Use of response surface methodology to study the effect of media composition on aflatoxin production by Aspergillus flavus

Aflatoxins are one of the most important secondary metabolites. These extrolites are produced by a number of Aspergillus fungi. In this study, we demonstrate the effect of media components and enhanced aflatoxin yield shown by A. flavus using response surface methodology in response to different nutrients. Different components of a chemically defined media that influence the aflatoxin production were monitored using Plackett–Burman experimental design and further optimized by Box–Behnken factorial design of response surface methodology in liquid culture. Interactions were studied with five variables, namely sorbitol, fructose, ammonium sulfate, KH₂PO₄, and MgSO₄.7H₂O. Maximum aflatoxin production was envisaged in medium containing 4.94 g/l sorbitol, 5.56 g/l fructose, 0.62 g/l ammonium sulfate, 1.33 g/l KH₂PO₄, and 0.65 g/l MgSO₄·7H₂O using response surface plots and the point prediction tool of the DESIGN EXPERT 8.1.0 (Stat-Ease, USA) software. However, a production of 5.25 μg/ml aflatoxin production was obtained, which was in agreement with the prediction observed in verification experiment. The other component (MgSO₄.7H₂O) was found to be an insignificant variable.     

In vivo evidence for two active nuclease motifs in the double-strand break repair enzyme RexAB of Lactococcus lactis

In bacteria, double-strand DNA break (DSB) repair involves an exonuclease/helicase (exo/hel) and a short regulatory DNA sequence (Chi) that attenuates exonuclease activity and stimulates DNA repair. Despite their key role in cell survival, these DSB repair components show surprisingly little conservation. The best-studied exo/hel, RecBCD of Escherichia coli, is composed of three subunits. In contrast, RexAB of Lactococcus lactis and exo/hel enzymes of other low-guanine-plus-cytosine branch gram-positive bacteria contain two subunits. We report that RexAB functions via a novel mechanism compared to that of the RecBCD model. Two potential nuclease motifs are present in RexAB compared with a single nuclease in RecBCD. Site-specific mutagenesis of the RexA nuclease motif abolished all nuclease activity. In contrast, the RexB nuclease motif mutants displayed strongly reduced nuclease activity but maintained Chi recognition and had a Chi-stimulated hyperrecombination phenotype. The distinct phenotypes resulting from RexA or RexB nuclease inactivation lead us to suggest that each of the identified active nuclease sites in RexAB is involved in the degradation of one DNA strand. In RecBCD, the single RecB nuclease degrades both DNA strands and is presumably positioned by RecD. The presence of two nucleases would suggest that this RecD function is dispensable in RexAB.     

Kolmogorov complexity of epithelial pattern formation: The role of regulatory network configuration

The tissues of multicellular organisms are made of differentiated cells arranged in organized patterns. This organization emerges during development from the coupling of dynamic intra- and intercellular regulatory networks. This work applies the methods of information theory to understand how regulatory network structure both within and between cells relates to the complexity of spatial patterns that emerge as a consequence of network operation. A computational study was performed in which undifferentiated cells were arranged in a two dimensional lattice, with gene expression in each cell regulated by identical intracellular randomly generated Boolean networks. Cell–cell contact signalling between embryonic cells is modeled as coupling among intracellular networks so that gene expression in one cell can influence the expression of genes in adjacent cells. In this system, the initially identical cells differentiate and form patterns of different cell types. The complexity of network structure, temporal dynamics and spatial organization is quantified through the Kolmogorov-based measures of normalized compression distance and set complexity. Results over sets of random networks that operate in the ordered, critical and chaotic domains demonstrate that: (1) ordered and critical networks tend to create the most information-rich patterns; (2) signalling configurations in which cell-to-cell communication is non-directional mostly produce simple patterns irrespective of the internal network domain; and (3) directional signalling configurations, similar to those that function in planar cell polarity, produce the most complex patterns, but only when the intracellular networks function in non-chaotic domains.     

Reproducibility of swallow-induced cortical BOLD positive and negative fMRI activity

Functional MRI (fMRI) studies have demonstrated that a number of brain regions (cingulate, insula, prefrontal, and sensory/motor cortices) display blood oxygen level-dependent (BOLD) positive activity during swallow. Negative BOLD activations and reproducibility of these activations have not been systematically studied. The aim of our study was to investigate the reproducibility of swallow-related cortical positive and negative BOLD activity across different fMRI sessions. We studied 16 healthy volunteers utilizing an fMRI event-related analysis. Individual analysis using a general linear model was used to remove undesirable signal changes correlated with motion, white matter, and cerebrospinal fluid. The group analysis used a mixed-effects multilevel model to identify active cortical regions. The volume and magnitude of a BOLD signal within each cluster was compared between the two study sessions. All subjects showed significant clustered BOLD activity within the known areas of cortical swallowing network across both sessions. The cross-correlation coefficient of percent fMRI signal change and the number of activated voxels across both positive and negative BOLD networks were similar between the two studies (r ≥ 0.87, P < 0.0001). Swallow-associated negative BOLD activity was comparable to the well-defined "default-mode" network, and positive BOLD activity had noticeable overlap with the previously described "task-positive" network. Swallow activates two parallel cortical networks. These include a positive and a negative BOLD network, respectively, correlated and anticorrelated with swallow stimulus. Group cortical activity maps, as well as extent and amplitude of activity induced by volitional swallowing in the cortical swallowing network, are reproducible between study sessions.     

Unconscious orientation processing

Recent findings have shown that certain attributes of visual stimuli, like orientation, are registered in cortical areas when the stimulus is unresolvable or perceptually invisible; however, there is no evidence to show that complex forms of orientation processing (e.g., modulatory effects of orientation on the processing of other features) could occur in the absence of awareness. To address these questions, different psychophysical paradigms were designed in six experiments to probe unconscious orientation processing. First we demonstrated orientation-selective adaptation and color-contingent orientation adaptation for peripheral unresolvable Gabor patches. The next experiments showed the modulatory effects of perceptually indiscriminable orientations on apparent motion processing and attentional mechanisms. Finally we investigated disappearance patterns of unresolvable Gabor stimuli during motion-induced blindness (MIB). Abrupt changes in local unresolvable orientations truncated MIB; however, orientation-based grouping failed to affect the MIB pattern when the orientations were unresolvable. Overall results revealed that unresolvable orientations substantially influence perception at multiple levels.     

Decreased Expression of Transporters Reduces Folate Uptake across Renal Absorptive Surfaces in Experimental Alcoholism

In this study, we examined the mechanistic insights of folate reabsorption during alcoholism, considering enhanced renal excretion as one of the major contributing factors to alcohol-induced folate deficiency. Male Wistar rats were fed 1g/kg body weight/day ethanol (20% solution) orally for 3 months. The results on characterization of the folate transport system in renal basolateral membrane (BLM) suggested it to be a carrier-mediated, acidic pH-dependent and saturable one. Chronic ethanol feeding decreased the uptake mainly by increasing the K _m and decreasing the V _max of the transport process at the BLM surface. At the molecular level, reduced folate transport activity in renal tissue during chronic ethanol ingestion was attributable to decreased expression of reduced folate carrier (RFC) and folate binding protein (FBP). Antibodies against RFC protein revealed a parallel change in RFC expression in both brush border and BLM surfaces during chronic alcoholism. Such findings highlight the role of downregulation of RFC and FBP expression and provide mechanistic insight into the observed reduced folate transport efficiency at renal absorptive surfaces in alcoholism, which may result in low blood folate levels commonly observed in alcoholics.     

Screening and identification of extracellular lipase-producing thermophilic bacteria from a Malaysian hot spring

Seven lipase-producing thermophilic bacteria (ST 1, ST 4, ST 6, ST 7, ST 8, ST 9 and ST 10) were isolated from the Setapak hot spring in Malaysia. The crude extracellular lipases recovered by ultrafiltration of cell-free culture supernatant were reacted in an olive oil mixture and their lipolytic activities were compared. Identification of the bacteria was carried out using the Biolog system and biochemical tests. Strain ST 7 that exhibited the highest lipolytic activity of 4.58 U/ml was identified as belonging to the Bacillus genus. Strain ST 6 with an activity of 3.51 U/ml, was identified as Ralstonia paucula. The lipolytic activities of strains ST 1, ST 4, ST 8, ST 9 and ST 10 were 2.39, 1.84, 2.38, 1.80 and 2.62 U/ml respectively. Strains ST 1, ST 4, and ST 10 were identified as Ralstonia paucula while strains ST 8 and ST 9 were Bacillus spp. Strains ST 7 and ST 9 were tentatively identified as Bacillus thermoglucosidasius, Bacillus stearothermophilus or Bacillus coagulans, whereas strain ST 8 was tentatively identified as Bacillus subtilis.     

Plant–Microbe Symbiosis: What Has Proteomics Taught Us?

Beneficial microbes have a positive impact on the productivity and fitness of the host plant. A better understanding of the biological impacts and underlying mechanisms by which the host derives these benefits will help to address concerns around global food production and security. The recent development of omics‐based technologies has broadened our understanding of the molecular aspects of beneficial plant–microbe symbiosis. Specifically, proteomics has led to the identification and characterization of several novel symbiosis‐specific and symbiosis‐related proteins and post‐translational modifications that play a critical role in mediating symbiotic plant–microbe interactions and have helped assess the underlying molecular aspects of the symbiotic relationship. Integration of proteomic data with other “omics” data can provide valuable information to assess hypotheses regarding the underlying mechanism of symbiosis and help define the factors affecting the outcome of symbiosis. Herein, an update is provided on the current and potential applications of symbiosis‐based “omic” approaches to dissect different aspects of symbiotic plant interactions. The application of proteomics, metaproteomics, and secretomics as enabling approaches for the functional analysis of plant‐associated microbial communities is also discussed.