Role of ATRX in chromatin structure and function: implications for chromosome instability and human disease

Functional differentiation of chromatin structure is essential for the control of gene expression, nuclear architecture, and chromosome stability. Compelling evidence indicates that alterations in chromatin remodeling proteins play an important role in the pathogenesis of human disease. Among these, α-thalassemia mental retardation X-linked protein (ATRX) has recently emerged as a critical factor involved in heterochromatin formation at mammalian centromeres and telomeres as well as facultative heterochromatin on the murine inactive X chromosome. Mutations in human ATRX result in an X-linked neurodevelopmental condition with various degrees of gonadal dysgenesis (ATRX syndrome). Patients with ATRX syndrome may exhibit skewed X chromosome inactivation (XCI) patterns, and ATRX-deficient mice exhibit abnormal imprinted XCI in the trophoblast cell line. Non-random or skewed XCI can potentially affect both the onset and severity of X-linked disease. Notably, failure to establish epigenetic modifications associated with the inactive X chromosome (Xi) results in several conditions that exhibit genomic and chromosome instability such as fragile X syndrome as well as cancer development. Insight into the molecular mechanisms of ATRX function and its interacting partners in different tissues will no doubt contribute to our understanding of the pathogenesis of ATRX syndrome as well as the epigenetic origins of aneuploidy. In turn, this knowledge will be essential for the identification of novel drug targets and diagnostic tools for cancer progression as well as the therapeutic management of global epigenetic changes commonly associated with malignant neoplastic transformation.     

A multiplex RTi-PCR reaction for simultaneous detection of Escherichia coli O157:H7, Salmonella spp. and Staphylococcus aureus on fresh, minimally processed vegetables

In this work, a new multiplex single-tube real-time PCR approach is presented for the detection of Escherichia coli O157:H7, Salmonella spp. and Staphylococcus aureus, three of the more frequent food-borne bacterial pathogens that are usually investigated in a variety of food matrices. The study includes the design and specificity testing, of a new primer and probe specific for Salmonella spp. Reaction conditions were adjusted for the simultaneous amplification and detection of specific fragments in the beta-glucuronidase (uidA, E. coli) and Thermonulease (nuc, Sta. aureus) genes, and in the replication origin sequence (oriC, Salmonella spp.). Melting-curve analysis using a SYBR Green I RTi-PCR approach showed characteristic T(m) values demonstrating the specific and efficient amplification of the three fragments. Subsequently, a TaqMan RTi-PCR approach was settled, using FAM, NED and VIC fluorescently labelled specific probes for an automated detection. It was equally sensitive than uniplex RTi-PCR reactions in Sta. aureus and E. coli O157:H7, using same amounts of purified DNA, and allowed detection of 10 genome equivalents in the presence of 10(2) or 10(4) genome equivalents of the other two pathogens. Finally, it was tested in artificially inoculated fresh, minimally processed vegetables, revealing a sensitivity of 10(3)CFUg(-1) each of these pathogens in direct detection, following DNA extraction with DNeasy Tissue Kit (Qiagen). The multiplex RTi-PCR developed scored the sensitivity recognised for PCR in food and it allows a high-throughput and automation, thus it is promising as a rapid and cost-effective test for the food industry.     

Nonstarter lactobacilli isolated from soft and semihard Argentinean cheeses: genetic characterization and resistance to biological barriers

Nonstarter lactic acid bacteria isolated from Argentinean cheeses were identified and characterized by focusing on their resistance to biological barriers, along with other physiological features of potential interest, in the search for future probiotic organisms. Lactobacilli were enumerated and isolated from semihard and soft cheeses made with multistrain Streptococcus thermophilus starters. Lactobacilli counts in 1-week-old cheeses were between 10(5) and 10(7) CFU/g and then reached 10(7) CFU/ g in all 1-month samples, while streptococci were always above 10(9) CFU/g. A total number of 22 lactobacilli isolates were retained, identified, and characterized by in vitro tests. Species identity was determined by carbohydrate metabolism and species-specific PCR assays. Genetic diversity was explored by random amplified polymorphic DNA (RAPD) PCR analysis. The Lactobacillus strains were assigned to the species L. casei, L. plantarum, L. rhamnosus, L. curvatus, L. fermentum, and L. perolens. All the strains studied tolerated 25 ppm of lysozyme, and most of them showed resistance to 0.3% bile. After incubation in gastric solution (pH 2.0), counts decreased by several log units, ranging from 3.2 to 7.0. The strains were able to grow in the presence of bile salts, but only three isolates were capable of deconjugation. The nonstarter lactobacilli that were assayed fermented the prebiotic substrates (especially lactulose and inulin). Some strains showed high cell hydrophobicity and beta-galactosidase activity, as well as inhibitory activity against pathogenic bacteria. It was concluded that most of the lactobacilli isolated in this study demonstrated resistance to biological barriers and physiological characteristics compatible with probiotic properties, which make them suitable for further research in in vivo studies aimed at identifying new probiotic organisms.     

Hydrogen Peroxide and Calcium Chloride Added to Irrigation Water as a Strategy to Reduce Bacterial Populations and Improve Quality of Fresh Mushrooms

The quality and value of fresh mushrooms are often diminished by the presence of high bacterial populations that cause a brown, blotchy appearance. The objective of the present research was to evaluate the addition of hydrogen peroxide and/or calcium chloride to irrigation water as a means to reduce total bacterial populations on fresh mushrooms. Crops were grown using commercial mushroom growing practices except for the addition of 0.75% hydrogen peroxide and/or 0.3% calcium chloride irrigation water added to the crop starting 11 d after the casing layer was applied on top of mushroom compost. Irrigation water without the added treatments acted as the control. Mushrooms were aseptically sampled from the production beds for enumerating bacterial counts. Total aerobic bacterial populations were determined by standard microbiological plating procedures. Mushroom whiteness (L‐value) and color (delta E) after harvest and postharvest storage were measured using a Minolta chromameter. Harvested mushrooms were separated by treatment and weighed to record yield. Mushrooms irrigated with water (control) had 7.3 log colony‐forming units (CFU) of aerobic bacterial populations per gram of fresh mushroom tissue. Compared with the control, irrigation with 0.75% hydrogen peroxide and 0.3% calcium chloride reduced the bacterial populations on fresh mushrooms by 87% (6.4 log CFU/g). Irrigation with hydrogen peroxide and calcium chloride significantly enhanced mushroom whiteness after harvest as well as after 6 d of postharvest storage at 12 °C. The irrigation treatments did not have a significant effect on crop yields; hence, the addition of hydrogen peroxide and calcium chloride to irrigation water was demonstrated to have good potential as a practical strategy to reduce bacterial populations and to improve the quality of fresh mushrooms.     

An overview of small-scale food fermentation technologies in developing countries with special reference to Thailand: scope for their improvement

Small-scale food fermentation technologies in developing countries have evolved through years of experience rather than through scientific breakthroughs. Many small-scale manufacturers are, therefore, reluctant to accept change and modify fermentation processes. Upgrading the quality and safety of fermented foods, while reducing their production cost and maintaining their authenticity and uniqueness, is of utmost importance. One strategic approach, which has been successfully implemented for the improvement of small-scale soy sauce fermentation in Thailand, is the consortium approach. This approach has allowed industry to work closely in sharing knowledge and common problems, which in turn has provided scientists with the research direction that would best benefit the industry. This consortium approach has brought about changes in the methodologies used in the production of soy sauce, by shortening processing times, introducing fiberglass tanks as bioreactors instead of the traditionally used small earthenware containers and introducing cost-effective waste management systems. One barrier to the application of starter cultures in the small-scale fermentation industry is the loss of uniqueness of fermented products. However, the advent of molecular biology techniques has allowed science to tailor starter cultures to the specific requirements of the manufacturer. Using the techniques of molecular biology, it has been shown that microflora of a specific product vary according to origin and sensory quality. A cell bank is being developed to serve as a resource base for Thai fermented pork sausage in order to facilitate the application of starter cultures in the manufacture of that product.     

Low prevalence of Listeria monocytogenes in foods from Italy

Listeria monocytogenes is an important foodborne pathogen that causes gastrointestinal disorders, and, especially in immunocompromised people, serious extraintestinal diseases, such as septicemia and meningitis, as well as abortion in pregnant women. Many foods, from both plant and animal origin, have been involved in listeriosis outbreaks. This article reports the results of a 12-year survey (1993 through 2004) on the presence of L. monocytogenes in several kinds of food marketed in Italy. Of 5,788 analyzed samples, 121 (2.1%) were contaminated with L. monocytogenes. The highest prevalence was found in smoked salmon (10.6%) and in poultry meat samples (8.5%) and the lowest in red meat (0.3%). L. monocytogenes was not found in 154 samples of fresh seafood products. Fifty-two isolates were also serotyped by the agglutination method. The most common serotypes detected in the 52 strains tested were 1/2a (36.5%), followed by 1/2c (32.8%), 1/2b (13.5%), 4b (11.5%), 3a (3.8%), and 3b (1.9%). The results of the present study showed low levels of L. monocytogenes in the analyzed samples. A total of 61.5% of the 52 L. monocytogenes strains analyzed belonged to serotypes 1/2a, 1/2b, and 4b, namely the serovars that are most commonly involved in extraintestinal human listeriosis outbreaks. In the ready-to-eat samples, these three serotypes were 40.0% (1/2a), 17.1% (1/2b), and 14.3% (4b). This finding highlights the need to implement strict hygienic measures during the production, distribution, and sale of foods to reduce the risk of foodborne listeriosis in humans to an acceptable level.     

Processing, Nutritional Evaluation, and Utilization of Whole Mesquite Flour (Prosopis laevigata)

ABSTRACT:  Basic crops such as wheat and maize are not cultivated easily in the central highlands of Mexico, but pods of mesquite ( Prosopis laevigata ), a N₂−fixing plant that dominates the vegetation, could be used as an additional food source. The physicochemical and functional properties and nutritional value of whole mesquite pod flour dried at 60 °C and 70 °C and muffins prepared with the mesquite flour were investigated. Drying temperature did not change the true protein concentration of the whole mesquite pod flour, that is, 97%. The digestibility for pod toasted at 70 °C was 78% compared to 77% at 60 °C, tannin content was 58% and 48% while trypsin inhibitor was 347 and 495 unit of inhibition (UI)/g sample, respectively. Functional characteristics of whole mesquite pod flour were similar to those of bean flour (Phaseolus vulgaris) , and superior to those of whole wheat flour (Triticum sp ) . Mesquite proteins were high in tryptophan and histidine. Sensorial analysis scores for products made with whole mesquite flour were high. The results of this work suggested that whole mesquite pod flour could be used as a supplement for human consumption.     

Occurrence of fungicide and insecticide residues in trade samples of leafy vegetables

The aim of this work was to report on a total of 23 fungicides and insecticides residues in 75 green and leafy vegetables (Swiss chards, Spinaches and Lettuces) collected from Ourense (NW Spain) by Spring 2007. The pesticides in the study samples were extracted with acetonitrile; the extracts were then cleaned-up by solid-phase extraction and concentrated before determination by PTV (Programmable Temperature Vaporization Injector) – GC-ITMS. Use of analyte protectants mixtures provided the best results in terms of effective compensation for matrix-induced enhancement effect. Pesticide residues were determined above the maxima residue limits (MRL) in 15 of the 75 analyzed samples, with a total of 18 violations of the MRL (three of the samples did not fulfil with two different pesticide MRL). The highest concentrations of fungicides were found in lettuce (procymidone, 12 mg/kg) and the highest concentrations of insecticides were found in Swiss chard (cypermethrin, 6 mg/kg). More positives for fungicides were detected and at larger concentrations than insecticides, especially for lettuces. Accumulation of pesticides in lettuces is higher than for the other leafy vegetables. The findings of this study pointed to the following recommendation: the need for a monitoring program for residues of iprodione and procymidone, together with cypermethrin, in food crops at the national level.