Silencing of the Laccase (lacc2) Gene from Pleurotus ostreatus Causes Important Effects on the Formation of Toxocyst-like Structures and Fruiting Body.

A wide variety of biological functions, including those involved in the morphogenesis process of basidiomycete fungi, have been attributed to laccase enzymes. In this work, RNA interference (RNAi) was used to evaluate the role of the laccase (lacc2) gene of Pleurotus ostreatus PoB. Previously, transformant strains of P. ostreatus were obtained and according to their level of silencing they were classified as light (T7), medium (T21) or severe (T26 and T27). The attenuation of the lacc2 gene in these transformants was determined by RT-PCR. Silencing of lacc2 resulted in a decrease in laccase activity between 30 and 55%, which depended on the level of laccase expression achieved. The silenced strains (T21, T26, and T27) displayed a delay in the development of mycelium on potato dextrose agar (PDA) medium, whereas in the cultures grown on wheat straw, we found that these strains were incapable of producing aerial mycelium, primordia, and fruiting bodies. Scanning electron microscopy (SEM) showed the presence of toxocyst-like structures. The highest abundance of these structures was observed in the wild-type (PoB) and T7 strains. However, the abundance of toxocysts decreased in the T21 and T26 strains, and in T27 they were not detected. These results suggest that the presence and abundance of toxocyst-like structures are directly related to the development of fruiting bodies. Furthermore, our data confirm that lacc2 is involved in the morphogenesis process of P. ostreatus.

Laccase Production from Agrocybe pediades: Purification and Functional Characterization of a Consistent Laccase Isoenzyme in Liquid Culture.

Laccases are valuable enzymes as an excellent ecological alternative for bioremediation issues because they can oxidize persistent xenobiotic compounds. The production and characterization of extracellular laccases from saprotrophic fungi from disturbed environments have been scarcely explored, even though this could diversify their functional characteristics and expand the conditions in which they carry out their catalysis. Agrocybe pediades, isolated from a disturbed forest, produces an extracellular laccase in liquid culture. The enzyme was purified, identified and characterized. Copper and hexachlorobenzene do not function as inducers for the laccase produced. Partial amino acid sequences were obtained by LC-MS/MS that share similarity with laccases from other fungi. Purified laccase is a monomer with a molecular mass between 55-60 kDa and had an optimum activity at pH 5.0 and the optimum temperature at 45 °C using 2,6-dimethoxyphenol (2,6-DMP) as substrate. The Km and Vmax also determined with 2,6-DMP were 100 µM and 285 µmol∙min-1∙mg-1, respectively, showing that the laccase of A. pediades has a higher affinity for this substrate than that of other Agaricales. These features could provide a potential catalyst for different toxic substrates and in the future laccase could be used in environmental recovery processes.

A review of the reproductive system in anuran amphibians.

Reproductive biology is an important topic that is well explored in many vertebrates, but information about frogs' reproductive mechanisms could be improved. Therefore, this review aims to provide organized and specific information on frog reproduction. First, we developed schemes that illustrate the general information regarding reproductive biological mechanisms in frogs in a specific way. Then, we described the physiological, histological, and morphological mechanisms of each organ of the reproductive system of male and female frogs. Finally, this manuscript may contribute to a broader understanding of anuran reproductive biology. Since, understanding frogs' reproductive system permits one to make a comparison with reproduction with other anurans.

The pollution effect on the denitrifying capacity of methanogenic sludges.

The denitrification process has been studied for biodegradation of some emerging contaminants (ECs). For this, anaerobic sludges from different Wastewater Treatment Plants (WTP) have been used; however, the biodegradation capacity can differ due to the contact they have had with various pollutants, given their origin. This work aims to evaluate the kinetic and metabolic capacity of two denitrifying sludges from different WTPs to biodegrade CH3COO--C and NO3--N. Denitrifying tests were carried out in batches with CH3COO--C (30 mg L-1) in a CN-1 relationship of 1.8 with sludge from a WTP of an educational center (WTP-A) and CH3COO--C (50 mg L-1) to a CN-1 of 1.4 with another from the WTP of Atotonilco de Tula, Hidalgo, México (WTP-B). The results showed that the biodegradation rate of CH3COO--C and NO3--N with the WTP-B sludge was 35 and 75% greater, respectively, compared to the WTP-A sludge. Therefore, we suggest that the consumption difference of substrate is attributable to the sludges of WTP, which have been exposed to a high concentration of a great variety of pollutants.

Biotransformation of 17ß-Estradiol through a Denitrifying Sludge.

17ß-estradiol (E2) is the natural estrogen with the most significant potential for endocrine disruption in the biota of aquatic ecosystems at trace concentrations. It is, therefore, essential to study treatments for water polluted with E2 that would guarantee its complete elimination and mineralization. Denitrification is a biological process shown to have the capacity to completely biodegrade drugs, such as ampicillin. This work is aimed to evaluate the biotransformation of 17ß-estradiol by employing a denitrifying sludge. The assays performed were: (I) abiotic with 3.5 mg E2-C L-1 and (II) denitrifying with 10 mg CH3COO--C L-1 as the reference, 10 mg E2-C L-1 as the sole electron donor, and a mixture of (mg L-1) 10 E2-C with 10 CH3COO--C at C N-1 of 1.1. The E2-C and NO3--N consumption efficiencies were greater than 99%, and HCO3--C and N2 production yields were close to 1 in all assays. The denitrifying sludge could biodegrade up to 10 mg E2-C L-1 as the sole electron donor and when mixed with 10 mg CH3COO--C L-1. No intermediate metabolites were generated from the process.

Metabolomics: A New Approach in the Evaluation of Effects in Human Beings and Wildlife Associated with Environmental Exposition to POPs.

Human beings and wild organisms are exposed daily to a broad range of environmental stressors. Among them are the persistent organic pollutants that can trigger adverse effects on these organisms due to their toxicity properties. There is evidence that metabolomics can be used to identify biomarkers of effect by altering the profiles of endogenous metabolites in biological fluids or tissues. This approach is relatively new and has been used in vitro studies mainly. Therefore, this review addresses those that have used metabolomics as a key tool to identify metabolites associated with environmental exposure to POPs in wildlife and human populations and that can be used as biomarkers of effect. The published results suggest that the metabolic pathways that produce energy, fatty acids, and amino acids are commonly affected by POPs. Furthermore, these pathways can be promoters of additional effects. In the future, metabolomics combined with other omics will improve understanding of the origin, development, and progression of the effects caused by environmental exposure.

Differential expression and immunoreactivity of thyroid hormone transporters MCT8 and OATP1C1 in rat ovary.

Thyroid hormones (THs) regulate several physiological processes in female mammals, many of which are related to reproduction such as steroidogenesis in the ovary, oocyte and granulosa cells maturation, follicular development and differentiation, and ovulation. THs actions require the presence of THs transporters to facilitate their cellular uptake and efflux. MCT8 and OATP1C1 are the principal THs transporters. The aim of the present study was to determine the gene expression and cellular localization of MCT8 and OATP1C1 in the rat ovary during the diestrus-II cycle phase. Ovaries of virgin adult rats were histologically processed. Reverse Transcription-PCR and immunohistochemistry analyses for MCT8 and OATP1C1 were done. MCT8 gene expression level was significantly higher (P ≤ 0.01) than that of OATP1C1 in the rat ovary. MCT8 and OATP1C1 were found in all types of ovarian cells but with different immunoreactivity. MCT8 showed stronger immunoreactivity in tertiary and Graafian follicles, corpus luteum and blood vessels, whereas OATP1C1's immunoreactivity was stronger in stroma cells, tunica albuginea, and blood vessels. Our results provide evidence that THs and their transporters are both necessary for ovarian function and that any alteration in these transporters could interfere with reproductive processes such as ovulation and steroidogenesis, compromising fertility.

Genotoxicological response of the common carp (Cyprinus carpio) exposed to spring water in Tlaxcala, México.

This study evaluated the genotoxic impact of anthropic activities in Huactzinco Spring, using Cyprinus carpio as a biomonitor. In situ and in vivo experimental designs were compared by means of simultaneous 2-week exposures. The water from the spring generated mean micronuclei frequency values (108.6 ± 32 MN/1,000) and DNA fragmentation values (143.4 ± 35 au) which were statistically higher than those for the negative control (10.9 ± 6 MN/1,000 and 67.6 ± 23 au). The in situ and in vivo experiments supported one another. The comet assay proved to be the most sensitive test, with an EC50 value (11.4 % ± 3.4 %) being less than that determined for the micronuclei test (54.8 % ± 3.2 %). The results of this study confirm the usefulness of C. carpio as an environmental contamination biomonitor, and suggest that Huactzinco Spring water constitutes a latent risk to human health and the environment.

Antigenotoxic potential of aqueous extracts from the chanterelle mushroom, Cantharellus cibarius (higher Basidiomycetes), on human mononuclear cell cultures.

Cantharellus cibarius is one of the most important wild, edible, and ectomycorrhizal mushrooms growing at La Malinche National Park, Tlaxcala, Mexico; therefore, the assessment of its biological properties is of great interest to know its potential as an alternative treatment to chemopreventive strategies when it is consumed as part of a diet. Comet assay was used to evaluate the antigenotoxic properties of several concentrations of aqueous extracts (0.0125, 0.025, 0.05, 0.1, and 0.2% w/v) prepared at room temperature (22 ± 2°C). As a test system we used human mononuclear cells exposed to methyl methanesulphonate (MMS) in vitro according to 3 different protocols: previous, simultaneous, and posterior. Previous (0.0125%) and simultaneous (0.1%) treatments resulted in the highest inhibitory efficiency. In the former, the cells assessed showed a tail length of 94.9 ± 64 µm; in the latter, the tails measured 106.2 ± 40 µm. Resulting percentages of reduction in damage were 236% and 196.1%, respectively. We did not obtain a dose-dependent response. The mean tail length for each protocol (previous, 133.1 ± 80 µm; simultaneous, 127.8 ± 57 µm; posterior, 146.3 ± 74 µm) was statistically significant with regard to the positive control (MMS).

Exposición infantil al plomo en sitios contaminados / Children exposure to lead in contaminated sites

OBJETIVO: Determinar el grado de la exposición infantil al plomo en diversos tipos de sitios contaminados. MATERIAL Y MÉTODOS: El estudio se realizó de junio 2008 a diciembre 2009 en cuatro sitios de México: metalúrgica de Ávalos, Chihuahua.; metalúrgica de Morales, San Luis Potosí (SLP); zona alfarera en La Trinidad, Tlaxcala, y sitio minero en Cedral, SLP. Se cuantificó plomo en polvo y se realizó un biomonitoreo humano en niños de la comunidad. RESULTADOS: Los valores obtenidos de plomo en polvo exterior superaron el límite establecido de 400 mg/kg para suelos residenciales en un intervalo de valores para los cuatro sitios de 62 a 5 187 mg/kg. En cuanto al monitoreo biológico, todas las poblaciones presentaron valores extremos, desde los 22 µg/dL en Cedral, 31 µg/dL en Morales, y 32 µg/dL en Ávalos, hasta los 52 µg/dL en La Trinidad. Es importante señalar que encontramos una correlación positiva y significativa entre los valores de plomo en polvo y plomo en sangre en todos los sitios de estudio (p<0.001). CONCLUSIÓN: Estos sitios son un ejemplo de los riesgos en salud relacionados con la exposición a plomo en México; por consiguiente, se requiere de un programa nacional de salud pública dirigido a reducir la exposición a este metal en poblaciones vulnerables.

OBJECTIVE: To assess the exposure to lead in children living in various types of contaminated sites. MATERIALS AND METHODS: The study was conducted from June 2008 to December 2009 at four sites in Mexico: Avalos metallurgical, Chihuahua; Morales metallurgical, San Luis Potosí (SLP); Trinidad pottery area, Tlaxcala and Cedral mine site, SLP. These sites contain different sources of lead. The metal levels were quantified in outdoor dust and in peripheral blood of children. RESULTS: Lead dust concentrations exceed the National Guidelines for residential soils (400 mg/kg) in a range of values for the four sites from 62 to 5 187 mg/kg. Regarding biological monitoring, the studied children showed maximum lead blood levels of 22 µg/dL in Cedral, 31 µg/dL in Morales, 32 µg/dL in Avalos, and 52 µg/dL in Trinidad. It is important to mention that in all the studied sites, a significative positive correlation was found between blood lead levels and the lead concentrations in dust. CONCLUSION: These sites are an example of the health risks related to lead exposure in Mexico; therefore, there is an urgent need for a national public health program aimed at reducing lead exposure in vulnerable populations.

[Children exposure to lead in contaminated sites]. / Exposición infantil al plomo en sitios contaminados.

OBJECTIVE: To assess the exposure to lead in children living in various types of contaminated sites. MATERIALS AND METHODS: The study was conducted from June 2008 to December 2009 at four sites in Mexico: Avalos metallurgical, Chihuahua; Morales metallurgical, San Luis Potosí (SLP); Trinidad pottery area, Tlaxcala and Cedral mine site, SLP. These sites contain different sources of lead. The metal levels were quantified in outdoor dust and in peripheral blood of children. RESULTS: Lead dust concentrations exceed the National Guidelines for residential soils (400 mg/kg) in a range of values for the four sites from 62 to 5 187 mg/kg. Regarding biological monitoring, the studied children showed maximum lead blood levels of 22 µg/dL in Cedral, 31 µg/dL in Morales, 32 µg/dL in Avalos, and 52 µg/dL in Trinidad. It is important to mention that in all the studied sites, a significative positive correlation was found between blood lead levels and the lead concentrations in dust. CONCLUSION: These sites are an example of the health risks related to lead exposure in Mexico; therefore, there is an urgent need for a national public health program aimed at reducing lead exposure in vulnerable populations.

DNA damage in blood cells from children exposed to arsenic and lead in a mining area.

In this work, we studied the frequency of DNA damage in children living in Villa de la Paz, Mexico, a mining site contaminated with arsenic and lead. DNA damage in blood cells was assessed using the Comet assay, and the results were compared to those found in children living in a less exposed town (Matehuala). In Villa de la Paz, high concentrations of arsenic and lead in surface soil and household dust were found. All of the soil samples had concentrations above 100 mg/kg of arsenic, and 58% of the samples were higher than 400 mg/kg of lead (these concentrations are used as intervention guidelines by the United States Environmental Protection Agency). In agreement with the environmental results, urinary arsenic in children living in Villa de la Paz (geometric mean 136 microg/g creatinine) was significantly higher than that found in children living in Matehuala (34 microg/g creatinine). Blood lead levels were also significantly higher in children from Villa de la Paz (11.6 microg/dL) than in children from Matehuala (8.3 microg/dL). The results of the Comet assay showed that the tail length and the tail moment in children living in Villa de la Paz were higher than those observed for children in Matehuala (P<0.05). Taking all the data into account, our study has shown increased DNA damage in children exposed to arsenic and lead in the mining site of Villa de la Paz.

Overview of human health and chemical mixtures: problems facing developing countries.

In developing countries, chemical mixtures within the vicinity of small-scale enterprises, smelters, mines, agricultural areas, toxic waste disposal sites, etc., often present a health hazard to the populations within those vicinities. Therefore, in these countries, there is a need to study the toxicological effects of mixtures of metals, pesticides, and organic compounds. However, the study of mixtures containing substances such as DDT (dichlorodiphenyltrichloroethane, an insecticide banned in developed nations), and mixtures containing contaminants such as fluoride (of concern only in developing countries) merit special attention. Although the studies may have to take into account simultaneous exposures to metals and organic compounds, there is also a need to consider the interaction between chemicals and other specific factors such as nutritional conditions, alcoholism, smoking, infectious diseases, and ethnicity.