Assessment of the working range and effect of sodium dichloroisocyanurate on Pseudomonas aeruginosa biofilms and planktonic cells

Sodium dichloroisocyanurate (NaDCC) is a chemical agent that acts against microorganisms in a manner similar to that of sodium hypochlorite by releasing free available chlorine. NaDCC has been approved by the WHO for the emergency treatment of water and by the US EPA for routine treatment of water. Previous studies assessing the effectiveness of NaDCC for the treatment of water implied that NaDCC should have a wide array of disinfecting effects beyond the treatment of planktonic cells in potable water. In this study the biocidal effects of NaDCC against Pseudomonas aeruginosa cells in different growth modes including planktonic cells and biofilms were explored. The data showed that a 60% dilution of the standard NaDCC solution was effective in the treatment of both P. aeruginosa planktonic cells and biofilms.     

Morning-evening preference: sleep pattern spectrum and lifestyle habits among Japanese junior high school pupils

We surveyed the sleep-wake patterns and lifestyle habits in a sample of Japanese first to third year junior high school children (n=638, age 12 to 15 yrs), of whom 29.3% were evening type, 64.1% intermediate type, and 6.6% morning type in preference. The morningness-eveningness (M-E) score was lower (more evening typed), 16.1 vs. 15.4 in first compared to third year students. There were significant gender differences, with girls showing a greater evening preference. Evening preference was associated with longer sleep latency, shortened sleep duration during schooldays and weekends, bad morning feeling, and episodes of daytime sleepiness. In contrast, morning preference was associated with higher sleep drive and better sleep-wake parameters and lifestyle habits. Our results suggest the morning preference should be promoted among junior high school children to increase the likelihood of more regular sleep-wake patterns and lifestyle habits.     

Carbon isotopes support the presence of extensive land floras pre-dating the origin of vascular plants

Multiple lines of evidence indicate that Earth's land masses became green some 2.7 Ga ago, about 1 billion years after the advent of life. About 2.2 billion years later, land plants abruptly appear in the fossil record and diversify marking the onset of ecologically complex terrestrial communities that persist to the present day. Given this long history of land colonization, surprisingly few studies report direct fossil evidence of emergent vegetation prior to the continuous record of life on land that starts in the mid-Silurian (ca. 420–425 Ma ago). Here we compare stable carbon isotope signatures of fossils from seven Ordovician–Silurian (450–420 Ma old) Appalachian biotas with signatures of coeval marine organic matter and with stable carbon isotope values predicted for Ordovician and Silurian liverworts (BRYOCARB model). The comparisons support a terrestrial origin for fossils in six of the biotas analyzed, and indicate that some of the fossils represent bryophyte-grade plants. Our results demonstrate that extensive land floras pre-dated the advent of vascular plants by at least 25 Ma. The Appalachian fossils represent the oldest direct evidence of widespread colonization of continents. These findings provide a new search image for macrofossil assemblages that contain the earliest stages of land plant evolution. We anticipate they will fuel renewed efforts to search for direct fossil evidence to track the origin of land plants and eukaryotic life on continents further back in geologic time.     

Carbonic anhydrase inhibitors. Synthesis of 2,4,6-trimethylpyridinium derivatives of 2-(hydrazinocarbonyl)-3-aryl-1H-indole-5-sulfonamides acting as potent inhibitors of the tumor-associated isoform IX and XII

A series of 2-(hydrazinocarbonyl)-3-aryl-1H-indole-5-sulfonamides possessing various 2-, 3- or 4- substituted phenyl groups with methyl-, halogeno- and methoxy-functionalities, or a perfluorophenyl moiety, has been derivatized by reaction with 2,4,6-trimethylpyrylium perchlorate. The new sulfonamides were evaluated as inhibitors of four mammalian carbonic anhydrase (CA, EC 4.2.1.1) isoforms, that is, CA I, II (cytosolic), CA IX and XII (transmembrane, tumor-associated forms). Excellent inhibitory activity was observed against hCA IX with most of these sulfonamides, and against hCA XII with some of the new compounds. These compounds were generally less effective inhibitors of hCA II. Being membrane impermeant, these positively-charged sulfonamides are interesting candidates for targeting the tumor-associated CA IX and XII, as possible diagnostic tools or therapeutic agents.     

Pronounced Effect of the Nature of the Inoculum on Biofilm Development in Flow Systems

Biofilm formation renders sessile microbial populations growing in continuous-flow systems less susceptible to variation in dilution rate than planktonic cells, where dilution rates exceeding an organism''s maximum growth rate (μ_max) results in planktonic cell washout. In biofilm-dominated systems, the biofilm''s overall μ_max may therefore be more relevant than the organism''s μ_max, where the biofilm μ_max is considered as a net process dependent on the adsorption rate, growth rate, and removal rate of cells within the biofilm. Together with lag (acclimation) time, the biofilm''s overall μ_max is important wherever biofilm growth is a dominant form, from clinical settings, where the aim is to prevent transition from lag to exponential growth, to industrial bioreactors, where the aim is to shorten the lag and rapidly reach maximum activity. The purpose of this study was to measure CO₂ production as an indicator of biofilm activity to determine the effect of nutrient type and concentration and of the origin of the inoculum on the length of the lag phase, biofilm μ_max, and steady-state metabolic activity of Pseudomonas aeruginosa PA01 (containing gfp), Pseudomonas fluorescens CT07 (containing gfp), and a mixed community. As expected, for different microorganisms the lengths of the lag phase in biofilm development and the biofilm μ_max values differ, whereas different nutrient concentrations result in differences in the lengths of lag phase and steady-state values but not in biofilm μ_max rates. The data further showed that inocula from different phenotypic origins give rise to lag time of different lengths and that this influence persists for a number of generations after inoculation.Microbial growth in batch cultures has been studied for a long time, and the observed phases have been designated the lag phase, the acceleration phase, the exponential phase, the retardation phase, the stationary phase, and the phase of decline although not each culture displays all of the mentioned phases (16). In contrast to batch cultures and static (no flow) biofilms (e.g., those that form in 96-well plates), the increase in biofilm cells in a flowing environment is a net process that is dependent on the irreversible adsorption rate of cells to the surface, the growth rate of the microorganisms, and the removal rate of cells lost to the bulk flow (18). There are numerous benefits for the cells in biofilms, e.g., protection against antimicrobials and the opportunity for and proliferation by continuous cell dispersion. There is also a possible competitive advantage if cells colonize surfaces at multiple sites and grow in such a manner that the resulting three-dimensional architecture exposes the maximum biofilm surface area to surrounding nutrients. The most successful colonizers would therefore be the cells with the ability to adhere to the surface (and stay adhered) and to start multiplying at maximum rate. The process of events from being free-floating cells to the so-called permanently surface-attached phase involves early steps including reversible attachment and a phenotypic change in the cells from a planktonic state to a sessile state, with concomitant changes in gene expression; these steps contribute to a lag phase that will occur before maximal growth/biofilm development can take place (23). Clearly, the ability to progress from the lag phase to a fast-growing phase, as well as the duration of the lag phase, is an important determinant of biofilm function and has an impact in a diverse range of environments, often with implications for infection or contamination control, as well as in industrial processes.At the cell level, an extended lag phase and slower growth create the risk that the cells will be displaced by faster-growing microcolonies, as was demonstrated by Klayman et al. (13) in dual species biofilms. A microorganism''s competence in dominating a surface area can therefore be evaluated by comparing the lag phases and maximal growth rates (μ_max) of a biofilm growth curve. Knowing a bacterial population''s specific growth rate is a requirement for its cultivation at optimum rates in a chemostat or other continuously fed bioreactor. A key assumption for this type of cultivation is that wall growth has a negligible effect, which is in stark contrast to systems where surface-associated growth dominates. Indeed, while dilution rates exceeding an organism''s μ_max results in cell washout in a conventional chemostat setting, biofilm formation enables microbial populations to persist at dilution rates much higher than the organism''s μ_max.Biofilm growth rates have been determined by various techniques, such as fluorescence in situ hybridization (FISH) (12, 31), measuring the incorporation of radioactive substances like [³H]thymidine and ³²P (8, 9), microscopy (3, 13, 17), measuring the total increase in biofilm mass (both cells and extracellular polymeric substances) (24, 28), colorimetric 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenyl-amino)carbonyl]-2H- tetrazolium hydroxide (XTT) assays (26), or measurement of amide II bands, as determined by attenuated total reflectance-Fourier transform infrared spectroscopy (6). Some of the above-mentioned techniques suffer the drawback that the biofilms have to be sacrificed with sampling or that the measured increases do not distinguish between live and dead matter in the biofilm (i.e., increases measured might not represent an accurate increase in viable cell numbers).In this study a carbon dioxide evolution measurement system (CEMS) (15) was used to track the biofilm development rate in real time. The advantage of using this system is that the measured rates represent the metabolic activity of the active cell mass and can be done nondestructively for any biofilm-forming microorganism. In the past, measurement of oxygen uptake rates has been used for determination of growth rates in batch cultures (19) and of the localized growth rate in biofilms (32). CO₂ measurements by a gas chromatograph have been used to determine growth rates in batch systems (4), but to our knowledge this is the first time that CO₂ measurements have been used to determine whole-biofilm specific growth rates. We applied this technique to compare the biofilm μ_max values for two well-described pseudomonads and a mixed microbial community when the organisms are grown on different nutrients and to test the premise that the origin of the inoculum has an impact on early biofilm development.     

Controversies in the laboratory diagnosis of celiac disease in children; new haplotypes discovered

The latest consensus on celiac disease in 2008, under the auspices of the International Societies of Pediatric Gastroenterology, Hepatology and Nutrition, shows that HLA DQ2/DQ8 typing indicates the highest negative predictive value for celiac disease, which would exclude the diagnosis of celiac disease. In Romania, there are no studies on the implication of HLA-DQ2/DQ8 in celiac disease in children. The aim of our study was to analyze the significance of genetic tests, with a focus on negative HLA-DQ2/DQ8 cases, as well as to determine the main haplotypes involved in celiac disease in children. We tested in 37 children with old celiac disease, confirmed based on the presence of intestinal villi changes on duodenal biopsy, the IgA anti-tissue transglutaminase antibodies (TgA-IgA) by ELISA and the IgA anti-endomysium antibodies (EmA-IgA) by indirect immunofluorescence, compared to HLA-DQ2/DQ8 typing by polymerase chain reaction (PCR). In 25 children, the determined HLA haplotypes predominantly belonged to DQ2, and in 3 children we report the presence of a new haplotype, DR3-DQ2/DR4-DQ8, formed by pattern 1, DR3-DQ2-the DQA1*0501 and DQB1*0201 alleles, and pattern 5, DR4-DQ8-the DQA1*0301 and DQB1*0302 alleles. In 9 children, genetic tests were negative for celiac disease. The identification of HLA-DQ2/DQ8 provides additional data in the diagnosis of celiac disease, but a rigid algorithm in the diagnosis of celiac disease has no practical applicability.     

IgG-F-actin antibodies in celiac disease and dermatitis herpetiformis

Anti-actin antibodies are found in 52-85% of patients with autoimmune hepatitis or chronic active hepatitis and in 22% of patients with primary biliary cirrhosis. In patients with celiac disease, anti-actin antibodies correlate with the degree of villous atrophy. Studies on their involvement in celiac disease and dermatitis herpetiformis in Romania have not been done. The purpose of this study was to evaluate of the quality of IgG anti-F-actin antibodies (IgG-AAA) tests compared with IgA tissue transglutaminase antibodies (IgA-TgA) having IgA endomysial antibody (IgA-EmA) as gold standard in celiac disease and dermatitis herpetiformis and to see if there is any relationship between them. The study included 70 pediatric patients with celiac disease under gluten-free diets and 10 adult patients with dermatitis herpetiformis, during 2010. The IgG-AAA antibodies levels were determined by ELISA. Assessing the qualities of IgG-AAA compared to IgA-TgA, we obtained the following values sensitivity (Se) 27.8%, specificity (Sp) 79.4%, respectively Se 88.9%, Sp 79.4% in celiac disease and Se 33.3%, Sp 100%, respectively Se 100%, Sp 100% in dermatitis herpetiformis. Also, there was a prevalence of 24.3% and 30% of IgG-AAA in the two groups of patients, but no statistically significant associations were found. Therefore, we concluded that IgG-AAA can not replace IgA-TgA in children patients with celiac disease under gluten-free diets and in adult patients with dermatitis herpetiformis. AAA-IgG serum activity in both diseases exist, but without a relationship of association with them.     

Protective effects of a plant histaminase in myocardial ischaemia and reperfusion injury in vivo

Grass pea seedling histaminase (a copper-diamine oxidase) was found to exert a significant cardioprotection against post-ischaemic reperfusion damage. Electrocardiogram (ECG) recordings from the rats subjected in vivo to ischaemia and reperfusion showed ventricular tachycardia (VT) and ventricular fibrillations (VF) occurring in 9 out of 12 untreated rats whereas no ventricular arrhythmias were found under histaminase (80U/kg body weight) treatment (n=16 rats). Computer-assisted morphometry of the ischaemic reperfused hearts stained with nitroblue tetrazolium showed the extension of damaged myocardium (area at risk and infarct size) significantly reduced in rats treated with histaminase, in comparison with the non-treated rats, whereas no protection was found with the semicarbazide inactivated histaminase. Biochemical markers of ischaemia-reperfusion myocardial tissue damage: malonyldialdehyde (MDA), tissue calcium concentration, myeloperoxidase (MPO), and apoptosis indicator caspase-3 were significantly elevated in untreated post-ischaemic reperfused rats, but significantly reduced under histaminase protection. In conclusion, plant histaminase appears to protect hearts from ischaemia-reperfusion injury by more than one mechanism, essentially involving histamine oxidation, and possibly as reactive oxygen species scavenger, presenting good perspectives for a novel therapeutic approach in treatment of ischaemic heart pathology.