Induction of mitotic crossing-over in diploid strains of Aspergillus nidulans using low-dose X-rays

As a contribution towards detecting the genetic effects of low doses of genotoxic physical agents, this paper deals with the consequences of low-dose X-rays in the Aspergillus nidulans genome. The irradiation doses studied were those commonly used in dental clinics (1-5 cGy). Even very low doses promoted increased mitotic crossing-over frequencies in diploid strains heterozygous for several genetic markers including the ones involved in DNA repair and recombination mechanisms. Genetic markers of several heterozygous strains were individually analyzed disclosing that some markers were especially sensitive to the treatments. These markers should be chosen as bio-indicators in the homozygotization index assay to better detect the recombinogenic/ carcinogenic genomic effects of low-dose X-rays. ©FUNPEC-RP.

In Vivo Genotoxicity Assessment of Titanium Dioxide Nanoparticles by Allium cepa Root Tip Assay at High Exposure Concentrations

The industrial production and commercial applications of titanium dioxide nanoparticles have increased considerably in recent times, which has increased the probability of environmental contamination with these agents and their adverse effects on living systems. This study was designed to assess the genotoxicity potential of TiO2 NPs at high exposure concentrations, its bio-uptake, and the oxidative stress it generated, a recognised cause of genotoxicity. Allium cepa root tips were treated with TiO2 NP dispersions at four different concentrations (12.5, 25, 50, 100 mu g/mL). A dose dependant decrease in the mitotic index (69 to 21) and an increase in the number of distinctive chromosomal aberrations were observed. Optical, fluorescence and confocal laser scanning microscopy revealed chromosomal aberrations, including chromosomal breaks and sticky, multipolar, and laggard chromosomes, and micronucleus formation. The chromosomal aberrations and DNA damage were also validated by the comet assay. The bio-uptake of TiO2 in particulate form was the key cause of reactive oxygen species generation, which in turn was probably the cause of the DNA aberrations and genotoxicity observed in this study.

Components of resistance to Fusarium head blight in wheat detected in a seed germination assay with Microdochium majus and the relationship to FHB disease development and mycotoxin accumulation from Fusarium graminearum infection.

Components of partial disease resistance (PDR) to fusarium head blight (FHB), detected in a seed-germination assay, were compared with whole-plant FHB resistance of 30 USA soft red winter wheat entries in the 2002 Uniform Southern FHB Nursery. Highly significant (P <0·001) differences between cultivars in the in vitro seed-germination assay inoculated with Microdochium majus were correlated to FHB disease incidence (r = -0·41; P <0·05), severity (r = -0·47; P <0·01), FHB index (r = -0·46; P <0·01), damaged kernels (r = -0·52; P <0·01), grain deoxynivalenol (DON) concentration (r = -0·40; P <0·05) and incidence/severity/kernel-damage index (ISK) (r = -0·45; P <0·01) caused by Fusarium graminearum. Multiple linear regression analysis explained a greater percentage of variation in FHB resistance using the seed-germination assay and the previously reported detached-leaf assay PDR components as explanatory factors. Shorter incubation periods, longer latent periods, shorter lesion lengths in the detached-leaf assay and higher germination rates in the seed-germination assay were related to greater FHB resistance across all disease variables, collectively explaining 62% of variation for incidence, 49% for severity, 56% for F. graminearum-damaged kernels (FDK), 39% for DON and 59% for ISK index. Incubation period was most strongly related to disease incidence and the early stages of infection, while resistance detected in the seed germination assay and latent period were more strongly related to FHB disease severity. Resistance detected using the seed-germination assay was notable as it related to greater decline in the level of FDK and a smaller reduction in DON than would have been expected from the reduction in FHB disease assessed by visual symptoms.

Enzyme reactions as index of freshness of Fish and Shellfish

An attempt has been made in this study to screen some fish muscle enzymes to assess their potential worth in testing the degree of freshness of fish. A problem with routine enzyme activity determinations is the complexity of the method of enzyme assay. Hence, in the present study as far as possible simple assay techniques were adopted. Several species were screened to assess the possibility of employing this procedure on a large scale. It is hoped that findings of this study will lead to the development of meaningful criteria in testing the freshness of fish. This thesis has been divided into five chapters

Capillary assay device with internal hydrophilic coating

The present invention provides assay devices having a unitary body with an exterior surface, the unitary body being substantially transparent to visible light and formed from a material having a refractive index in the range 1.26 to 1.40, the refractive index being measured at 20 °C with light of wavelength 589 nm, and wherein the unitary body is formed from a hydrophobic material, and at least two capillary bores extending internally along the unitary body, wherein at least a portion of the surface of each capillary bore includes a hydrophilic layer for retaining an assay reagent, and wherein the hydrophilic layer is also substantially transparent to visible light to allow optical interrogation of the capillary bores through the capillary wall. The present invention also provides assay systems including such assay devices, methods of performing an assay using such assay devices and method of method for manufacturing such assay devices.

Genotoxicity of Brosimum gaudichaudii measured by the Salmonella/microsome assay and chromosomal aberrations in CHO cells

The root bark of Brosimum gaudichaudii Trecul (Moraceae) is popularly used for treatment of vitiligo. In the present study the mutagenic activity of the aqueous and methanolic extract as well as of the n-butanolic fraction of this medicinal plant were evaluated using Salmonella typhimurium assays, TA100, TA98, TA102 and TA97a strains, while the clastogenic effect in Chinese hamster ovary (CHO) cells in the G(1)/S, S and G(2)/S phases of the cell cycle. The results showed mutagenic activity of the aqueous extract against TA102 in the presence of S9, and of methanolic extract, with and without metabolic activation. TA100 mutagenicity was only observed for the methanolic extract in the absence of S9. The n-butanolic fraction did not present mutagenic activity. In CHO cells only the methanolic extract induced a significant increase of chromosomal aberrations in the G(1)/S and S phases, whereas a decrease in the mitotic index was observed in the G(1)/S and G(2)/S phases. No clastogenicity was observed for the aqueous extract. The furocoumarins (psoralen and bergapten) presented in the extracts might contribute to the mutagenicity. The lower activity of the aqueous extract was probably due to the presence of smaller amount of furocoumarins compared to the methanolic extract. (C) 2002 Elsevier B.V. Ireland Ltd. All rights reserved.

Lymphocyte transformation assay for C neoformans antigen is not reliable for detecting cellular impairment in patients with Neurocryptococcosis

Background: Cryptococcus neoformans causes meningitis and disseminated infection in healthy individuals, but more commonly in hosts with defective immune responses. Cell-mediated immunity is an important component of the immune response to a great variety of infections, including yeast infections. We aimed to evaluate a specific lymphocyte transformation assay to Cryptococcus neoformans in order to identify immunodeficiency associated to neurocryptococcosis (NCC) as primary cause of the mycosis.Methods: Healthy volunteers, poultry growers, and HIV-seronegative patients with neurocryptococcosis were tested for cellular immune response. Cryptococcal meningitis was diagnosed by India ink staining of cerebrospinal fluid and cryptococcal antigen test (Immunomycol-Inc, SP, Brazil). Isolated peripheral blood mononuclear cells were stimulated with C. neoformans antigen, C. albicans antigen, and pokeweed mitogen. The amount of H-3-thymidine incorporated was assessed, and the results were expressed as stimulation index (SI) and log SI, sensitivity, specificity, and cut-off value (receiver operating characteristics curve). We applied unpaired Student t tests to compare data and considered significant differences for p<0.05.Results: The lymphotoxin alpha showed a low capacity with all the stimuli for classifying patients as responders and non-responders. Lymphotoxin alpha stimulated by heated-killed antigen from patients with neurocryptococcosis was not affected by TCD4+ cell count, and the intensity of response did not correlate with the clinical evolution of neurocryptococcosis.Conclusion: Response to lymphocyte transformation assay should be analyzed based on a normal range and using more than one stimulator. The use of a cut-off value to classify patients with neurocryptococcosis is inadequate. Statistical analysis should be based on the log transformation of SI. A more purified antigen for evaluating specific response to C. neoformans is needed.

Detection of the single nucleotide polymorphism (rs2227307) in the human interleukin 8 gene using a pcr-rflp assay

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Study of Salmonella typhimurium mutagenicity assay of (E)-piplartine by the Ames test

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Mutagenic activity in waste from an aluminum products factory in Salmonella/microsome assay

The mutagenic activity of waste material originating from an aluminum products factory was determined by the Salmonella/microsome assay, using the bacterial strains TA100, TA98 and YG1024. The material was obtained by sweeping the factory floor at the end of the work shift. Organic compounds were extracted by ultrasound for 30 min in dichloromethane or 70% ethanol. After evaporation of solvent, these extracts were dissolved in dimethylsulfoxide, and tested for the mutagenic activity at varying concentrations. All the extracts from the factory had mutagenic activity, especially in the YG1024 strain, suggesting the presence of aromatic amines, later confirmed by chemical analysis. The TA98 strain also showed mutagenic activity, though it did not exhibit the highest mutagenicity index observed with the YG1024 strain. In TA100, mutagenic activity was not observed. This study should serve as an alert to management and those who are occupationally exposed, and as a warning that this type of waste should not be discarded in the environment without any control. (C) 2004 Elsevier Ltd. All rights reserved.

In vivo clastogenicity assessment of the Austroplenckia populnea (Celastraceae) leaves extract using micronucleus and chromosome aberration assay

The clastogenic effect of the A. populnea leaves extract was tested in vivo on bone marrow cells of Wistar rats by evaluating the induction of chromosome aberrations and micronuclei induction on polychromatic erythrocytes. The extract was administered by gavage at doses of 300, 600 and 900mg/kg body weight. Experimental and control animals were submitted to euthanasia 24 h after the treatment. Under the conditions used, A. populnea leaves extract did not induce decrease in mitotic index and did not induce a statistically significant increase in the mean number of micronucleated polychromatic erythrocytes or chromosome aberrations in the bone marrow cells of Wistar rats. © 2007 The Japan Mendel Society.

Padronização e aplicação da técnica de ELISA (Enzyme-linked immunosorbent assay) indireto com anticorpo de captura para a detecção de anticoepos contra o cicovírus suíno tipo 2

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Improvement of an enzyme immunosorbent assay for detecting antibodies against Dioctophyma renale

An available enzyme-linked immunosorbent assay (ELISA) was studied for the detection of anti-Dioctophyma renale antibodies in the sera of dogs using, detection of parasite eggs in urine sediment as a reference test. ELISA uses a soluble antigenic preparation of esophagus of D. renale and the optimal dilutions of the antigen, serum and conjugate were determined by means of checker board titration, using positive (n=13) and negative (n=27) reference serum. The specificity and sensitivity of the ELISA were 93.8% and 92.3% respectively and the kappa index was good (0.76). These results suggest that ELISA described may prove to be an effective serological test for detecting dogs infected and exposed to this parasite mainly dogs that are not eliminating parasite eggs through their urine.

Fluorescence polarization assay, competitive enzyme-linked immunosorbent assay (ELISA-C) and indirect ELISA for the diagnosis of brucellosis in buffaloes (Bubalus bubalis)

The objective of the present study was to compare the performance of three serological tests for diagnosis of Brucella abortus infections in buffaloes (Bubalus bubalis). Serum samples collected from 696 adult females were submitted to the competitive enzyme-linked immunosorbent assay (ELISAC), (I-ELISA), fluorescence polarization test (FPA), 2-mercaptoethanol test (2-ME) and complement fixation test (CFT). The gold standard was the combination of CFT and 2-ME, considering as positive the reactors in both CFT and 2-ME, and as negative those non-reactors. ROC analyses were done for C-ELISA, I-ELISA and FPA and the Kappa agreement index were also calculated. The best combinations of relative sensitivity (SEr) and relative specificity (SPr) and Kappa were given by C-ELISA (96.9%, 99.1%, and 0.932, respectively) and FPA (92.2%, 97.6 and 0.836, respectively). The C-ELISA and FPA were the most promising confirmatory tests for the serological diagnosis of brucellosis in buffaloes, and for these tests, cut-off values for buffaloes may be the same as those used for bovines.