Genomic analysis reveals the biotechnological ability of Enterococcus italicus to produce glutathione

Through the analysis of the recently available genome shotgun sequence of Enterococcus italicus DSM 15952^T type strain (Accession PRJNA61487, ID 61487), we found the presence of a gene encoding a bifunctional enzyme, termed γ-GCS-GS or GshF, involved in glutathione production and not influenced by feedback inhibition. The gshF gene exhibited high nucleotide and amino acid sequence similarity to other reported sequences from the Enterococcus genus and was constitutively expressed both in osmotic shock or in common cultural conditions. Several experimental studies concerning the culture medium, physiological stress, cell extract obtainment, and scaling-up showed that in selected conditions E. italicus was able to accumulate up to 250 μM of intracellular glutathione, which represented the main thiol group present into the cells. This is the first report regarding the production of glutathione by E. italicus, a species that could be used as a safe adjunct culture for glutathione-enriched dairy foods.     

Environmental light and endogenous antioxidants as the main determinants of non-cancer ocular diseases

The human eye is constantly exposed to sunlight and artificial lighting. Exogenous sources of reactive oxygen species (ROS) such as UV light, visible light, ionizing radiation, chemotherapeutics, and environmental toxins contribute to oxidative damage in ocular tissues. Long-term exposure to these insults places the aging eye at considerable risk for pathological consequences of oxidative stress. Furthermore, in eye tissues, mitochondria are an important endogenous source of ROS. Over time, all ocular structures, from the tear film to the retina, undergo oxidative stress, and therefore, the antioxidant defenses of each tissue assume the role of a safeguard against degenerative ocular pathologies. The ocular surface and cornea protect the other ocular tissues and are significantly exposed to oxidative stress of environmental origin. Overwhelming of antioxidant defenses in these tissues clinically manifests as pathologies including pterygium, corneal dystrophies, and endothelial Fuch's dystrophy. The crystalline lens is highly susceptible to oxidative damage in aging because its cells and their intracellular proteins are not turned over or replaced, thus providing the basis for cataractogenesis. The trabecular meshwork, which is the anterior chamber tissue devoted to aqueous humor drainage, has a particular susceptibility to mitochondrial oxidative injury that affects its endothelium and leads to an intraocular pressure increase that marks the beginning of glaucoma. Photo-oxidative stress can cause acute or chronic retinal damage. The pathogenesis of age-related macular degeneration involves oxidative stress and death of the retinal pigment epithelium followed by death of the overlying photoreceptors. Accordingly, converging evidence indicates that mutagenic mechanisms of environmental and endogenous sources play a fundamental pathogenic role in degenerative eye diseases.     

Molecular diagnosis of cystic echinococcosis in humans from central Poland

The identity of the causative agent of cystic echinococcosis (CE) in humans from central Poland receiving treatment between 2000 and 2010 was determined. A total of 47 samples obtained after hepatectomy were examined and protoscoleces were identified in wet preparations in 27 cases. Using DNA extracted from the samples, two mitochondrial regions (nad1 and cox1 genes) were amplified and the nad1 fragment was sequenced. This PCR analysis confirmed the presence of Echinococcus species in 30 cases and nad1 sequence alignments showed identity with the G7 (pig) strain, Echinococcus canadensis. These data demonstrate that the pig strain of this parasite is the most frequent causative agent of human cystic echinococcosis in central Poland.     

Rab35: GEFs,GAPs and Effectors

Rabs are the largest family of small GTPases and are master regulators of membrane trafficking. Following activation by guanine‐nucleotide exchange factors (GEFs), each Rab binds a specific set of effector proteins that mediate the various downstream functions of that Rab. Then, with the help of GTPase‐activating proteins, the Rab converts GTP to GDP, terminating its function. There are over 60 Rabs in humans and only a subset has been analyzed in any detail. Recently, Rab35 has emerged as a key regulator of cargo recycling at endosomes, with an additional role in regulation of the actin cytoskeleton. Here, we will focus on the regulation of Rab35 activity by the connecdenn/DENND1 family of GEFs and the TBC1D10/EPI64 family of GTPase‐activating proteins. We will describe how analysis of these proteins, as well as a plethora of Rab35 effectors has provided insights into Rab35 function. Finally, we will describe how Rab35 provides a novel link between the Rab and Arf family of GTPases with implications for tumor formation and invasiveness .      

Accumulation of polyhydroxyalkanoates by halophilic archaea isolated from traditional solar salterns of India

Extremely halophilic archaeal isolates obtained from brine and sediment samples of solar salterns of Goa and Tamil Nadu, India were screened for accumulation of polyhydroxyalkanoates (PHA). Seven polymer accumulating haloarchaeal strains (TN4, TN5, TN6, TN7, TN9, TN10 and BBK2) were selected based on their growth and intensity of fluorescence when grown on 20 % NaCl synthetic medium supplemented with 2 % glucose and incorporated with Nile red dye. The polymer was quantified by conversion of PHA to crotonic acid which gave a characteristic absorption maxima at 235 nm. On the basis of phenotypic and genotypic characterization the cultures TN4, TN5, TN6, TN7, TN10 and BBK2 were grouped under genus Haloferax whereas isolate TN9 was grouped under the genus Halogeometricum. Growth kinetics and polymer accumulation studies revealed that the culture Halogeometricum borinquense strain TN9 accumulates PHA maximally at the mid-log phase, i.e. 5th day of growth (approx. 14 wt% PHA of CDW). Analysis of the polymer by IR, ¹H NMR and ¹³C NMR confirmed it to be a homopolymer of 3-hydroxybutyrate.     

Bacterial composition of soils of the Lake Wellman area, Darwin Mountains, Antarctica

The aim of this study was to examine the bacterial composition of high latitude soils from the Darwin–Hatherton glacier region of Antarctica. Four soil pits on each of four glacial drift sheets were sampled for chemical and microbial analyses. The four drifts—Hatherton, Britannia, Danum, and Isca—ranged, respectively, from early Holocene (10 ky) to mid-Quaternary (ca 900 ky). Numbers of culturable bacteria were low, with highest levels detected in soils from the younger Hatherton drift. DNA was extracted and 16S rRNA gene clone libraries prepared from samples below the desert pavement for each of the four drift sheets. Between 31 and 262 clones were analysed from each of the Hatherton, Britannia, and Danum drifts. Bacterial sequences were dominated by members of the phyla Deinococcus-Thermus, Actinobacteria, and Bacteroidetes. Culturable bacteria, including some that clustered with soil clones (e.g., members of the genera Arthrobacter, Adhaeribacter, and Pontibacter), belonged to Actinobacteria and Bacteroidetes. The isolated bacteria are ideal model organisms for genomic and phenotypic investigations of those attributes that allow bacteria to survive and/or grow in Antarctic soils because they have close relatives that are not tolerant of these conditions.     

Optimization of a Method for Chromatin Immunoprecipitation Assays in the Marine Invertebrate Chordate Ciona

Chromatin immunoprecipitation (ChIP) assays allow the efficient characterization of the in vivo occupancy of genomic regions by DNA-binding proteins and thus facilitate the prediction of cis-regulatory sequences in silico and guide their validation in vivo. For these reasons, these assays and their permutations (e.g., ChIP-on-chip and ChIP-sequencing) are currently being extended to several non-mainstream model organisms, as the availability of specific antibodies increases. Here, we describe the development of a polyclonal antibody against the Brachyury protein of the marine invertebrate chordate Ciona intestinalis and provide a detailed ChIP protocol that should be easily adaptable to other marine organisms.