invertebrates in lower-order streams, and thus points to the need to preserve riparian vegetation.

Giant cell tumours of the small bones of the hands and feet: long-term results of 30 patients and a systematic literature review.

Giant cell tumours (GCTs) of the small bones of the hands and feet are rare. Small case series have been published but there is no consensus about ideal treatment. We performed a systematic review, initially screening 775 titles, and included 12 papers comprising 91 patients with GCT of the small bones of the hands and feet. The rate of recurrence across these publications was found to be 72% (18 of 25) in those treated with isolated curettage, 13% (2 of 15) in those treated with curettage plus adjuvants, 15% (6 of 41) in those treated by resection and 10% (1 of 10) in those treated by amputation. We then retrospectively analysed 30 patients treated for GCT of the small bones of the hands and feet between 1987 and 2010 in five specialised centres. The primary treatment was curettage in six, curettage with adjuvants (phenol or liquid nitrogen with or without polymethylmethacrylate (PMMA)) in 18 and resection in six. We evaluated the rate of complications and recurrence as well as the factors that influenced their functional outcome. At a mean follow-up of 7.9 years (2 to 26) the rate of recurrence was 50% (n = 3) in those patients treated with isolated curettage, 22% (n = 4) in those treated with curettage plus adjuvants and 17% (n = 1) in those treated with resection (p = 0.404). The only complication was pain in one patient, which resolved after surgical removal of remnants of PMMA. We could not identify any individual factors associated with a higher rate of complications or recurrence. The mean post-operative Musculoskeletal Tumor Society scores were slightly higher after intra-lesional treatment including isolated curettage and curettage plus adjuvants (29 (20 to 30)) compared with resection (25 (15 to 30)) (p = 0.091). Repeated curettage with adjuvants eventually resulted in the cure for all patients and is therefore a reasonable treatment for both primary and recurrent GCT of the small bones of the hands and feet.

Sudan Black B treatment reduces autofluorescence and improves resolution of in situ hybridization specific fluorescent signals of brain sections.

Interference by autofluorescence is one of the major concerns of immunofluorescence analysis of in situ hybridization-based diagnostic assays. We present a useful technique that reduces autofluorescent background without affecting the tissue integrity or direct immunofluorescence signals in brain sections. Using six different protocols, such as ammonia/ethanol, Sudan Black B (SBB) in 70% ethanol, photobleaching with UV light and different combinations of them in both formalin-fixed paraffin-embedded and frozen human brain tissue sections, we have found that tissue treatment of SBB in a concentration of 0.1% in 70% ethanol is the best approach to reduce/eliminate tissue autofluorescence and background, while preserving the specific fluorescence hybridization signals. This strategy is a feasible, non-time consuming method that provides a reasonable compromise between total reduction of the tissue autofluorescence and maintenance of specific fluorescent labels.