Seasonal variation in nutrient limitation of microbial biofilms colonizing organic and inorganic substrata in streams

Humans have increased the availability of nutrients including nitrogen and phosphorus worldwide; therefore, understanding how microbes process nutrients is critical for environmental conservation. We examined nutrient limitation of biofilms colonizing inorganic (fritted glass) and organic (cellulose sponge) substrata in spring, summer, and autumn in three streams in Michigan, USA. Biofilms were enriched with nitrate (NO₃ ⁻), phosphate (PO₄ ³⁻), ammonium (NH₄ ⁺), NO₃ ⁻ + PO₄ ³⁻, NH₄ ⁺ + PO₄ ³⁻, or none (control). We quantified biofilm structure and function as chlorophyll a (i.e., primary producer biomass) and community respiration on all substrata. In one stream, we characterized bacterial and fungal communities on cellulose in autumn using clone library sequencing and denaturing gradient gel electrophoresis to determine if community structure was linked to nutrient limitation status. Despite oligotrophic conditions, primary producer biomass was infrequently nutrient limited. In contrast, respiration on organic substrata was frequently limited by N + P combinations. We found no difference between biofilm response to NH₄ ⁺ versus NO₃ ⁻ enrichment, although the response to both N-species was positively related to water column PO₄ ³⁻ concentrations and temperature. Molecular analysis for fungal community composition suggested no relationship to nutrient limitation, but the dominant members of the bacterial community on cellulose were different on NO₃ ⁻, PO₄³, and NO₃ ⁻ + PO₄ ³⁻ treatments relative to control, NH₄ ⁺, and NH₄ ⁺ + PO₄ ³⁻ treatments, which matched patterns for biofilm respiration rates from each treatment. Our results show discrete patterns of nutrient limitation dependent upon substratum type and season, and imply changes in bacterial community structure and function may be linked following nutrient enrichment in streams.     

Automated Bayesian model development for frequency detection in biological time series

Background

The ability to perform quantitative studies using isotope tracers and metabolic flux analysis (MFA) is critical for detecting pathway bottlenecks and elucidating network regulation in biological systems, especially those that have been engineered to alter their native metabolic capacities. Mathematically, MFA models are traditionally formulated using separate state variables for reaction fluxes and isotopomer abundances. Analysis of isotope labeling experiments using this set of variables results in a non-convex optimization problem that suffers from both implementation complexity and convergence problems.

Results

This article addresses the mathematical and computational formulation of ¹³C MFA models using a new set of variables referred to as fluxomers. These composite variables combine both fluxes and isotopomer abundances, which results in a simply-posed formulation and an improved error model that is insensitive to isotopomer measurement normalization. A powerful fluxomer iterative algorithm (FIA) is developed and applied to solve the MFA optimization problem. For moderate-sized networks, the algorithm is shown to outperform the commonly used 13CFLUX cumomer-based algorithm and the more recently introduced OpenFLUX software that relies upon an elementary metabolite unit (EMU) network decomposition, both in terms of convergence time and output variability.

Conclusions

Substantial improvements in convergence time and statistical quality of results can be achieved by applying fluxomer variables and the FIA algorithm to compute best-fit solutions to MFA models. We expect that the fluxomer formulation will provide a more suitable basis for future algorithms that analyze very large scale networks and design optimal isotope labeling experiments.     

The reduction of Raf-1 protein by phosphorothioate ODNs and siRNAs targeted to the same two mRNA sequences

Two sets of 20-mer phosphorothioate-modified oligodeoxynucleotide DNAs (sODN) and 21-mer or 22-mer small interfering RNAs (siRNAs), targeted to the same coding sites in raf-1 mRNA, were compared for their abilities to reduce the amount of endogenously expressed Raf-1 protein in T24 cells. The amount of Raf-1 protein was monitored by careful quantitation of Western blots. We found that the siRNAs were somewhat less effective than the S-ODNs in reducing the Raf-1 protein level 20 hours after a 4-hour transfection. The siRNA duplexes were characterized by circular dichroism (CD) spectra, and melting temperatures (Tm) were obtained for the siRNA duplexes and DNA x RNA hybrids formed by the S-ODNs. The S-ODNs differed in their effectiveness, the S-ODN that formed the more stable hybrid being the more effective in reducing the Raf-1 protein level, but the two siRNAs were equally effective despite a difference in Tm of about 20 degrees C. Finally, the siRNAs and S-ODNs had a comparable nonspecific effect on a nontargeted (Bcl-2) protein. Our data add to others in the literature that show it can be difficult to select siRNAs that are more effective than antisense ODNs in downregulating endogenously expressed proteins.     

Canadians’ Perceptions of Food,Diet, and Health – A National Survey

Background

Poor nutrition is harmful to one’s health as it can lead to overweight and obesity and a number of chronic diseases. Understanding consumer perceptions toward diet and nutrition is critical to advancing nutrition-related population health interventions to address such issues. The purpose of this paper was to examine Canadians’ perceived health and diet status, compared to their actual health status, and general concern about their own diet and beliefs about health. Also analyzed were some of the perceived barriers to eating “healthy” foods, with a focus on the availability of “healthy” processed foods.

Methods

Two surveys were administered online to a group of Canadian panelists from all ten provinces during May 2010 to January 2011. Thirty thousand were invited; 6,665 completed the baseline survey and 5,494 completed the second survey. Panelists were selected to be nationally representative of the Canadian adult population by age, sex, province and education level, according to 2006 census data.

Results

Approximately one third of Canadians perceived their health or diet to be very good while very few Canadians perceived their health or diet to be very poor. While the majority of Canadians believed food and nutrition to be very important for improving one’s health, fewer Canadians were concerned about their own diets. The majority of Canadians reported difficulty finding “healthy” processed foods (low in salt and sugar and with sufficient vitamins and minerals). Many also reported difficulty finding healthy foods that are affordable.

Conclusion

Although consumers believe that nutrition is one of the most important factors for maintaining health, there are still a number of attitudinal and perceived environmental barriers to healthy eating.     

Identification of the synaptic vesicle glycoprotein 2 receptor binding site in botulinum neurotoxin A

Botulinum neurotoxins (BoNTs) inhibit neurotransmitter release by hydrolysing SNARE proteins. The most important serotype BoNT/A employs the synaptic vesicle glycoprotein 2 (SV2) isoforms A-C as neuronal receptors. Here, we identified their binding site by blocking SV2 interaction using monoclonal antibodies with characterised epitopes within the cell binding domain (H_C). The site is located on the backside of the conserved ganglioside binding pocket at the interface of the H_CC and H_CN subdomains. The dimension of the binding pocket was characterised in detail by site directed mutagenesis allowing the development of potent inhibitors as well as modifying receptor binding properties.     

Interfacial friction and substrate deformation mediate long-range signal propagation in tissues

Biomechanics and Modeling in Mechanobiology - Tissue layers can generally slide at the interface, accompanied by the dissipation due to friction. Nevertheless, it remains elusive how force could...     

A single amino acid determines the catalytic efficiency of two alkenal double bond reductases produced by the liverwort Plagiochasma appendiculatum

Alkenal double bond reductases (DBRs) catalyze the NADPH-dependent reduction of the α,β-unsaturated double bond of many secondary metabolites. Two alkenal double bond reductase genes PaDBR1 and PaDBR2 were isolated from the liverwort species Plagiochasma appendiculatum. Recombinant PaDBR2 protein had a higher catalytic activity than PaDBR1 with respect to the reduction of the double bond present in hydroxycinnamyl aldehydes. The residue at position 56 appeared to be responsible for this difference in enzyme activity. The functionality of a C56 to Y56 mutation in PaDBR1 was similar to that of PaDBR2. Further site-directed mutagenesis and structural modeling suggested that the phenol ring stacking between this residue and the substrate was an important determinant of catalytic efficiency.     

Current methods for molecular typing of Campylobacter species

Campylobacter remains one of the most common bacterial causes of gastroenteritis worldwide. Tracking sources of this organism is challenging due to the large numbers of human cases, and the prevalence of this organism throughout the environment due to growth in a wide range of animal species. Many molecular subtyping methods have been developed to characterize Campylobacter species, but only a few are commonly used in molecular epidemiology studies. This review examines the applicability of these methods, as well as the role that emerging whole genome sequencing technologies will play in tracking sources of Campylobacter spp. infection.